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·Fibrocystic Disease of the Pancreas

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·Fibrocystic Disease of
the Pancreas
Report of the Eighteenth Ross
PEDIATRIC
RESEARCH
CONFERENCE
Issued by Ross Laboratonas itarterly M&R Laboratories:
Columbus 16, Ohio

·No text on this page.

·The Eighteenth Ross Pediatric Research Confezence, a symposium on Fibrocz cric Disease of the Pancreas, was held under the auspices of the Depart. ment of Pediatrics of the State Unirersity of Lona College of Medicine, at lowa City, in the Confer. ence Room of the lowa Unuersity Hospital School for Seserely Handicapped Children, September 30- October 1, 1955 Cochairmen CHARLES D Nay. MD. Chairman, Department nt Pediatrics Stare Prisersity of lous College of Medicine, low's Cuts Rongat C FRAZILA, MD Assistant Professor, Department of Pediatrics. State Unilersity of loud College of Medicine, loua Cry Speakers ERIKA BRUCK, VID Assistant Clinical Professor of Pediatrics, The Untersity of Bufalo School of Medicine HARIOS CHILDS MID Assistant Professor of Pediatnes, The Johns Hopkins Valtersity School of Medicine, Baltimore R>KRT E COOKE, MD Associate Professor of Pediatrics and Physiology, Yale Unuersty School of Medicine, New Haren PAUL & D SIXI'AGNESE, MID 45sistant Professor of Pedratnes Columbia Unitersity, College of Physicians and Surgeons, New York LA MASIA DISCHE, MD, Associare Professor of Biochemistry Columbia Unnerud), College of Physicians and Surproni, Sex torh TERY AVICHT, MD Associate Professor in Medicine, anderbilt Uniterszy Schonl of Vedicine, Vashtille RY+4915 H. LAYDING. HD Assistant Professor of Pathology and Pediatrics, The Children's Hospital Research Foundation, Concinnon CRARLES U Long, VD 41sacrale Professor of Pediatrics, The Untersity of Bufalo School of Medicine STEPHEW foyce, JR., 1D Instructor in Pediatrics, Unuetsy of Southern California School of Medicine Las Angeles HAARY SHACRMAN, SID. unstant Professor of Pedrainca at the Children's Hospital, Hanard Medical School, Boston T

·The purpose of these conferences is to assist in the correlation of
the latest research information on specific topics of general interest
in the practice of pediatrics, and to stimulate further research by
the exchange of information These objectives coincide with the
intention of the scientific staff of Ross Laboratories (formerly
M&R Laboratories), manufacturers of Similac®, to keep informed
of current developments in pediatric research It has become appar
ent that a large audience is interested in the conferences, and for
this reason Ross Laboratories publish this report
The conference transcript has been edited
and this report prepared by Robert G
Frazier, MD Department of Pediatrics
State University of lowo College of Medicine
Copyright 1956 by Ross Laboratories
Printed in USA
540221 Ta
Quotation permitted if pourte schnowledged

·Contents Embryology as Related to Fibrocystic Disease of the Pancreas 13 Exocrine Function in Fetal Life 18 Fibrocystic Disease of the Pancreas with Cirrhosis of the Liver 26 Sweat - Dasze Physiology 34 Studies of Sweat in Fibrocystic Disease of the Pancreas 45 Diagnosis of Fibrocystsc Disease of the Pancreas & Partial Pancreatic Insufficiency The Chemistry of Mucus 53 59 Histochemistry of Mucus and Exocrine Glands Role of Staphylococcus 69 Metabolic Studies of Fibrocystic Disease of the Pancreas 73 Respiratory Acidosis Basic Physiology 75 Cardiac & Pulmonary Computations in Fibrocystic Disease of the Pancreas 29 Genetics 87 ·CLAUDE A VILLE, JR., PhD. JOSEF WARKANY, M.D. Assistant Professur of Biologicul Chemistry, Harvard Medical School, Boston Professor of Research Pediatrics, University of Cincinnati College of Medicine Participants DOROTHY H. ANDERSEN, M.D Associate Professor of Patholugy. Columbia Untersity, College of Physicians and Surgeons, New York GIULIO J. BARBERO, M.D. Assistant Professor in Pediatrics, Unilersity of Pennsyliana School of Medicine, Philadelphiu HERBERT J. BARTELSTONE, D.D.S Assistant Professor of Pharmacology, Columbia Unitersity, College of Physicians and Surgeons, New York EARL H. BAXTER, M.D Chairman, Department of Pediatrics, The Ohio State Unilersity College of Medicine, Columbus SADIVEU P. BESSMAN, MD. Associate Professor of Pediatrics, The University of Maryland School of Medicine, Balnmore JOHN MCRAIC, MD Associale in Pathology of the Children's Hospital, Harvard Medical School, Boston GORDOY E. Gius, D. Associate Professor of Pediatncs, Unuersity of Nebrasha College of Medicine, Omaha LEE FORREST HILL, M.D. Chairman, Department of Pediamncs, Blank Memonal llospital, Des Moines L. ESMETT HOLT, JR., MD Professor of Pediatrics New York University College of Medicine, New York Douchas E. JOHNSTOVE, M.D Instructor in Pediatrics The University of Rochester, School of Medicine and Dentistry 1 ALHRICHT JOVES, M.D. 4ssistant Professor of Pediatrics, Unitersity of Pennsylıania School of Medicine, Philadelphia CURS P RATSAMPES, M.D Assistant Professor of Pediatnes. The Untersity of Rochester, School of Medicine and Dentistry BENJAMIN KRAMER, ND Professor of Clinical Pediatrics How }ark Unuersity College of Medicine, New York RUSTIN MCINTUSI, VID Carpenter Professor of Pediatrics, Columbia Unuersity, College of Physicians and Surgeons, Aew York PAUL R. PATTERSOY, MD Chairman, Department of Pediatrics, Union Unrersity, Albany Medical College WYNWE SHARPLES, VD Chairman, Board of Trustees, Children's Exourine Research Foundation, Philadelphia SAMLEL SPECTOR, 1.D Associate Professor of Pedroncs, Festern Reserse Untersity School Medicine, Cleteland Wolr W'. ZCELZER, XD Professor of Pediatric Research, r'ayne Unnersity College of Medicine, Detroit

·"But concerning the sneal-bread, we
deelare nothing for a certainty, only
we suspect that Obstructions, if not a
Scyrrhus, may sometimes invade that
part. But this we delegate, to the
enquiry of others.”
Glitian, Fit De Rachitide Swe Morbe Puerif qur
Vulgo The Rickets Dicitur, Storatus London (1658), cited in
Ruhrah. Pediatrics of the Past, New York Paul
Hoeber, 1925

·No text on this page.

·"But concemmy the sweat-bread, we
declare nothing for a certainty, only
we suspect that Obstructions, if not a
Scirrhus, may sometimes invade that
part. But this we delegate, to the
enquiry of others.”
Glisson, F, et al De Rachitide, Sive Morbe Puerili, qui
Vulgo the Rickets Dicitur, Tratatus London (1650), cited in
Ruhrah, J Pediatrics of the Past, New York, Paul B
Hoeber, 1925

·No text on this page.

·Embryology as Related to Fibrocystic Disease of the Pancreas Embryology of the pancreas and the parotid and submaxıllary glands Dr. Josef W'ARKANY: In the fifth week of gestation, the 5 mm embryo shows the beginning of development of the pancreas by two outgrowths from the endodermal lining of the gut. At about the same time, the epithelial anlage of the respiratory system appears as an outgrowth of the pharyngoesophageal border. At six weeks the anlage of the parotid has also become discernible, and somewhat later the submaxillary gland makes its appearance. During the seventh week, the 17 mm embryo shows fusion of the two pancreases into a single organ (figure 1.) The ventral pancreas forms the head, and its duct becomes the main outlet for the organ, the pancreatic duct of Wirsung. The dorsal duct becomes the accessory duct of Santorini. At this time the epithelial lining of the duodenum, the esoph. agus and the rectum grows more vigorously than does the intestinal tube, resulting in the well-known temporary occlusion of these seg. ments as indicated by the black bands (figure 1). Such physiologic occlusion of the lumen has not been found in the pancreatic ducts. The celiac plexus has now become recognizable in its prevertebral position By 10 weeks of gestation histologic differentiation has begun. In the pancreas we see formation of acini, which originate as terminal or lateral buds from the ducts The islands of Langerhans are present at 14 weeks, and in the second half of fetal life there develops a relative preponderance of islands compared to acinar tissue. The islands, like the acini, are products of differentiation from the ducts. In the intestine, Lieberkuhn's crypts are well developed at 16 weeks, and Brunner's glands make their appearance during the sixth month. Meconium has reached the ileocolic junction at this time, however, it is not seen in the rectum before the end of the fifth fetal month Mucid cells are present in the salivary gland at 16 weeks Sweat glands become discernible as epithelial buds, yet they do not develop lumina before the seventh month of fetal life (figure 2). 13 ·The cells of Paneth, which occur in the human fetus not only in the crypts but also on the villi, do not appear before the seventh month. These cells contain about their nucleus large, acidophilic granules, which are discharged into the lumen under the influence of pilocarpine Zymogen granules of the pancreas react to pilocarpine in a similar manner. 17 mm CR LENGTH Lung- Stomach Liver Coeliac Plexus O 4 8 36 38 12 16 20 24 28 32 Fertilization Age in Weeks Figure 1 Development of organs involved in cysuc hbrous of the pancreas, by the seventh week of fetal life Enzyme appearance Trypsin can be found in the duodenum in the fourth month Pancreatic trypsin has been found in a few human fetuses at 16 weeks, but in tno cases it was absent at 22 and 23 weeks. Pancreatic amylase has been found as early as 22 neeks, but it may be absent even at term. Pancreatic tipace is present at 32 weeks Proteolytic enzymes are produced by the pancreatic acını beginning with the fifth fetal month.

·It seems that these enzymatic activities begin at about the midpoint of gestation. It is understandable that accurate data on this point cannot easily be gathered from human ſetuses. However, a study has been made by Sampson of the changing quantity of trypsinogen in the pancreas of the fetal pig. The rapid development of the tryptic action, beginning in the seventh week and reaching almost adult activity in the tenth week, is shown in figure 3. Seven weeks of gestation represents the midpoint of pregnancy in the pig. 120 mm CR LENGTH Sweat Glands Skin Istet Sec Acinus Tunica Muscularis -Meconium -Intestinal Glands Myenteric Plexus 8 12 16 20 24 28 32 Fertilization Age in Weeks 36 38 Figure 2. Development of crocrne glands in the sixteenth week of fetal life The time at which pancreatic trypsin is found in the human fetus is in agreement with the measurement of trypsinogen activity in the fetal pis, indicating a parallelism in enzyme activation. It seems that pancreatic trypsin activity can be demonstrated soon after meconium appears in the intestine ·00100 00090 2.00080 3.00070 -00060 Refractivity due to split products of proteins 00050 3-00040 00 00020 .00010 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 Adult Longth of fortua in millimetres 1 } á s ibi 14 1 Age in Weeks Figure 3 Development of tryptic activity in the pancreas of the letal pig Reprinted from The nurnal of Biological Chemistry, 38 315, 1919 Experimental pancreatic fibrosis The progress made in experimental mammalian teratology dur ing the past 15 years has not contributed further knowledge concerning the pathogenesis of crystic fibroas of the pancreas Congenital anomalies of soft tissues can now lur produced in rats and mice by a variety of teratogenic agents, but the pancreas is not 3mong the organe so influenced deliberate effort to produce prenatal fibrous of the pancreas was made by Lee. Kaplan, Wiseman and myself It has been shown that ahliteration of the pancreatic acinar ticar, acrompanied hy ·fibrous proliferation, can be produced by injection of dl-ethionine, These findings suggested the posibility of producing fibrosis of the pancreas in fetuses of pregnant rats by the injection of ethionine during pregnancy, Several variations of treatment and dosage were tried. Although the ethionine treatment destroyed the mothers' pan. creatic tis-ue, that of their fetuses grew and developed normally. Definite retardation of growth was observed in many litters. It may be that not enough ethionine reached the ſetal pancreas. or that the placenta changed the ethionne into a harmless material. or posibly, that ethionine is not effective because metabolism in the pancreas of the fetus is different from that in the adult animal. This experimental failure is regrettable, since the production of prenatal fibrosis of the pancreas would provide a useful tool for the investi. gation of fetal physiology.

·Exocrine Function in Fefal Life DR CLAUDE A. VILLEE: During the past two decades, research in biochemistry has done a great deal to elucidate the role of enzymes in the economy of the cell. As our ideas of the number and amount of these different enzymes have grown, it has become clear that most, if not all, of the protein material in a cell is actually enzy malic in nature. At the present time it would then he dillicule to conceive of a living cell without these batteries of enzymes. In each cell there are enzymes for the synthesis and breakdown of complex molecules of carbohydrate, protein, and lipid; for the intermediary metabolism of carbohydrates, fatty acids, and amino acids, for the release of energy from substrates and the transfer of this energy to substances such as adenosine triphosphate and other energy-rich compounds Each of these enzymes we now believe is regulated by a single gene Experiments in the field of biochemical genetics have established the so-called "one gene • one enzy me" theory as a sound working ho pothesis. There is experimental evidence to support the concept that the fernalized egg has all or most of these enzymes, and shat each daugh ter cell receives its quota during the process of cell division By mechanisms quite unknown at present, additional enzyme molecules are synthesized within each cell so that the growing mass of cells is continually supplied with an adequate amount of metabolic machinery Enzymes essential for cell survival Studies of the metabolism of human fetal tissue conducted in our laboratory have shown that the fundamental pathways of intermediary metaboli-m are well established at six to eight weeks of life in the human ſetus. This is as early as we have been able to make obmerations Furthermore, these studies hase ahonn that there is little change in the rates of activity along most of these metabolic pathways from then until term The rates of orygen consumption. glucose and pyruvale utilization, and lactale production are all relatutely constant from eight seeks until term in fetal Inver, heart. kidney. Jung and cerebral corter Some changes in enzyme actity were oberied in the course of the e studies. For erample, the enzymatic mechant-me for the synthesis of glycogen and the secretion of glucose are alernt from ·the liver until about the thelfth week of fetal life. Following their appearance they soon reach the adult lesel As deselopment progresses, increase in the ability of the liver to produce glycine, to phosphorylate glycerol so it can be utilized, and to carry on transamination reactions was also obverved We have no comparable observations on the metabolism of exocrine giands, but there is no reason to doubt that, as development proceeds, the rates of oxygen consumption, glucose and pyruvate utilization, lactate production, and other intermediary metabolic reactions are relatively constant in these tissues also. Specific enzymes In addatson to the enzymes that are common to all cells, there are others that are pecubar to specific tissues of the body. Examples of these are the many digestive enzymes produced by the salivary glands, the pancreas and the gastric and intestinal mucosa Secretion of such enzymes is not necessary for the survival of the cell. In view of this, it is not surprising to find enzymes absent when the tissue is first differentiated and appearing later in develop ment. Investigators are not in complete agreement on when these enzymes can first be demonstrated in the human fetus. Needham lamented this fact in 1931, the same situation seems to exist today Salivary amylase, or plyalın, has been identified in the saliva of human fetuses as early as the fourth month of gestation The gastric mucosa of the fetus produces some repain, rennin and hydrochloric acid beginning with the fifth month. Trypsinogen and lipase are secreted in the rancreas by the fifth month But most investigators cannot demonstrate the presence of pancreatic amylase until birth or perhaps even shortly after hirth The enzymes produced by intestinal glands have been said hy come investigators to be present as early as the fifth month, but this has been denied by others. It has been reported that the gastric mucosa of the term infant is about eight times richer in pepsin than that of the premature infant The difference between term and premature infants in tryptic activity of the pancreatic extract is even more marked. It can be inſerred from these results that the amount of pepsin and trypsin produced is much greater in the last month of gestation than earlier Their chief role before birth would appear to be that of digesting and liquefying meconium. lature of the biochemical lesion The hereditary nature of fibrocystic disease of the pancreas umgests that it may belong to that slacs of diseases named inbom ·errors of metabolism by Garrod in 1909. In these, one of the enzymes in a cell is altered, deficient or absent as the result of an action of a mutant gene. The disease would be comparable to alcaptonuria, albinism or glycogen storage disease of the liver. The finding that trypsin is deficient or lacking would suggest at first glance that there is some impairment in the synthesis of this enzyme. However, pancreatic lipase and amylase are concurrently deficient or absent. In all the other examples of inbom error, the change in a single hereditary unit or gene leads to the deficiency of a single enzyme, We might hypothesize that the action of this altered gene leads to some impairment in an enzyme system in the pancreas that is involved in a step common to the synthesis of all three enzymes. This would suggest that the primary biochemical lesion in fibro. cystic disease is the result of a mutant gene which acts to decrease or to eliminate the ability of the pancreas to synthesize all three digestive enzymes Aleconium ileus would then occur because the enzymes are not present in amounts required to liquefy the mecona vum, and impaired digestion and absorption would follow. Sull unexplained by this hypothesis are the viscid mucous congestion in the bronchial tree, and the atrophy of the acınar tissue of the pancreas A second hypothesis postulates that the mutant gene alters one of the enzymes involved in the production of mucoprotein secreted by the mucous glands. The action of the altered enzyme would cause the production of longer and more polymerized molecules than normal, resulting in a more uscid solution This viscid mucus would lead to the plugging of the bronchial tree and of the ducts of the pancreas The plugging of the pancreatic ducts would lead to atrophy of the acinar tissue and deficiency pancreatic digestive enzymes, ohservations that can be made when the pancreatic duct is experimentally tied off It is consistent with this formulation that the deficiency of pancreatic enzyme is frequentls not complete, and includes all of the pancreatic digestive enzyme Other clinical manifestations would follow from the decreased production of the pancreatic enzy me The absence of lipase is probably the most important one, resulting in impaired digestion and absorption of fate, which nould lead to the falls er sometimes aleried, and particularly to the deficiencie of vitamina A and a. and cholesterol. The absorption of theme fat snuble substances would be impaired along with that of lipida This latter theory of the etiology of the deser does not imme diatels suggest any nen metliod of treatment. rien iſ an abnormal molecular configuration of mucoprotein were found, for there is no ·way known at present to delete or alter any of the intracellular enzymes of the body. Discussion DR. DOUGLAS E. JOHNSTOVE: I would like clarification of the evidence that trypsin production in premature infants versus term infants is deficient Dr. VILLEE: Werner from Stockholm made pancreatic extracts and tested their tryptic activity; the differences were quite marked There was no overlapping of the ranges in activity of the extracts from term infants and from prematures. De L. EMMETT HOLT, JR ; May and Vance did assays of duodenal contents and were not able to demonstrate a defect in the enzymes of the premature. Clement A. Smith made observations en tirely in agreement with these, and quite different from those of Werner DR HARRY SHWACHM 4Y: We have studied some prematures weighing 11/2 to 5 lb and found approximately the same activity in the premature as in term infants. This has been verified indirectly by noting a rise in plasma amino acids following a whole protein feeding DR JOHNSTONE. We have found that in a premature infant weighing 2 lb the first meconium plug had as much tryptic activity as that of any term infant we studied. De Villie: I don't think there is necessarily disagreement here The studies have been dealing with two different things, the amount of tryptic activity found in pancreatic extract, and the amount of tryptic activity found in duodenal juice The latter would probably be much more to the point, because it is the trypsin in the duodenal juice that is effective. not in the pancreas DR ROBERT E. COOKE I should think. also, that analysis of the extract of the pancreas simply measures the amount present at any one moment, whereas in the synthetic process, the rate is probably the critical factor. and not the amount in the gland at any one moment. DR DOROTHY H. ANDERSEY: I should like to inquire as to what data is available concerning the earliest age at which the pancreatic lesion of cystic fibrosis has been found at postmortem examination In our experience, the earliest well-authenticated lesion ap peared around the eighth fetal month. We have had a couple of premature infants who died of meconium ileus after having been ·errors of metabolism by Garrod in 1909. In the-e, one of the enzymes in a cell is altered. deficient or absent as the result of an action of a mutant gene. The disease would be comparable in alcaptonuria, albinism or glycogen storage disease of the liver. The finding that trypsin is deficient or lacking would sugges! at first glance that there is some impairment in the synthesis of this enzyme. Honeses, pancreatic lipase and amylase are concurrently deficient or absent. In all the other examples of inbom error, the change in a single hereditary unit or gene leads to the deficiency of a single enzyme We might hypothesize that the action of this altered gene leads 10 some impairment in an enzyme system in the pancreas that is involved in a step common to the synthesis of all three enzymes This would suggest that the primary brochemical lesion in fibro. cystic disease is the result of a mutant gene which acts to decrease or to eliminate the ability of the pancreas to synthesize all three digestive enzymes. Neconium ileus would then occur because the enzymes are not present in amounts required to liquefy the meconium, and impaired digestion and absorption would follow. Still unexplained br this hypothesis are the viscid mucous congestion in the bronchial tree and the atrophy of the acinar tissue of the pancreas. A second hypothesis postulates that the mutant gene alters one of the enzymes involved in the production of mucoprotein secreted by the mucous glands The action of the altered enzyme would cau-e the production of longer and more polymerized molecules than normal resulting in a more viscid solution This viscid mucus hould lead to the plugging of the bronchial tree and of the ducts of the pancreas The plugging of the pancreatic ducts would lead to atrophy of the acinar tissue and deficiency of pancreatic digestive enzymes, observations that can be made when the pancreatic duct is experimentally tied off It is consistent with the formulation that the deficiency of pancreatic enzyme is frequentis not complete. and includes all of the pancreatic digestile enzymes Other clinical manifestahons would follon from the decreased production of the pancreatic enzi me The absence of Jipase is probably the most important one. resulting in impaired digestion and absorption of fats, which would lead to the fatny lover sometimes obversed, and particularly to the deficiencia of vitamins A and a and cholesterol The absorption of these fat soluble substances would be impaired along with that of lipid:. This latter theory of the etiology of the disea-e does not imme diately suggest any new method of treatment, esen of an abnormal molecular configuration of mucoprotein were found, for there is no ·had been in cardiac failure, with marked chest and gastrointestinal symptoms. With apparent clinical improvement, he no longer showed so high a concentration of iodide. These data on parotid secretion of iodide, I believe, contradict the possibility that there is a defect in energy production in secretor, tissue. 20 NORMAL CINC FINOSIS PATIENTS OF 10 NUMBER NO o SJ 10 الد 101-15 13-20 20 MLLIEQUIVALENTS OF WO 4- CHLORIDE PER 50 LITER 511- OF SALIYA Figure 1. Concentration of chloride in the parotid secretion of children. DR. COOKE: I am not sure I would agree with that interpreta. tion There are opposing arguments that can be based on the data on sweat. I do not think the calculation of free energy exchange can be based on the assumption that one anion determines the total work output of a gland We have some calculations for whole sweat; in thee the chloride concentration at times actually runs inversely to the energy requirements for the whole secretion. DR VILLEE: I should like to ask whether the volume of salına secreted by the two groups is the same DR BESSMAY. The volume is a little greater, on the average, in patients with fibrocystic disease than in the control children. DR BENJAMIN H. LAVDING: Can it be considered established that all erocrine glands are affected in this disease; or if not, can we arrive at a consensus about the glands with which we are concerned? This might give us additional information from which to construct hypotheses.

·born in the eighth fetal month, and who had solvulus which must, by internal evidence have dated from about a month before DR CHARLES U. Lowe: With reference to the secretin theory, is it possible that what is lacking is the stimulus to cause an elabo. ration of the three enzyme? This is a middle ground between the two possibilities proposed. Dr VILLEE: I don't see how the secreton theory can explain plugging in the bronchial tree Dr. Gordov E. Gibs: In assays of the intestinal mucosa of patients who died, we have found normal secretin content I was hoping to hear a third hypothesis, namely, a deficiency of some factor in the mechanism of secretion that would also involve the sweat glands. DR. COOKE. That was a hypothesis that I was hoping to present, because I think there is another way that one might explain the concurrent findings in all the glands DR. SAMUEL P. BESSMAN We haye found that salivary secretion of chloride is not at all affected by secretın. Administration of secretın to normal children and to patients with fibrocystic disease of the pancreas did not affect the concentrating power of the salivary gland in either group Perhaps there is a basic lesion of secretory tissue in general that could cause all these abnormalities. We started with the assumption that there might be a defective mechanism for energy produc tion by secretory tissue, that the adenosine triphosphate generation was abnormal in these glands. There is a significant difference in parotid secretion of chloride, an there is in the sweat, between normal controls and patients with cystic fibrosis of the pancreas We have esidence suggesting that there is no energy for the retention of chloride, for keeping it in the blood, and in the patient with this disease it leaks into the parotid and other exocrine secretions Figure 4 shows the concentration of chloride in the parotid secretion of normal children (235) and those with cystic fibrosis of the pancreas (20) There is another son secreted by the parotid gland that is interesting in this respect--it is nodide In the normal individual the concentration of iodide in the parotid salma is between ten- and thirty-fold greater than the blood concentration We gave iodide to a large number of normal children and to four patients with cystic fibrosis of the pancreas, and measured the parotid secretion of sodide following this administration Figure 5 shows that these patients secrete odide în fugher concentration than do the normal indıraduals The triangles in this figure represent the jodide secretion alues obtained in a patient after about three weeks of therapy This patient ·DR. ANDERSEY; I should like to point out that in the glands on which there is general agreement regarding involvement, ie. pancreas, intestinal glands, sueat glands, salivary glands, respira. tory epithelium and biliary epithelium, there are three kinds of abnormalities. 1) morphologic concretions within the glands and ducts, as in pancreas, Inver, and intestional glands; 2) dilatation of the glands and ducts with excess secretion, but without actual concretions, or very rare ones as in the bronchial mucous glands and Brunner's glands In these the glandular secrp. fion appears. histologically, different from what is seen in the pancreas; 3) chemical abnormality of the secretion without associated morphological changes, as in the sweat glands, where under the microscope nothing abnormal is seen, and a chemist is required to discover the abnormality. In view of three different types of change. any theors based on tudy of only one gland is probably inadequate.

·DR. ANDERSEY; Are all agreed there is something wrong with the pancreas? DR. DI Sur'AGTESI: All of us agree that there are some patients in whom the pancreas functions normally. Whether these patients have some involvement of the pancreas that is not recog nizable clinically, because the methods are too gross, is something ne cannot say. Doo HOURS Higures Excretion of iodide in parutid saliva following oral admin Intration of potassium iodide. Large dots are alupe from patients with fibrorystir dreame of the pancreas DR ANDERSEY: Ihat prool is there of the disease pathologically, of the pancreas is normal? DR. Wolf W. ZUELZER That is very much to the point, we have no proof. I think all of us have occasionally seen lesions in the res. piratory tract simulating closels, if not indistinguishable from the pulmonary lesion seen in pancreatic fibrosis, without being able to demonstrate lesions in the pancrea: Pathologicalls, we have no way of knowing whether thes are related to this disease ·Among the living, case 4 has had 110 recurrence of gastrointestinal bleeding since operation, a period of three years, but the pulmonary status has deteriorated. Cases 2 and 6 are doing only moderately well from the pulmonary standpoint, but have not been greatly troubled by liver involvement Case 7 is doing well from all standpoints, although recently hypersplenism has begun to appear (table 1). Table 1 Dala on seien palients uith fibrocystic disease and cirrhosis of lurer Hepatouplond. Liver megaly biopsy Liver func Portal Hyper GI tion system splen. bleedtests pressure 30m ing Shunt ing opera. tion Case Ascites Group I 1 ? 3 + ijo z +++ N Cirrh Cirrh Circh Currh + 370mm NO 450min 570mm + + + + 'ታ * - + + + + to++ + + + Group 11 5 6 N + + + ND** ND. ND ND. ND 0 0 + 0 0 0 0 0 7 ND N 0 * Normal = less than 200 mm of water ** ND. = not done In all patients, hepatosplenomegaly was present. The liver was hard, wontender, and moderately enlarged, and its surface irregular; in several instances, nodules were felt. The spleen was palpated in most cases 3 to 5 cm below the costal margin, firm, nontender, and smooth In case 3, the spleen was enormously enlarged and occupied much of the left side of the abdomen. In the first four cases, biopsy of the liver revealed cirrhosis. Jaundice was absent in all. Although the symptoms of hypersplenism, gastrointestinal hleeding or ascites or a combination of the three, were much more cevere in Group I, all the patients presented evidence of portal hypertension On exploratory laparatomy in cases 1, 3 and 4, the pressure in the portal system was found to be markedly increased A surgical shunting procedure to relieve the increased tension was performed in these cases Pathologic changes The variations in the degree of cirrhosis, with the more severe lesions chiefly in the older children, offer an opportunity for study ·Fibrocystic Disease of the Pancreas with Cirrhosis of the Liver DR PIUI, A pu ST'AGVESE: Review of the medical literature indicates that hepatic changes described in patients with fibrocystic disease have been largely chance findings at autopsy, giving rise in the majority of cases to no clinical symptoms. It can be shonn, however, that in patients with fibrocystic disease of the pancreas the initial lesions may progress to severe and, at times, fatal clinical manifestations, due to a distinctive type of multilobular biliary cirrhosis with concretions. The studies summarized here were carried out in collaboration with Blanc, Clinical matcrial studied Data on 25 patients with fibrocystic disease show that, in the absence of clinical evidence of liver involvement, chemical determinations related to hepatic function are normal Seven patients have been observed with fibrocystic disease of the pancreas and clinically manifest cirrhosis of the Inser They have been divided into two groups Group I, four patients, with biopsy of the liver; and Group II, three patients in whom hepatic biopsy was not performed, but in whom the diagnosis of biliary cirrhosis was felt to be established on clinical grounds. When involvement of the liver was first diagnosed, the patients ranged in age from 1 to 10 years All seven patients had pulmonary disease characterized by generalized obstructive emphysema and chronic bronchopneumonia in varying degrees of severity In cases 1 and 4, this had not been present when the hepatic cirrhosis was diagnosed In all seien cases the concentration of sodium and chloride in the sweat was increa-ed Five of the seven had pancreatic deficiency Duodenal content in case I was extremely uscous, but had a level of tryptic activity in the low normal range. In case 2. tryptic activity of material obtamed on duodenal assay was normal at first, but later decreased to very low levels of the seven patients, three are now dead (ases 1 and 5 died of progrennile pulmonars disease, which in case I had not been present at the time of operation. Case 3 died of uncontrollable gastrointestinal hemorrhage.

·Among the living, case 4 has had 10 recurrence of gastrointestinal bleeding since operation, a period of three years, but the pulmonary status has deteriorated. Cases 2 and 6 are doing only moderately well from the pulmonary standpoint, but have not been greatly troubled by liver involvement, Case 7 is doing well from all standpoints, although recently hypersplenism has begun to appear (table 1). Table 1 Data on seen patients uith fibrocystic disease and cirrhosis oj lner Hepala- Splona tiver megaly biopsy Liver func. tion tests Portal system pressure Hyper-1.GI splen- bloed. ing Shunt. ing opera tron Case Asciten Group I 0 ? . Currh Curele Cirrh Curri N + + + 370mm ND. 450mm 570mm + 0 + 3 + + + + + + Group 11 0 5 6 +++ ND** ND + AD. N + N ND ND ND 0 0 + + ? 0 0 0 0 0 0 - * Normal = less than 200 mm of water **ND. not done In all patients, hepatosplenomegaly was present. The Irver was hard, nontender, and moderately enlarged, and its surface irregular; in several instances, nodules were felt The spleen was palpated in most cases 3 to 5 cm below the costal margin, firm, nontender, and smooth. In case 3, the spleen was enormously enlarged and occupied much of the left side of the abdomen. In the first four cases, biopsy of the liver revealed cirrhosis. Jaundice was absent in all Although the symptoms of hypersplenism, gastrointestinal bleeding or ascites or a combination of the three, were much more serere in Group I, all the patients presented eudence of portal hypertension On exploratory Japaratomy in cases 1, 3 and 4, the pressure in the portal sistem was found to be markedly increased A surgical shunting procedure to relieve the increased tension was performed in these cases. Pathologic changes The variations in the degree of cirrhosis, with the more severe lesions chiefly in the older children, offer an opportunity for study ·of the evolution of the hepatic lesions. The earliest changes seen in our series of patients with cystic fibrosis of the pancreas were noted at three days of age. Focal biliary cirrhosis with concretions On inspection of the capsule at autopsy, the initial lesion is seen as a varying degree of pitting, corresponding to small, stellate, depressed cirrhotic foci. Microscopically, this focal biliary cirrhosis with concretions is characterized by biliary proliferation, inflammatory reaction, concretions of amorphous eosinophilic material plugging the bile ductules, and absence of marked bile stasis. The nature of the concre. tions is not clear, but they show a morphologic and histochemical resemblance to those in the pancreas. Both may be interpreted as resulting from inspissated secretions. The concretions in either organ react negatively to Feulgen's test for nucleic acid, stain simi. larly with periodic acid-Schill reagent, do not show metachromasia. and have a similar pll curse by the methylene blue extinction test. Multilobular biliary curthosis with concretions with time, multiple foci merge Dilſuse portal changes take place. Multiple lobules are encircled and trapped by the fibrotic process. As fibrosis becames more extensive, the architecture is destroyed and the original foc. of concretions may entirely disappear, although, more commonly, occasional foci of eosinophilic concretions are found. Because of the focal character of the initial lesions, damage proceeds irregularly, and areas of preserved lobular pattern are found adjacent to massive foci of bile duct proliferation with fibrosis This multilobular biliary cirrhosis is followed by portal lesions and ends in remodeling of hepatic architecture with regeneratne nodules Typical of this kind of cirrhosis is the formation of irrego ular nodules larger than those in the usual type of portal and biliary cirrhosis. Retraction of fibrous tissue leads at times to deep clefts producing a hepar lohatum Characteristic also is the absence or limited degree of bile stasis At this late stage only. the spleen is enlarged and shows the chronic passive congestion with pulp cord fibrosis typical of portal hypertension. The name of multilobular biliary cirrhosis with concretions has been chosen as descriptre of these adianced hepatic lesions, because a group of luser lobules is involved in the process from the begin. . monolobular biltary cirrhosis, of cholestatic or cholangitic iary, cirrhosis inay also result ·from massive necrosis with collapse of the collagenous framework of the liver. Use of the terms mulalobular, biliary and concretions expresses the distinctive character of the cirrhosis in fibrocystic disease of the pancreas. Development of the clinical picture The first lesion is a focal biliary cirrhosis which gives rise to no clinical manifestations and is a chance finding at postmortem examination. Even when the lesions are extensive, their scattered character is such that, though a significant amount of hepatic paren chyma is affected, there is no clinical or laboratory evidence of involvement of the liver. In some instances the process becomes generalized Sucha patient goes on to a diffuse and severe hepatic cirrhosis, which combines scarring and regenerative nodules with diffuse portal changes. On physical examination, the liver is found to be moderately enlarged, hard, nontender, frequently nodular. An unusual feature is the apparent variation in the size of this organ as it descends further into the abdomen or recedes with each cycle of pulmonary infection, as a consequence of the differences in mechanı. cal pressure exerted through the diaphragm by alternation in the degree of distention of the lungs. Chemical tests of liver function and serum bilirubin are still normal After a varying length of time, a few months or a few years, the distortion and remodeling of the architecture of the parenchyma of the liver is such that portal hypertension eventually develops. It is at this stage that the condition becomes clinically manifest, 1 1 rubin is either normal or very slightly increased. Incidence The autopsy material for the twenty years, 1935-1955, was reviewed in an attempt to establish the incidence of hepatic lesions in this disease. Twenty-five of 116 patients with fibrocystic disease of the pancreas at Babies Hospital had cirrhotic lesions at autopsy, an incidence of 22 percent Sixteen of these had single or multiple lesions of focal biliary cirrhosis in nine, the changes were moore extensive. Cirrhosis of the liver, with portal hypertension, has been observed in 7 of 311 patients with fibrocystic disease of the pancreas, an incidence of a little more than 2 percent. A survey of the diagnostic files from 1919 through 1951 shows that hepatic cirrhosis nas entered among the final diagnose« in 28 ·cases. Seven of the patients had fibrocystic disease of the pancreas; in 10 others the cirrhosis of the liver was of unknown origin; in seven, atresia of the bile ducts was the basic disease; in tso, chronic acquired hemolytic anemia; in one, glycogen storage disease of an unusual type, and in one, schistosomiasis. Portal hypertension was present in ll of the 28 In our experience, therefore, a child with the syndrome of portal hypertension secondary to cirrhosis of the liver has a good chance of having fibrocystic disease of the pancreas. The concentra. tion of electrolytes in the sweat has been found to be characteristically increased above normal only in patients with fibrocystic disease, and the test is therefore of value in the differential diagnosis of cirrhosis. Etiology The histologic appearance suggests that the cycle is initiated by primary mechanical obstruction of bile ductules by what appear to be inspissated secretions, an expression presumably of the abnormality of secretory products of exocrine glands that is character. istic of the disease. While focal biltary fibrosis is a common postmortem finding in patients with fibrocystic disease of the pancreas, the diffuse multi. Tobular biliary cirrhosis with clinical manifestations is rare. fl'hat, then, triggers the passage from one type of hepatic lesion to the other? A varying degree of involvement of the affected area is characteristic of fibrocystic disease of the pancreas. In four of the patients described, the pulmonary disease was milder than average In four, the concentration of electrolytes in the sweat was alnormally elevated but in the lower range of the values character. istic of this condition. In two, pancreatic function was either within normal limits or normal at first, with deficiency appearing later It is possible, therefore, that patients with cystic fibrosis who des elop cirrhosis of the liver provide another example of the vari. ability of this disease It might be postulated that an added infec. tion, eg.. infectious hepatitis or nutritional insult, con adierse response by a luser which is already basically abnormal. It is difficult to incriminate any ingle factor in the elicitation of this response. Nutritional deficiency, ascending cholangitis in a partially obou structed biliary tree or toxicity from long continued administration of antibiotic drugs might all be postulated and may be present or alisent in an indısıdual. Certain it is that diffuse and clinically manifest hepatic cirrhosis in fibrocystic disease of the pancreas was rare some jears ago, but is being recognized with increasing frequency now that survival of palients is greater the condition does cause an ·nol occur, horrever, in young children, and it must be assumed that the main effert of age is longer exposure to a noxious agent and, consequently, increased chance of developing hepatic lesions in a progressive disease. It is too soon to try to prognosticate for patients who develop cirrhosis of the liver with portal hypertension, but current observa. lion would indicate that it is not impairment of hepatic function, but rather increased tension in the portal system and consequent gastrointestinal bleeding that represents their main hacard Surgical shunting procedures are the patient's best hope for survival, and should be undertaken if the severity of the bleeding or, at times, the degree of hypersplenism points to the necessity for surgical intervention. The outlook for patients surviving operation in which portal pressure 1e successfully releed is still not sell-defined. Summary Another major area, the hepatic, must be added to those already known to be frequently solved in this disease. The loralized and focal hepatic lesions, presumably resulting from primary mechanical obstruction, may progress to a pathologically distinct and widen spread multlobular biliary cirrhosts with concretions. Clinically. this advanced cirrhotic process maniſests itself in patients with fibrocystic disease of the pancreas by hepatosplenomegaly, and us characterized by the symptoms of portal hypestension and, usually an absence of icterus, Discussion DR. VERVON KUIGHT. In patients with acute respiratori infec. tion, where the liser comes down in the course of infection, is there an associated period of abnormal function of the her? DR DI SANT'AGSESE As far as I know, chetnical tests of hepatic function shon ng changes under such circumstances Mechanical factors seem to be largely responsible for the apparent enlargement, DA SYNACHVAN. I should like to ask whether dietary or other therapy can be related to the severity of the cirrhos24. We hase enrountered cirrhosia of the liver less frequently, nor do I recall ever seeing a child with portal hypertension, secondary to the hepatie cilange in fibrocystic disease.

·cases Seven of the patients had fibrocystic disease of the pancreas; in 10 others the cirrhosis of the liver was of unknown origin: in seven, atresia of the bile ducts was the basic disease; in two, chronic acquired hemolytic anemia; in one, glycogen storage disease of an unusual type, and in one, schistosomiasis. Portal hypertension was present in 11 of the 28. In our experience, therefore, a child with the syndrome of portal hypertension secondary to cirrhosis of the liver has a good chance of having fibrocystic disease of the pancreas. The concentra. tion of electrolytes in the sweat has been found to be characters istically increased above normal only in patients with fibrocystic disease, and the test is therefore of value in the differential diagnosis of cirrhosis. Etiology The histologic appearance suggests that the cycle is initiated by primary mechanical obstruction of bile ductules by what appear to be inspissated secretions, an expression presumably of the abnor. mality of secretory products of exocrine glands that is character. istic of the disease. Thule focal biliary fibrosis is a common postmortem finding in patients with fibrocystic disease of the pancreas, the diffuse multilobular biliary cirrhosis with clinical manifestations is rare. Whal, then, triggers the passage from one type of hepatıc lesion to the other? A varying degree of involvement of the affected area is char. acteristic of fibrocystic disease of the pancreas. In four of the patients described. the pulmonary disease was milder than average. In four, the concentration of electrolytes in the sneat was ab). normally elevated, but in the lower range of the values characterjatic of this condition in two, panereatic function was either within normal limite or normal at first, with deficiency appearing later. It is possible, therefore, that patients with cystic fibrosis who develop cirrhosis of the Inter provide another example of the var. ability of this disease it might be postulated that an added infec. tion, eg. infectious hepatitis or nutritional insult. could cause an aduerne response by a liser which is already basically abnormal. It is difficult to incriminate any single factor in the elicitation of this response Nutritional deficiency, ascending cholangitis in a partially obstructed billary tree or toxicity from long continued administration of antibiotic drugs might all be postulated and may be present or absent in an individual. Certain it that diffuse and clinically manifest hepatic cirrhosis in fibrocystie disease of the pancreas was rare some years ago, but is being recognized with increasing fre. quency now that suryaal of patients is greater the condition does ·of a child with hepatic disease from other cause; is this pathology specific before the end stage? DR. ANDERSEN; I think that the microscopic V-shaped lesions beneath the capsule are specific. I have never seen anything quite like this in any other disease. Some of the later, severe cirrhotic lesions are complex and I can not always be sure of their origin In our experience, the diagnosis of fibrocyctic disease has been made in several patients, of a type and degree that has required operation for portal hypertension. DR. SHITACHMAS: Do you think bropsy on someone who is not undergoing surgery is justified? DR. DI SANT AGVESE: We are not justified in performing s biopsy if patients have no symptoms of gastrointestinal bleeding. If severe gastrointestinal bleeding is present, I think we are obli. galrd to undertake surgery to relieve portal hypertension and then a biopsy of the Iner should be obtained. DR KNIGHT: If there were assumed a superimposed lesion on the liver that might precipitate arther disturbance and progression of cirrhosis, infectious hepatitis should be considered. The fact that we can use gamma globulin to alleviate the disease prophylactically indicates that children can have it at a very young age in sub. clinical form I wonder if the patient who already has focal lessons of the liver might suffer more damage from infectious hepatitis than the patient without such lesions DR ANDERSEN I would seem quite likely, but rather hard to lest ohjertrety DR DI SANT AGNESE Once portal hypertension starts it seems to develop very rapidly, although the precipitating mechanism is not clear DR CHARLES D Mar The recognition may be abrupt and the sequence of events rapid We have not actually succeeded in isolat ing, from scrutiny of the hostories, events which might be looked upon as precipitating DR BARBERO An analogy, regarding the precipitation of more diffuse liver patholog, may be provided by information we obtained from study of the Chinese unser fluke, Clonorchis sinensis This parasite produces a hillary type of cirrhosis, cholgangitis, and inflammatory involvement Ve found that these children in the Onent can live in a simbolic relation with the disease. As nutrition deteriorated where lach of food occurred, the chuldren developed intense liver disease, berame jaundiced, and showed progressive and massive enlargement of the her. This condition was reversed as mujrbon improved ·DR DI SAVT'AGA ESE. Vo definitive answers can be gaien be cause the number of patients is small. Two of our patients did not have any chnical signs of pancreatic deficiency when the cirrhosis with portal hypertension was manifest. Two others did not have much in the way of pulmonary infection, but had been diagnosed as having fibrocystic disease and kept on restricted diets for many years before the cirrhosis of the liver appeared. Dietary deficiency. therefore, may have an effect in some but not all patients. DR. GIULIO / BARBERO: Two of our sıx patients with biliary cirrhosis first came in without evidence of pancreatic disease and with only minimal evidence of pulmonary disease; they both had portal hypertension and nodular livers. Te have had identical twins who, very early in life, developed nodular livers which have progressively enlarged. They have shown signs of some degree of portal hypertension and hypersplenism, later developing signs of pancreatic deficiency, and now manifesting advanced pulmonary disease. DR. JOHY NI Crud. Of 181 instances of cystic fibrosis of the pancreas coming to autopsy, we have 10 that we have labeled focal biliary cirrhosis In only two of these was there obvious bile stases, and in none was there evidence of portal hypertension. We agree with Zuelzer that these cases of focal biliary cirrhosis are unusual and specific ones. The regenerative nodules that di Sant'. Agnese describes have not been found. The histological pictures of two of the cases that di Sant'Agnese reports here suggest that some other lesion has been superimposed on the liver of a patient with cystic fibrosis of the pancreas In reviewing our pathology records from 1920 until the present. He found the age at death of these patients is progressively increas ing, and thus we can expect a different pathologic picture over the years. increasing age, and most of the periportal fibrosis is accompanied by a hyperplasia of the bile ducts leading to local ibrary exthosis As di Sant'Agnese suggested. this hyperplasia of the bile ducts is probably secondary to the inspissation of bule in the ducts The nutritional status at the time of autopsy cannot be corre. lated with any particular change in the histologi of the loser. We do not subscribe to the published thesis that the nutritional status in cystic fibrosis of the pancreas is parucularly important in the deselopment of cirrhosis DR LOXE ls it possible in the early stages of this lesson 10 differentiate a liser from a child with fibrors clic disease from tha:

·are some my oepithelial cells which are scattered rather irregularly, and are thought to be able to contract, expel fluid, and contribute to the initial secretion, at least hy forcing out material already in the duct. They may contribute to the secretion pressure, said to be as high as 250 mm of mercury. There is a separate blood supply for the secretory portion as contracted with the tubular portion; I do not know the significance of this. Histochemistry Ifistochemical study of the sweat glands has revealed certain metabolic aspects which may be of importance. The secretory por. tion of the gland is faintly basophilic or acidophilic in staining characteristics, and contains many lipid and glycogen vacuoles. The Jack of melachromatic staining is suggestuse esidence that acid mucopolysaccharides are absent The prevence of abundant glycero. phosphatase and of alkaline pho-phatase, and the absence of acid phosphatase has been noted in eccrine sweat glands. It is of interest that these glands contain large amounts of glycogen, both in secre. tory and duct cells. By contrast, apocrine glands contain no gly. cogen. much less alkaline phosphatase, and some iron, The abundant lycogen in the secretory cells at the base of the eccrine sweat glands di appears with actus ity, suggesting that the glycogen may be used for energy for secretion. The role of the alkaline phosphatase has not yet been determined The distribution of sweat glands in various parts of the body is of some importance with regard to the practical measurement of sweat for diagnostic purposes. The iodine starch spot technique bas demonstrated that the number of sweat pores sanes consider. ably from one part of the body to another. The sweat pore count ranges from 350 on the dorsum of the hand down to 150 on the anterior chest It has further been observed that during moderate thermal sweating there is a ciclic discharge by larger and smaller numbers of sweat glands With increased stimulation each sweat gland puts out a larger amount of fluid. Thermal sweating is confined primarily to the trunk, extremities and head. The glands of the palms of the hands the soles of the feet and the axilla are primarily activated by emotion Pharmacology The innervation of the sweat glands has been demonstrated to due through the thorarolumbar outflow of the sympathetic nervous stem Of significance for our purposes is a consideration of the chemical mediator at the end of the postganglionic sympathetic ·Sweat - Basic Physiology Analomy DR. ROBERT E Cooke Sweat glands fall into two general groupa: eccrine glands which are present over most of the body surface, and the apocrine glands which are present primarily in the avilla and perianal region, the latter beginning to function at puberty. Most of this discussion will be devoted to the physiology of the ecerine glands. Grossly, the sneat gland may be divided into two portions, a secretory portion which has been called a glomerulus by analogy lo the kidney, and a tubular portion which has been likened to the tubules of the kidney (figure 6). The analogy of the glomerulus 1 Figure 6. Schematic drawing of the ansiomy of the swral gland Reprinted from Ar chives of Dermatology and Syphilology 57 907, 1918 inexact in that there is no known filtration mechanism in the sweat gland. The secretory portion consists of a coiled duci made up of a single layer of cuboidal cells. The collecting duet, on the other hand, as a double-layered tule, with a very dense, thick, cuticular inner border in the small intercellular canals of the latter there ·Flushing of the sweat gland is of some importance, since after two or three periods of sweating, sweat concentrations of sodium and chloride fall to a stable level. Potassium concentration falls considerably. This phenomenon has been descrihed as a washing out of retained concentrated sweat from the ducts Table 2 Factors decreasing the concentration of sodium and chloride in sueat Local Low skin temperature Acclimatization Flushing of gland Systemic Relative hyperadrenalism DCA, ACTIE Adrenal tumor Hyperthyroidism Increased k intake Na deficit Acclimalization Repeated muscular work Table 3 Factors increasing the concentration of sodium and chloride ah sual Systemic Local Relative hypoadrenalism Region-especially palmar Addison's Disease Vapor barrier Panhypopituitarım Shin temperature flugh NaCl intake Increased rate Elevation of rectal temperature I'rolonged sweating (fatiguel Flevation of deep akın temperature! Arterial occlusion CYSTIC FIBROSIS The systemic factors that cause an elevation of sodium and chloride concentrations in sweat are, in part, the converse of those previously listed table 31, Elevation of rectal temperature and of deep skın temperature may express themselves through their effects on rate of sweating, which I have listed among the local factors laterpretation of sweat tests for diagnostic purposes must be approached with some caution. The concentrations of electrolyte ·fibers to the eccrine sweat glands; this has heen demonstrated to he acetylcholine The sweat gland responds to drugs with muscarinic effect, such as pilocarpine or mecholyl, when injected intradermally. This response is blocked by atropine. By the use of a staining technique, it has recently been shown that specific cholinesterase is present in abundance around the secretory cells of eccrine glands, thereby indicating their cholinergic innervation By contrast, no specific cholinesterase could be demon. strated around the apocrine sweat glands. Even though a small amount of sweat may be produced by local administration of adrenergic drugs, there is no evidence for adren. ergic innervation of the sweat glands. Almost certainly the chemical transmitter is acetylcholine, thus putting the sweat gland in the same category with mucous glands, salivary glands, and the pancreas. We have attempted to measure the adrenergic sweating, but the amount is so small that quantitative measurements are impossible. It is our opinion that epinephrine may stimulate myoepithelial elements leading merely to evacuation of the ducts and not to true sweat formation The evidence for this effect in the apocrine sweat glands is excellent, for eccrine sweat glands, less convincing Factors affecting electrolyte composition With relation to fibrocystic disease, an important aspect of the physiology of sweat glands is a study of the factors influencing the composition of sweat The water, sodrum, potassium, chloride, lactate, bicarbonate and urea content, and the pH of sweat have been studied Lactate in sweat may have bearing on the provision of energy for secretion, but little is known about it Or greatest import to the problem of cystic fibrosis is a consideration of the factors that influence the sodium and chloride concentrations in sweat Factors decreasing the concentration of sodium or chloride in sweat may be divided somewhat arbitrarily between systemic and local, as in table 2. An increase of salt-and-water hormone or exo. genous DOCA-like steroids reduces markedly the concentration of sodium and chloride in the sweat It is likely that the significantly lower concentration of sodium in sweat in severe hyperthyroidism, as contrasted with that of mild hyperthyroidism or euthyroidism, is due to the relative hyperadrenocorticism that accompanies the former The action of potassium excess, sodium deficit and acclimatization may be mediated through an increased output of adrenal steroids In acclimatization, hyperadrenalism is only one of the factors concerned ·microEq/11/min again-t the rate of flow of sneat, the cure shown in figure 8 - obtained. There is a break in the curie, and above a llow rate of 5 m/r/min -odiuin excretion is proportional to sneal flow. The regression equalion for the le. for rate: grealer than 5 gm/11/min, shows the intercept of this line not at zero but at about –185 microEq/Al/min. This quantity was assunied to be the amount of sodium reabsorbed from the original precursor solu. tion per square meter per minute. The slope of that line. 68 6. actui. ally represents the number of microEg of sodium put into each gram of final sweat. The line for potassium intersects zero, indicating no reabsorption. 80 601 Concentration of sodium (Sna) meq per liter + . forearm sites •Right upper lower Left upper lower (Initial periods are encircled) X . 1 25 5 10 15 20 M2 minute Rate of flow of sweat (V) Grams per Figure 7 Relation of concentration of sodium in sweat to rate of flow of meat. Reprinted from The Journal of Clinical Investigation, 35 111, 1956 Conditions of low sodium intale are associated with low sodium concentrations in the sweat With low concentrations of sodium in the sweat, there may actually be less put into the precursor solution per unit of time, and considerably less reabsorbed The argument that low sweat electrolyte concentrations are due to increased reabsorption of electrolyte are not substantiated by these studies Data on various secretions are shown in table 1. The arrows indicate a rise in concentration with rate of sweating ·vary considerably from one region of the body to another. Palmar and axillary sweat base sery high sodium chloride concentrations, especially with intermittent sweating. In a study on the relationship Jetween whole body sheat and sweat collected under a vapor barrier, it was found that the concentration of electrolyte collected locally under a vapor barrier is alu ays higher than in whole body sweat, the differences being less marked at lower electrolyte concentrations Whether these findings are due in part to higher skin temperature and/or increased local rate must he determined In addition, it is possible that some back diffusion of water might eles ate concentrations. Prolonged sweating also leads to a rise in concentration as well as an occasional fall in rate. Exactly the same changes have been observed after arterial occlusion Neither excess water intake nor acute elevation of the plasma sodium and chloride concentrations with intravenous hypertonic salt affect the composition of sweat. Pitressin, which increases water reabsorption in the renal tubule, prohably does not affect water reab. sorption from the sweat duct, since electrolyte and nitrogen concen. trations are not altered Our experiments have corroborated findings to the effect that, with increasing skin temperature, there is a rise in chloride concentration to quite a marked degree, and sodium concentrations rise propornonately. The volume of sweat rises less acutely with increas ing skin temperature than does the concentration of chloride. In sweating induced by cholinergic drugs such as mecholy), there is less effect from changing skin temperature. The marked change in electrolyte concentration in one study on the effect of rate of su eating is shown in figure 7. As the rate of su eating increases, there is an increase in sodium concentration tending to approach a maximum of 56 mEq/1 The concentration of sodium in the salna also rices with increasing rate of flow Process of sucal formation Earlier work suggents that a fairly constant precursor solution of sweat is elaborated and as the secretion passes through the tubule, water and sodium may be reabsorbed Changes in sodum concen tration of sweat have been attributed to changes in the reabsorption of sodum by some portion of the seat gland, Such theoretical considerations are of great importance with regard to the problem of cystic fibroza There has been an atteint to gre some mathematical expresion to this seabeorptne phenomenon and to the total excretor; phenomenon by the secretory cells of the sheat glands. By plotting the rate of excretion of sodium in ·These data suggest that the precursor solution has a high con. centration of sodium when the concentration of sodium in the sweat is high and a low concentration of sodium when the concentration of sodium in the sneat is low, and that, consequently, reabsorption plays a minor role in determining the concentration of sodium in the sweat. Physiology of sweal in fibrocystic disease It was of some interest to us to determine whether the rate of sweating might be increased in patients with cystic fibrosis. Table 5 shows there is little difference between these patients and a control Table 4 Composition of exocrine secretions in normal indiriduals (Usoal Secretion (Ureal Plasma Concentration mEq/ Na CI K 125 Tears Saliva Sweat 1 02 9 1 75 140 2090 20 - 56 1013 10-40 16 20 9 Table 5 I olume of su eat Units mi/M/hr ml/100 cal Patients with Controls cystic fibrosis 152 211 274 Respiratory Water Loss 622 636 gm/kg/hr group The caloric turnover for some of the control subjects who tended to hyperventilate hecause they were not accustomed to aerosol equipment may be high. Respiratory water loss was about the ame in the two groups Three possible explanations for high concentration of sodium in the sweat of patients with cystic fibrosis are greater water reabsorption by the ducts of the sweat glands, less sodium reabsorption or high sodium concentration in the precusor solution.

·It is apparent that area is concentrated in the sweat of nor. mal individuals. Vrea, a very freely diffusable molecule, is assumed to enter the precursor solution by passive diffusion. Then, as water is reabsorbed, the urea concentration rices. With increase in the rate of flow, the ratio of the concentration of urea in steat to that in plasma has been found to remain quite constant at about 1.75 whereas concentrations of sodium and chloride rise with increase in rate of flow of sweat. 14001 Tor values of V> 5gm per M min fomitting initial periods) Ena = 68 6(+40)V-101 (+53) + 1200 1000 x B00F Rate of excretion of sodium (Ena Spa peq per M2 minute 600F 4004 Forearm sites • Right upper lower Left upper lower (Initiai periods are encircled) x 2001 5 20 10 15 Grams pep M2 minute Rate of flow of sweat (V) Figure 8. Relation of rate of exeretion of sodium in sweat and rate of How of sweat Reprinsed from The Journal of Clinical Insestigation, 35:114, 1956.

·These data suggest that the precursor solution has a high con. centration of sodium when the concentration of sodium in the sweat is high and a low concentration of sodium when the concentration of sodium in the sweat is low, and that, consequently, reabsorption plays a minor role in determining the concentration of sodium in the sneal. Physiology of sweat in fibrocystic disease It was of some interest to us to determine whether the rate of sweating might be increased in patients with cystic fibrosis. Table 5 shows there is little difference between these patients and a control Table 4 Composition of erocrine secretions in normal inderrduals (Ureal secretion [Ureo] Plasma K Tears Salısa Sweat Concentration mEq/l Na Ci 110 125 20-90 10 - 13 2056 1040 1.02 ? 16 20 9 1.75 Table 5 Iolume of su eal Units ml/M/hr ml/100 cal Patients with Controls cystic fibrosis 114 152 211 274 Respiratory Water Lose m/kg/hr 622 636 group The caloric turnover for some of the control subjects who tended to hyperventilate because they were not accustomed to aerosol equipment may be high Respiratory water loss was about the same in the two groups. Three possible explanations for high concentration of sodium in the sheat of patients with cystic fibrosis are greater water reabsorption by the ducts of the sweat glands, less sodium reabsorption, or high sodium concentration in the precusor solution, ·l'ater realisorption If urea concentration is a measure of water reabsorption by the sweat gland, the urea concentrations in the sweat should indicate whether there is greater or less water reabsorption by the ducts of patients with cystic fibrosis. The mean concentrations of urea in the sweat of the controls and of patients with cystic fibrosis of the pancreas were found to be essentially identical. The concentrations of urea in the plasma were also the same in the two groups, as were the ratios concentrations of urea in sueat to that in plasma (tahle 6). Table 6 Ratio of urea in su eat to urea in plasma Controls 585 mg/100 ml = 178 328 mg/100 ml 55 81 mg/100 ml = 181 308 mg/100 ml Patients with etic fibrosis Sodium rcabsorption Concerning the possibility that there is diminished sodium real. sorption in the sweat glands of patients with cystic fibrosis, it seems unlikely there would be a specific instance in which realisorption of sodium was diminished, since in other states where sodium concen. tration in the sweat is high, there may actually be greater sodium reabsorption, as shown by the regression equations (figure 8). Furthermore, any system based on reabsorption of sodium cannot be generalized to involve other glands, which is one of the prerequisites for solution of the problem of cystic fibrosja. Sodium concentration in the precursor solution By use of standard thermodynamic equations of the form Pla-ma concentration 1G = nRT In Sweat concentration ja (urte similar to that mm figure 9 is evolved When the sodium concentration rises in the sweat, there is actually an increased gain for the gland; AG is post tive Above & maximum of 50 to 55 mEq Na/l of sweat, AG de. creases. With a continued increase of sodium the sheat, the free energ) gain for the gland decreases until there ja no free energy gain for the gland itself

·In calculating 4G, the total free energy change, for sweat of
varying sodium concentrations, the relationship shown in figure 10
obtains. This is the AG for transport of all the materials comprising
the glandular secretion. As can be seen, the minimum free energy
change, the point at which minimum work is required for secretion,
is in the vicinity of a concentration of sodium in sweat of 90 to 100
mEq/1. Most of the work is due to the transport of water. The
sweat of patients with cystic fibrosis is secreted with considerably
less expenditure of energy than that of the individual whose sweat
has a normal concentration of sodium These same calculations can
be applied to the secretion of all glands.
AG
gcal/
30
20-
توه
ART in
[Na]
10
[Nols,
o
20
60
100
140
Figure 9. Free energy change (AG) for sodium in sweat when secreted
at various concentrations.
If speculation is permissible, it might be interesting to consider
the concept of some common denominator affecting all glands
responding to cholinergic stimuli, that common denominator being a

·geal/ 180 160- AGART In {{ }p [E] 140. 120. 20 GOJNads 100 140 Figure 10. Total free energy change (JG) for the sweat gland at various concentration of sodium on the weat. chortage of energy for secretory activity in the presence of a diminished energy supply, secretions will he formed that require a relatively low free energy change when less energy is available. tuo things will occur, a rise in concentration of solid materials and. If the load 14 great enough and concentration has attained an optimal level, a limitation of rate of secretion Such findings have been obtained when arterial occlusion was carried out What chemical systems would be insolved in such a deficit of energy are unknown, and considerably more nork must be done on the energeties of secretion to confirm or deny this hypothesis, ·Studies of Sweat in Fibrocystic Disease of the Pancreas Clinical findings Dr. PAUL A. DI SANT'AGNESE: la 1952 Kessler and Andersen reported that 5 of 10 patients hospitalized during a sesere hot spell in August, 1918, were patients with fibrocystic disease of the pan. creas They pointed out that such patients responded adversely to periods of high atmospheric temperature. Since there was already some evidence that in fibrocystic disease there may be a disturbance of multiple glandular structures, and since the sweat glands enter into the physiologic picture in a major way only during hot weather, the objectives of original studies were ino, to measure the amount and electrolyte composition of sweat in patients with this condition, and to attempt to relate the findings to clinical casualties and to the pathologic physiology of the disease The group of patients studied has now been increased to 90 with fibrocystic disease and to 151 controls, of which 135 are chil. dren 6 weeks to 16 years, and 16 are adults, 35 to 62 years of age. A description of the technique for collection has been published Seventy-nine of the patierts presented the classical clinical and laboratory features of fibrocystic disease in all these, one or more duodenal assays have shown the absence of pancreatic enzymes Eighteen have died subsequent to the studies. Autopsies of 14 showed the pathologic changes typical of the disease. In the remaining 11 patients, tryptic activity was still present in duodenal contents, hou. ever, the disease was strongly suspected on clinical grounds. Two of these died subsequent to the sweat studies, and autopsy in both has confirmed the diagnosis Specifically excluded from the control group, which included patients with a variety of conditions, were those with known adrenal insulficiency or renal disease and members of the families of patients with known fibrocystic disease The range of concentration of chloride was 60 to 160 mEq/1 in the patients with cystic fibrosis of the pancreas, with a mean of 106, and in the controls 4 to 60 mEq/l, with a mean of 32 (figure 11). The range of values for concentration of sodium in the sweat of the patients with fibrocystic disease was 80 to 190 mEq/1, with a mean of 133, and in the control subjects, 10 to 90 mEq/1, with a ·mean of 59. Age or color did not seem to make any difference in the concentrations of electrolytes in sneat. The mean volume of sweat was not significantly different be. tween the patients with fibrocystic disease and the control group of patients An effect of pancreatic deficiency or of chronic pulmonary dis. ease was excluded by the finding of the same concentration of 15 FIDROCYSTIC DISEASE of the pancreas 90 coses O NO OF CASES 60 CONTROLS 30 151 cases 180 200 MEQ / LITER S Figure 11. Concentration of chloride in the sweat of patients with fibrocytic dreane and of a control group Swrats from midaldomen or scapular region Reprinted from the Journal of the Amerxan Medial Association, 160 816, 1956 electrolytes in sheat in patients with fibrocystic disease with or with out pancreatic insulficiency or respuratory disease in addition, two groups of patients adults with acquired pancreatic deficiency and adults and children with a variety of chronic pulmonary diseases of other origin, were found to have normal concentrations of electro. lytes in the sweat The effects of the administration of pancreatin or antibiotics were also eliminated by similar methods. Data on a few patients with fibrocystic diseaue has been pub. lished showing a normal adrenal response to ACTH stimulation, a normal ability of the kidney to conserie base and a slight decrease in sweat electrolytes on administration of DOC:. These findings constitute evidence that impaired adrenai and renal function cannot ·he significant factors in the production of the abnormal concen. trations of electrolytes. Thirty-one patients of the group with fibrocystic disease had normal concentrations of sodium in the seruin; 10 patients with respiratory acidosis, either uncompensated or compensated, had normal concentrations of sodium in the serum. Six patients with cystic fibrosis of the pancreas, in acute dehydration during hot weather, had los concentrations of sodium and chloride in the serum. The concentrations of potassium were not altered, nor was pH. Heat appe which results abnormal corr. is a so-called pure salt depletion in which primanly the extracellular Nuid compartment is affected. The circulatory changes produced by salt depletion are much more severe than those in so-called pure water depletion, and produce an effect similar to the shock that follows extensive burns, trauma or hemorrhage. These studies suggest that acute loss of salt through the sweat should be suspected when patients with fibrocystic disease, with vomiting and dehydration, are seen during a period of hot weather Thırst is an unreliable symptom Hyperpyrexia, cardiovascular col. lapse, and, at times, coma represent further steps in salt depletion and bring about an acute medical emergency. The patient may be in imminent danger of death unless prompt and vigorous measures are undertaken to restore the depleted water and electrolyte stores. Physiologic saline is the first urgently needed solution; other repair solutions should be administered as indicated. Sweat in other conditions Some of the conditions in which results of the sweat test have been normal are shown in table 7 Sixty members of families of patients with known fibrocystic disease have been studied. Fourteen had abnormally high electrolytes in the sweat and of these, six had unusual susceptibility to long-lasting and severe respiratory Infections in three of these, varying degrees of generalized obstructive emphysema and chronic bronchopneumonia nere demonstrated roentgenographically it was possible to test all members of 15 families in the others, one or more indisiduals were missing. In tho of the 15, both parents were aflected; in three others, one of the parents and, in the remaining 10, electrolytes were normal in both parents Electrolytes in other body fluids The data in table & come from analysis of electrolytes in mixed saliva There is a statistically significant difference in the ·concentration of sodium and chloride between the patients with cystic fibrosis of the pancreas and the control group, which represent a variety of other conditions. This is not true of concentration of potassium Table 7 Some conditions with normal concentrations of electrolytes in the su eat Chronic Pulmonary Disease a. Bronchial asthma b. Familial dysautonomia c. Agammaglobulinemia d. Miscellaneous Idiopathic Celiac Disease Cirrhosis of the Liver Acquired Pancreatic Deficiency Table 8 Electrolytes in mixed salıra (mEq/L) Na CI 24 K 21 27 Cystic Mean Fibrosis of Pancreas SD Controls Mean SD Difference (M) SE. A ML (+62) 16 (+49) 8 197 (+ 6.1) 20 (53) (+107) 16 (106) 11 38 2.9 411 048 SEM Probability (P Number CIT or Casps Controls < 001 10 < 001 16 16 15 15 15 Studies of the parotid secretory rate done with a suction cup and graduated pipet reveal that patients with fibrocystic disease hase a significantly increased parotid secretory rate as compared to the control group (table 9). Iniestigated with negatise results were mixed salivary amylase. lysozyme and electrolytes in parotid salsa The siscosity of mixed and parotid salma was found to be slightly less than that of the control.

·Since most patients with fibrocystic disease have chronic lung disease and continually swallow mucous, it was difficult to know whether the electrolytes in the stomach come from the pulmonary or gastric secretions. The electrolytes in the duodenal contents were investigated and found to be quite normal Excretion of chloride or sodium in the stool, measured during balance studies, was not altered Table 9 Parotid secretory rate (ml/5 min) 0 30 Cystic Mean 6brosis of pancreas (16) SD Control Mean Eroup (23) SD Difference (M) SEAM ΑΛΙ SEAM Probability (P) Number CFP of cases Controls + 0194 0 125 + 010 0.175 0 052 336 < 001 16 - 23 Summary In cystic fibrosis of the pancreas there is a specific electrolyte abnormality of sweat. The concentrations of chloride and sodium in the sweat are markedly increased in patients with fibrocystic disease, as compared with normal individuals or patients with a variety of diseases In contrast, the volume of sweat is not different in the two groups Failure of the sweat glands to conserve salt clarified the unusual susceptibility of patients with fibrocystic disease to heat, through acute loss of salt in the sneat It also offered, from the practical standpoint, a means of treating heat casualties and of preventing their occurrence by the administration of additional salt to the diet, In all cases showing the characteristic anatomical changes of fibrocystic disease at autopsy. sheat tests (when performed) had shown an abnormality in the concentration of electrolytes. In about ·concentration of sodium and chloride between the patients with cystic fibrosis of the pancreas and the control group, which repte sent a variety of other conditions. This is not true of concentration of potassium. Table 7 Some condinons with normal concentrations of electrolytes in the sul eat Chronic Pulmonary Disease a. Bronchial asthma b. Familial dysautonomia c Agammaglobulinemia d. Miscellaneous Idiopathic Celiac Disease Cirrhosis of the Liver Acquired Pancreatic Deficiency Table 8 Electrolytes in mured salu (mEq/L) Na CI 24 к 31 Mean 27 Cyste Fibrosis of Pancreas Controls SD Mean (01) 16 (49) B 197 (+61) 20 (+53) SD (+10.7) 16 (+106) il 38 10 Difference (A.M) S.E.AM. 21 048 SEN Probability (1) Number CFT of Cases Controls < 001 16 001 16 To 15 15 15 Studies of the parotid secretory rate done with a suction cur and graduated pipet reveal that patients with fibrocystic drease have a significantly increased parotid secretory rate as compared to the control group (table 9) Investigated with negative results were mixed salivary amylase. lysozy me and electrolytes in parotid salsa. The viscosity of mixed and parotid saliva was found to be slightly less than that of the control, ·of determinations has not been great between patients and controls, and I wonder whether this could not be considered a serviceable clinical test for smaller institutions Dr. Lowe: D. Sant'Agnese has published some balance data on administration of docu for short periods of time to patients with fibrocystic disease. I would interpret the data as showing some change in the concentrahon of electrolytes in sweat in response to DOCA. I wonder if this might be an abnormal response Table 10 Sweat electrolytes Sodium mEq/ fube Chloride mEg/1 tube gauze gauze 12 102-198 1615 9 9 8 110 6-215 4 103 1-216.9 116 220 1624 156 8 1673 Fibrocystic patients (12) No of determinations Range Mean Control group (40) No of determinations Range Mean 10 11-101 5 31 10 882 258 39 31 9.3-1118 80 930 425 36 319 DR. DI SANT'AG\ESE: There is some response, but the response is not very great, and the concentrations of electrolytes are shll in the Very high range But with each one of these factors, salt restrichon, heat or DOCA, there was some decrease in the concentration of electrolytes in sweat. DR COOKE: I think the response of the sweat gland to DOCA is of some interest. In continual administration of DOCA to a normal individual, there is a striking reduction in the excretion of sodium and chloride via the kidney for two or three days This excretion returns to normal levels, but the electrolyte concentration in the sweat remains at a very low lesel, indicating an entirely different response from that of the kidney tubule DR BESSAY If there is a lesion in the generation of energy, it would be rather difficult to understand how the sodium of the sweat, even if there were some evaporation, is still greater than that of plasma DR. DI SAVT'AGVESE I should like to point out that the determinations in which the concentration of sodium and chloride in the sweat were higher than in the serum remain unexplained We have thought there might be a technical error, due to some cause such as evaporation ·99 percent of living patients with the typical pulmonary and pan. creatic findings, electrolyte abnormalities also existed in the sneat. Patients with adrenal insufficiency have not been tested. The diagnosis has been limited, up to the present, to that fully manifested disease in patients with complete pancrearic deficiency, respiratory disease and abnormal sueat electrolytes. However, gradation in or absence of involvement of the pancreas, lungs, liver and, perhaps, the sneat glands is inherent in this condition. Of course, it is possible that in every instance all susceptible organs are affected, but perhaps sometimes to an extent too limited to give rise lo clinical manifestations. It must he concluded that the generalized disease called fibro. cystic disease of the pancreas may occur in patients in whom pancreatic function is normal, and that the presence of tryptic activity in the duodenal juice in normal concentration does not negate the diagnosis. The mixed salary and the parotid secretion studies indicate that there are three different defects in exocrine secretions that need to be explained a probable abnormality of mucous secretion giving u reasonable explanation for the pancreatic, hepatic and possibly the pulmonary symptoms; an abnormal concentration of electrolytes in eccrine sweat and mixed saliva, and, finally, an increased paroud secretory rate. Therefore, these exocrine glands, different in function and in the products they elaborate, are affected in different ways. It would seem that dysfunction of the autonomic nervous system may be the common denominator responsible for this wide spread disturbance. Discussion Dr. BARRERO. We have attempted to find a simple method for the collection of sweat which will give results comparable to the Darling technique of patch collection, An infant less than six months of age. previously washed, is placed in an Isolette, on a plastic sheet With the Isolette set at 90° F and 50 percent humidity, in a period of 5 to 25 minutes the sweat that has collected on the plane sheet can be aspirated with a syringe. With an older child, a heat cradle and a 100-wait bulb are placed over the trunk. It is obvious that such a gros, technique las a serious potential error of evaporation. Table 10 compares the salues obtained from patients with fibro. cistic disease and a control group in the gauze patch technique of Darling and his shat I have called the tule method. The overlap ·Diagnosis of Fibrocystic Disease of the Pancreas and Partial Pancreatic Insufficiency DR. HARRY SHWACHMAN: In the past the diagncsis of fibrocystic disease of the pancreas has been based largely on tests of pancreatic function. More recently it has become obyious that a few of the children who succumb with this disease have demonstrated none of the changes in pancreatic function during life that we consider characteristic of the disease This, of course, immediately throws doubt on the validity of the laboratory examination of pancreatic function in making the diagnosis Among complicating factors are the adequacy of the duodenal Auid specimen, and the accuracy of the laboratory determination itself. Table 11 lists some of the duodenal fluid measurements They are divided into those which provide a direct measurement of pancreatic function and those which are indirect measures of pancreatic function In addition to the reduced volume of duodenal secretion that occurs in pancreatic insufficiency, there may be increased viscosity and change in pH. If there is complete pancreatic failure, the stimulation of the pancreas by intravenous secretin or by intraduodenal installation of olive oil no longer seems to have effect. The enzyme determinations in the fasting state do not necessarily reflect what reactions the various tests will show following stimulation If there is dummished fasting activity on repeated occasions, it may still be possible to demonstrate a rise in enzyme activity following the stimulation I think this has important implications in attempting to evaluate balance studies on patients with this disease who, in their fasting state. show no activity, yet, after stimulaton with food or with olive oil or secretin, produce some measurable activity. In reporting the enzyme activity or other measurements of duo. denal ſud, it is important to describe exactly what is being measured Our experience has shown that the level of activity of a single enzyme may not be representative of others present in duodenal fluid, ·DR. COOKE: Studies on palmar sweat show that intermittent sweating gives concentrations of electrolyte that are several times the concentrations when the sweating is profuse; these con. centrations may be of the order of twice the isotonic concentration of sodium. Urea concentrations are also tremendously elevated, about 1,000 mg/100 ml. When there is profuse sweating there is a constant flow, with minimal opportunity for reabsorption of water back through the duct or into the skin or wherever else it goes. The urea concentration is useful as a control for evaluation of electrolyte concentrations Schwartz finds the ratio of the concentrations of sweat urea to plasma urea is very close to 1.75, despite extreme variations in levels of urea in plasma. In order to draw the conclusion that the sweat is hypertonic, we would have to be certain there is not a very high urea concentrafron in that same sweat, to ensure that there had been no back diffusion or evaporation. DR. ZACHARIAS DISCHE: Has anyone determined the lactic acid content of the secretions from patients with cystic fibrosis? If there is a defect of the energy transfer in this diseased tissue, we might expect a decrease in lactic acid formation during secretion This may be a very easy way of getting some conclusive indications of the mechanisms involved DR. COOKE. T'e are in the process of determining concentrations of lactic acid. I avoided the lactic acid problem in my discussion because, at the moment, it seems to be in a confused state with regard to concentrations in sweat Investigators who have attempted calculations feel that there is enough lachc acid in a liter of sweat to account for the energy requirement for a dilute sweat. What the actual source of the lactic acid is, whether the plasma or the metabolism of the gland, remains to be determined DR. DI SANT'AGVESE: May I ask Cooke to comment on the solu tions to be used in patients who have acute salt loss? Dr. COOKE. I should think expansion of extracellular volume initially with something resembling extracellular fluid, namely saline, would be the obvious first step Certainly, it is worthwhile to expand volume initially because the hyponatremic patient may have very marked elevation of the serum potassium, even though he is short of potassium, and he is particularly susceptible to potassium intoxication. Replacement with a potassium containing solution is the next step. If there are symptoms or signs of water intoxication, which 1 do not believe your patients exhibited, a more rapid elevation of sodium concentration with the use of three percent sodium chloride Latime ernension ·abnormal values obtained by the usual measures of pancreatic func. tion Table 12 shows the ranges that we have found in the various measurements. It would seem that even the roughest test for pan. creatic function would, perhaps, be useful, even though it did not necessarily indicate intermediate ranges. Table 12 Duudenal fluid findings Test Healthy Complete Celiac Partial paninfants and pancreatic croatic in disease insufchildren sufficiency ficiency Volume, ml/hr 8-25 20.15 1.10 5-25 pl 60 8.4 60-84 35-8.1 5084 Bilc 20-125 15 75 10-125 10 125 over 3 in Viscosity over 3 in approxi. below 3 below 3 87 percent mately lime, min patients 70 percent patients Amylase, units 1 20 420 01 020 under 6 mos 0 0 0 0 Lipase, unsts 3-10 310 0-1 010 Protease, units 18-10 18 70 0-25 0-70 Carboxy peptidase, mg 0 10 100 10.100 0-0 40 000-100 A-naphthol Chymotrypsin 11 65 11 65 0-5 0 65 mg B-naphthol *Based on examination of appro nimately 250 patents The other talues are based on a study of orez 1,009 panents The numbers refer to tange of achlity of the enzymes as defined and determined in our laboratory The viscosity test can be carried out in about 90 percent of the patients examined It cannot be done on all because of the technical difficulties We do not centrifuge or filter the specimen hut measure it, using an Ostwald viscometer immediately after collection In most patients with compete pancreatic insufficiency, the viscosity time is well over three minutes Since amylase is absent under six months of age in all children, there is no need for the determination of the amylase in the diag. nosis of disease in small infants The method used for this deter. mination is the standard procedure described by Free and Meyers.

·Indirect measurements, such as nitrogen and fat balances are useful. These are probably not of great importance in diagnostic work, but are of fundamental importance in an understanding of the disease. All the diagnostic methods that have been devised are not listed here. For example, the antitrypsin titrations reported by the group in England we found of no great help. Measurement of excretion of jodine in the urine after administration of Lipiodol ® by mouth has not proven reliable as a diagnostic procedure. Table 1 Laboratory tests for pancreatic function Direct Indirect Duodenal Fluid Metabolic balance Volume Before and after Nitrogen and fat intake & stimulation output Bicarbonate Before and after Stool examination or pil stimulation (1) Gross examination (see Viscosity Metabolic balance) Enzyme activity (amylase, lipase, Excess bulk with increase proteases) in total fat and nitrogen (1) Fasting state (2) Microscopic examination (2) After stimulation Fat (a) intraduodenal olive oil Starch (b) intravenous secretin Proteins (Schmidt test) (3) Enzymes-trypsin Absorption tests (1) Fat (a) cream chylomicron counts fatty acids (b) vitamin A esters vs alcohol and aldehyde serum levels of Vitamin A (2) Proteins (a) gelatin or casein absorption tests (b) 7181 labelled casein link al response (1) To dietary adjustment (2) To an adequate potent pancreatin preparation Clinical response appears at the end of this table because this may give a clue as to whether pancreatic insufficiency exists or not This is certainly not a very scientific way of demonstrating any. thing, but I think that when there is doubt we might get some help by this technique There has been considerable discussion about the tremendous range of activity between the average normal and the definitely ·with normal sweat electrolytes. Have the latter children been repeatedly tested to be sure these values were correct? DR. SHFACHMAN: We examined 90 parents, and found about 21 to have abnormal concentrations of electrolytes in the sweat. We found two pairs of parents where both father and mother had abnor. malities. Repeat sweat tests have been performed in the atypical cases. DR DI SANT'AGNESE. In our studies, 100 percent of patients who have died and have shown pancreatic change at autopsy have had an abnormal sweat. At Babies Hospital we have not seen patients with cystic fibrosis who did not have abnormal sweat; however, I feel sure that a patient with cystic fibrosis may have normal sweat electrolytes DR ZUELZER: How much variation within the same individual is observed over a period of time? Is there any progression of the sheat anomaly comparable to that in the enzyme anomalies in the duodenal juice? DR DI SANT'ACNESE. While there was some variation in repeated testing of one individual a few weeks or a few months apart, there has been no evidence so far that time or increase in age makes a difference in electrolyte concentration DR. ROBERT G FRAZIĻR In trying to set up some of our diag. nostic methods for the assay of enzymes in pancreatic juice, we studied the gelatin viscosity determination as described by Leubner and Shwachian We found that the accuracy of the method was markedly decreased by small vatanions in the ratio of initial 10 final viscosity When the proteolytic activity was carefully determined by both the gelatın viscosity method and the Andersen-Early method simul taneously, the results were comparable It is my feeling that the Andersen-Early modification of the Ferm method is simple to per. form, reliable in its ability to mea-ure proteolytic achivity, and requires no special equipment The viscosimetric method has the potential for greater accuracy, but this is difficult to achieve DR 5 ALBRIGHT JOVES I would like to report some daia obtained from duodenal intubations Material aspirated from the duodenum during a routine intubation mas consist of a mixture of duodenal and gastric juices A study was made to determine whether it is important to exclude gastric contents from the duodenum during the collection of duodenal juice for enzymatic analysis Sixty-three patients, none of whom had cystic fibrosis of the pancreas, were selected bilumen Aller-Abboit tube, with the openings so located that gastric and duodenal juices could be separately aspirated, was used A sample of Juodenal juice was collecteit by routine intubation Then the stomach was emptied as completely ·! The method for lipase utilizes olive oil. The protease method employs casein as a substrate, and the liberated tyrosine is measured. Carboxypeptidase and chymotrypsin are two enzymes of interest because they have not been measured regularly in patients with pancreatic insufficiency. Determination of the former is not now one of our routine measurements, but we measured it in a series of patients and found that in complete pancreatic insufficiency this enzyme is deficient, just as is protease. Chymotrypain is of interest because it will attack certain proteins, including casein, much more effectively than trypsin. Chymotrypsin attacks gelatin less eſficiently than trypsin The standard procedure used routinely in our laboratory for determination of proteolytic activity is the Andersen-Early modifi. cation of the Fermi method. We also use a method based on the change in viscosity of a gelatin substrate in the presence of duodenal fluid. In this method the amount of enzyme present can be calcu. lated from the change in viscosity in the one-hour incubation period. We feel that this test is a more sensitive measure of proteolytic activity than is the Andersen-Early test, in which the end point depends on liquefaction of well over 80 percent of the substrate. We studied a fairly large number of children with all types of conditions and have not encountered any condition in which the concentration of electrolytes in the sweat is as high as it is in this disease. We found three or four patients with fibrocystic disease in whom the concentration of electrolytes in the streat was in the normal range. 1 should like to emphasize the great importance of taking very careful and accurate histories and, alo, of having the faculty of doubting certain statements that are made by parents who are well. intentioned but who, perhaps do not have the experience necessary to report accuratelt on obsersations concerning the development of their infant, Discussion DR STEPHEY W. ROYCE I should like to knon whether Shnachman has any figures on abnormalities in the sweat of the parents of his child patients I think such data might be used to set up & screening test for parents Di Sant'Agnese cited abnormalities of sweat electrolytes in 100 percent of palients, while Shwachman stated he found three or four ·The Chemistry of Mucus DR ZACHARIAS DISCHE Clues to the understanding of the chemistry of mucus came from a know ledge of the chemistry of related compounds. These are primarily the so-called blood group substances, which were discovered at the beginning of the twentieth century and were characterized initially on the basis of their serologic behavior. It was soon found that the gastric juice, mucosa of the stomach and other mucoid substances from the intestines displayed some features, including the specificity, of the serologic behavior of blood group substances Urine and other substrates, such as saliva, were also found to contain these compounds In 1946 it was found that the blood group substances obtained from the mucosa of the stomach of the pig contained a sugar that up to this time had not been found in animal tissue. This was so-called fucose, a methylpentose or 6-desoxy-hexose. About one or two years later we succeeded in developing a sensitive and simple method of detecting and determining this type of sugar. Mucus of the respiratory tract and all kinds of mucus of the intestinal tract and of the urogenital tract were found to contain compounds similar in composition to blood group substances consisting of galactose, hexosamine, later found to be a mixture of two different kinds of hexosamine, and fucose Quantitative determination indicated that the amount of fucose is very similar in all these compounds in human mucus, the amount of fucose varies between 13 and 18 percent of the total sugar The ratio between the galactose and fucose moieties is between two and three to one; for hexosamine, and fucose These are combined in such a way that the aldehyde group of carbon 1 of one sugar is attached to one of the hydroxyl groups of another sugar, forming long chains of repeating units There is convincing evidence that fucose is attached to the hydroxyl of another sugar so that its hydroxyl groups are all free Mucopolysaccharides of this type are also associated with serum proteins, particularly globulins, and are present in almost all animal tissues Without attempt at strict definition, this combination of 59 ·as possible, and kept emptied by frequent aspiration during the collection of a second duodenal specimen Determination of pH and enzymatic analyses for trypsin, amylase and lipase were made on the two samples; comparison showed no important or consulent differences between the two specimens. It is concluded that the customary method of collecting one duodenal specimen for enzy. matıc analysis is satisfactory. Dr. May: At what age does the anomaly of secretion of sweat appear? Have measurements been made at birth? DR. SHWACHMAY: We have been able to measure sweat in infants between four and five weeks of age, and have found the defect present. We have not been able to get enough sweat from the smaller infant. Dr DI SANT'AGNESE: We have recently examined a number of children about six weeks of age with the same results. DR BARBERO: We have studied the infants with meconium ileus shortly after the convalescent period, at 7 to 10 days of age, and these have shown abnormal sweat electrolyte concentrations. However, the production of sweat in these infants appeared to be the exception to the rule, since we have had no success in collecting sweat generally before two to three months of age DR. SHWACHMAN: It has been shown that meconium from patients with pancreatic fibrosis is abnormal in its chemical composition Our observations have confirmed the presence of protein, which is abnormal Normal meconium will not precipitate when protein precipitating agents are added, the abnormal meconium in these patients will This can be of practical value at the time of operation to determine whether the condition is atresia or meconium ileus, a 20 percent trichloroacetic acid solution can be utilized as a precipitating agent to test the meconium.

·A similar analysis of mucus from patients with cystic fibrosis is shown in table 11. In the ratios of ſucose to galactose and of hexosamine to galactose, there is no fundamental difference as com. pared with the normal fluid ercept, perhaps, in one instance where the fucoce 19 low. Table 13 Amount and composition of the mucord frachon of the duodenal fluid of normal indruiduals All values in mg/100 ml of duodenal fluid Specimen Protein Glucose Galactose Fucose Herosamine Mucopoly saccharido % of mucus 1 19.0 470 395 51.5 IT 218 70 310 190 030 217 11.5 210 111 IV 19 8 28 8 67 18 5 1 265 20 21.2 107 داد VI 130 13,5 13 S 177 Table 14 1mount und composition of the mucoid fraction of the duo. denal fluid from patients uuh cystic fibrosis of the pancreas All values in mg/100 ml of duodenal fluid Mucopoly Hero saccharide Specimen Protein Glucose Galactose Fucose % of mucus 156 #3 66 JI 291 90 300 32.0 377 0.28 111 662 125 215 45 1V 130 11: 150 228 25 samino - 100 The ralios between ſucose and galactose and between hexosamine and galactose in both groups, with one erreption, shon no significant difference This one very simple measure so impreselle as a differentiating criterion with blood group substances, showed ·mucopolysaccharide and protein has been called a mucoprotein or, if the content of the total unit in carbohydrate is low, glycoprotein Mucoproteins are not so much characterized by the amount of carbohydrate but by the fact they do not have the properties we expect of the more familiar proteins. For example, they are very often not precipitable by hydrochloric acid, nor by trichloroacetic acid, which is one of the most powerful and complete precipitants The mucoproteins can be classified by the nature of the mucopoly. saccharide. Apparently, fucose is able to condition the surface of the poly. saccharide and, in a combination of polysaccharide with the protein, to condition the surface of the protein. This is essentially the same role played by the residues of amino acids in the protein molecule. The fact that the content of fucose varies to a great degree within various types of mucus in a specific species, between individuals of the same species and between species bears out this supposition. Analyses of mucus in duodenal fluid In our studies, we found the content of fucose in mucoids from serum may vary, in the different mucoproteins and in different indu viduals, particularly in disease, up to 200 or 300 percent. It is a mobile component that may be of major importance. It seemed possible that the change in viscosity of the duodenal flud in patients with cystic fibrosis could be due to a difference in the degree of polymerization of the mucopolysaccharides, which might be associated with differences in the ratios of various sugars This might offer the opportunity to differentiate it from the duodenal fluid of normal children The result of the analysis of the intestinal fluid of normal indaviduals is shown in table 13. The mucoprotein fraction of the fluid contains fucose, hexosamme, galactose and one or two other hexoses calculated as glucose There is almost no relation between the amount of "glucose and the amount of galactose The "glucose" in the duodenal fluid comes from other sources than the muco. proteins. There is an equivalence between fucose and galactose, except in one instance where the difference is beyond the limit of error of the method. The lack of a stoichiometric relation of herosamine and galactose can probably be explained by the presence of another type of polysaccharide found in all forms of mucin This is şıalic acid, which is a hydroxy-keto acid of known structure. It is apparently combined with hexosamine, and this excess of hexosamine over galactose may be due to the inclusion of sialic acid.

·Histochemistry of Mucus and Exocrine Glands Dr. BEVJAVIV H. LANDIC: Some but perhaps not all mucus. secreting glands of endodermal origin are affecled in cystic fibrosis of the pancreas, and all suscepuble glands may not be affected in all patients. Whether mucus glands arising from other germ layers can be affected depends on the outcome of as yet unresolved disputes about the embryologic and functional positions of the salivary glands and on studies on the genitourinary tract glands. I do not think we can say whether the pattern of involvement is conditioned by the fact that those glands most clearly affected are structures with ducts which would be more easily obstructd mechanically We decided to confine our studies to airway, pancreas, biliary tree, stomach, small intestine, and colon, where the specific defect involves mucus, utilizing selected blocks of these tissues, from a group of patients of varying age with fibrocystic disease and a group of control patients All the tissues were fixed in cold acetone and embedded in paraffin, for the reason that this is a method which allows study of some enzymes as well as the mucin. We used 40 blocks in all, and carried sections of each block through a total of 10 procedures. We chose to devote most attention to mucopoly. saccharides, chiefly because histochemical methods for the mucopolysaccharides are much better developed than are those for proteins. The classical histochemical procedure for the demonstration of the carbohydrate moieties of mucoproteins is the periodic acid leukofuchsın, or Pis procedure Periodic acid oxidizes the carbo. hydrates to carbonyl groups which can bind leukofuchsin as a red dye for demonstrating acidic groups, such as the groups of carboxylic acids like mucic and glucuronic acids, or ester sulfate, He used a basic thrazıne dje, toluidine blue (TB). Aldehyde-fuchsın (AF) stains both acid groups and mucin generally The mechanism of the more general inucin-staining power is not known, but there is some evidence the tripheny Imethane dyes an a class have an affinity for mucins Results of tissue stains 4 shown in table 15, there are no significant differences which me tan recognize belween the staining powers of mucus from normal 63 ·no differences between the mucopolysaccharides from normal indı. viduals and from patients with cystic fibrosis. Discussion DR. GIBBS: We have determined the serum B-glucuronidase in normal infants and children (25) and in patients with fibrocystic disease of the pancreas (15). There is an elevation of borderline statistical significance in the serum levels of children with fibro. cystic disease; this is probably explained by the pulmonary infection We have also determined the serum mucoproteins in the two groups by the method of Winzler. There was no difference found by this method. DR. BESSMAN We have made a somewhat similar investigation of the salivary mucoprotein We measured hexosamine, acetylhexoscamine and sialıc acid in several patients with cystic fibrosis and in a number of normal children There was no distinguishing feature between the two groups. DR BARBERO We have investigated the protein fraction of duo denal fluid on controls and in patients with fibrocystic disease After dialysis and concentration of duodenal Nuid, the proteins have been separated by paper electrophoresis At a pH of 85 ve have obtained a band moving in the opposite direction from that in controls This could be a different protein or a difference in polymerization. We have not tested salma.

·patients and patients with cystic fibrosie, with regard to pas, tolui. dine blue or aldehi de fuch-in staining. The last three columns shou the effect of a variety of oxidizing agents, Sodium bismuthate and lead tetra-acetate are two reagents which produce approximately the same results as periodic acid. that will con. hich can be th this disease may show a greater tendency to take this stain, but this is the only finding suggesting difference. This may indicate some link in the mucus of patients with cystic fibrosis which is more easily ruptured by sulfuric acid-permanganate. Table 17 Tolurdine blue binding al cartons plt 3 4 5 6 7 ! If it Controls Pancreatic ducts Stomach Snjall intestin Brunner's glands Colon Laryny Bule ducts ftit 1 it + 1 It It + 17 ++ + to to Pancreatic fibrosis Pancreatic ducts Stomach Small intention Brunner's glands Colon it! Hit It Ill + to to + tn + + to it + to 1+ to + + + to + ++ to +++ 10 + +to It +10 Bronchioles Bile sunt Galliolashda #to #to +10 - l'able fuse the findings of the preliminary attempts to study the lobbits of these mucuses, simply by extracting them with water Ir 21 hour sl &ring pH There are no obvious differences. Table 17 how the results of staining with toluidine blue in 12072083 al 211005 pH values, more strongly acidic groups hind ·Table 15 Basic procedures and effects of oxidants PAS TB AF Bis KMnO. AF PbS + to + Controls Pancreatic ducts Stomach Small intestine Brunner's glands Colon Laryny Bile ducts + ++ ++ ++ ++ + ++ It It + 1 it! ++ ++ +++ II 1 + + + ++ +1 + to ++ ++ ++ Pancreatic fibrosis Pancreatic ducts Stomach Small intestine Brunner's glands Colon Larynx Bronchioles Bile ducts Gallbladder I H+ I It! I ++ | Ito ++ + to — ++ ++ ++ ++ ++ +++ + to - ++ to +++ ++ ++++ to + + ++ + to + + to 1 - + to + + to it It It + Table 16 Extrachon at different pll-Pas slaining 25 4.5 65 8 5 + ++ + ++ ++ ++ ++ +++ + + + + Controls Pancreatic ducts Stomach Small intestine Brunner's glands Colon Laryny Bile ducts l'ancreatue obro- Pancreatic ducts Stomach Small intestine Brunner's glands Colon Larynt Bronchioles Bile ducts Gallbladder ++ ++ ++ + ++ t. + ++.+ ++ ++ ++ ++ +++ ++ + ++ ++ + +,+ + +,+ ·toluidine blue at lower pll Again, there is a rather questionable increase in toluidine blue binding generally in the mucus of the patients with cystic fibrosis It is more apparent at higher pH, sug. gesting that these mucins have a higher content of a weakly acidıc group, possibly carboxyl, as opposed to strongly acid sulfate ester, I do not know of any phosphate ester groups normally occurring in the mucins, except for glycogen. Table 18 shows results from tests with pepsin, trypsin, hyal. uronidace, and streptokinase plus streptodornase (skn). There may Le slightly greater resistance to digestion with trypsin of the mucins of patients with fibrocystic disease. Table 19 Miscellaneous determination Top SII SS Acetylation-PAS + TH!!!! + it + + + + + + I t It It It Bt it! Controls Pancreatic celle Pancreatic ducts Stomach Small intestine Brunner's glands Colon Laryny Bile ducts Pancreatic fibrosis Pancreatic ducts Stomach Small intestine Brunner's glands Colon Larynx Bronchioles Bile ducts Gallbladder + +, + - + Table 19 completes the miscellaneous analyses which we did. Lipa e is present in surviving acınar cells in patients with fibrocystic disease in apparently normal amounts. The sulfhydral and disulfide contents shows no differences, which may indicate the presence of normal quantities of chymotrypsinogen. Acetylation is a procedure that will block the oxıdızıng reactions of the muruses. The acetylating ability of both groups is about the same ·Table 18 Digestibility of mucins in vitro Pepsin 3 hrs. Pepsin 20 hrs. Trypsin 3 hrs Trypsin 20 hrs #to 1 十一十十十十一十 2 + + + + D p Esloot D D Controls Pancreatic ducts Stomach Small intestine Brinner's glands Colon Laryn Bile ducts Tancreatic fibros Pancreatic surts Stomach Small intestine Brunner's gland- Colon Larynx Bronchioles Bile ducts Gallbladder + ta + ++ ++ In + + + to T! 31 oa Hit soit + 13 +10 - Hyal 4 firs SKD 4 hrs SKD 22 hrs, + ++ ++ ++ ++ ++ ++ It Controls Pancreatic dieta Stomach Small intestine Brunner's glands (plon Larynx Bile durts Pancreatic fibros Pancreatic duris Stomar Small intatie Brunner's plands Colon larynt Bronchioles Bile ducts Gallbladder +++++++ 10 + ++ ++ ++to+ ++ + to ++ tv ++ + ++ * means sections totally diokset ·Role of Staphylococcus DR. VERXOX KNIGIIT: There has been a considerable lack of appreciation of what is meant by the term "a pathogenic staphy. lococcus." Because there is no good test of pathogenicity in animals, such as one finds available with the pneumococcus, pathogenicity of the staphylococcus is defined bacteriologically. The test that wall distinguish most consistently the disease producers from the nondisease producers is the coagulase test. indicating pathogenicity in the sense that coagulase positive strains most commonly are isolated from cases of staphylococcus infection. At least four hemolysins have been separated from the staphy: lococcus, but the one that exhibits the most complete hemolysis on the blood agar plate is alpha hemolysm. It correlates very well with production of coagulase, and with virulence, as estimated hy isolation from cases of human infection. Another characteristic of importance is the fermentation of mannitol, also fairly reliably associated with the pathogenicity of staphylococci. Still another test which indicates potential pathogenicity is the suspectibility of staphy lococci to bacteriophage. Staphylococci that produce coagulase can be phage-typed, those that do not produce coagulase cannot. Staphylocci can be divided into three groups by phage type, which correspond extremely closely with the three serologic types discovered by Cowan, The staphylococcus isolated from outbreaks of food poisoning and from pastries and contaminated materials generally exhibits, in addition to these characteristics, a capacity to liquefy gelatin. It has not been observed that the specific toxins elaborated by staphylococci in artificial media play an important role in the sympiomatologs of the infection in man. Role in cystic fibrosis The relationships between pathogenic staphylococci and cystic fibrosis comprise a field where speculation must hold because there are few facts available which provide information on the problem, Though often present in the throat we would not expect chronic infection with the pneumococcus or streptococcus, because these organisms stimulate an immune response suſficient to terminate infection. The gram-negatne bacıllı are in a minority in cultures of the nose and throat and therefore apparently not usvall available to cause respirator infection 69 ·Summary and conclusions From the 1600 slides studied to date, we hale a few hints that the mucins of patients with cystic fibro-is may contain more numerous weakly acidic groups; they may be slightly more resistant to tryptic digestion; they may have some kind of a bond which is more easily broken by sulfuric acid-permanganate. We can do very little in histochemistry to characterize the pro. lein moieties of these mucoproteins, and we therefore cannot say that they are not abnormal. At any rate, studies on these protein moieties should be pursued Our largely negative results may have some value in indicating aspects of mucoprotein chemistry where any differences, if they exist, do not seem to lie There are many enzymes which might possibly le involved in abnormal mucus formation, if such, in fact, exists. Carbonic anhy. drase is one that might be involved in the sweat gland dysfunction. Preliminary studies have shown that the carbonic anhydrase in. hibitor, Osamar, does not altes the electrolyte concentration in snear toward the pattern seen in fibrocystic disease. Discussion Dx DI STACHESE We have made one biopsy of the pan. creas at a time when the patient had apparently normal pancreatic function. There was no evidence of ohstruction of any kind, but there were inflammatory changes. I should like to raise the question of the frequency of involvement of the bile duct, it has been said that some involvement 2x present in all patients, DR. LAVDIVC: The lesion in the pancreas is not advanced to the same degree in all areas at any one time. Turthermore, in any Een grea there is a slowls progressing lesion, as shown by the number of parents #ho have been ludied clincally, »ho show progressive loss of iryptic activity in the duodenal Hud A normal ares of pancreas cannot be taken as representative of the whole I do not know whether the basic trouble hes in the ducts or aciai, which is really the root of the problem They both seem to be affected in established lesions In nen borns with meconium peus, the acını sectn to be affected first, because plugs can be seen in acini, without obiidus plug in the decea While the balsary tree is much more commonly affected than has been generally thought, a gmificant degree of pathology is quite rare ·studies at Nashsille and New York City. There is a similar degree of resistance to several of our commonly used drugs These resistant strains were predominately group III phage reactors. Although staphylococci were apparently not natively highly susceptible to chloramphenicol, they seem to have less capacity to become resistant to it in rito. In ritro we can unquestionably demonstrate emerging resistance to chloramphenicol after prolonged exposure This is one of the drugs we believe to have a definite place in the treatment of staphylococcal infection Table 20 Antimicrobial susceptibiluy of staphylococci New York, 1953-51--516 strains Na-hulle, 1955 -617 strains Penicillin Tetracyclines percent percent N. Y. Nash Ny. Nash Resistant 72 58 66 51 Intermediate 13 21 Susceptible 15 20 33 18 Streptomycin percent Nash. NY 66 52 1 33 45 1 3 Chloramphenicol percent NY Nash. Erythromycin percent NY Nash, Resetant 9 1 2 86 91 2 6 Intermediate Susceptibile 5 8 97 92 Erythromycin is not used widely, therefore, a high proportion of strains of staphylococci are still highly susceptible to this drug We found that patients who received no antibiotics during hoepitalization had less tendency to acquire the drug resistant strains in contrast to patients treated with tetracyclines and penicillin This represents an astounding influence of medication in an epidemiologic situation. I do not know of any natural situation where differences in the rate of exchange of bacterial fora are so remarkably distinct In early studies there was no predominance of group III phage types in the drug-resistant strains, and the organisms were mostly ·The staphylococcus will grow almost any wherc, grow well through a wide range of pH and in the presence of many substances that interfere with the growth of other organisms. A medium containing 7.5 percent sodium chloride is effective in isolating staphylococci from mixed cultures. This suggests that something in the salt concentration of the pulmonary secretions might be favorable to the growth of staphylococcus, such as has been described in the sweat and saliva in patients with cystic fibrosis of the pancreas. This parasite produces a different type of inflammatory reaction in tissues than does any other organism. Although staphylococci produce a glutinating antibodies when they infect humans, there is no durable immunity. With a staphylococcic infection, there in Tutle or no impetus on the part of the body to cure itself, as far as can be determined with present methods, In the symbiotic relationship which develops in patients with chronic pulmonary dsease and staphylococcal infection, the flareups come along, but the staphylococcal infection tends to remain localized in contrast to infection with highly virulent group A streptococci. Thus, this organism fits well the situation of cystic brosis. Our evidence suggests that, although we are now able 10 ch tinguish among pathogenic staphylococci with our phage-typing eſcorts, any one which produces coagulase and has these other characteristics has the capacity to produce infection 31 man There does not seem to be a specific sub-group of staphvlococc) which can be implicated in this disease. It is likely, iſ there is some factor at work such as heavy ex. posure to one kind of staphylococci or treatment with a particular drug that these patients will respond with changes in staphy fococcal flora in much the same way as do normal people who are carriers of staphylococci. Epidemiology In a recent study of adult patients on a ko-putal Ward, cultures of the nove and throat were obtained every day for fre monthThe results show that there is a considerable umformity in the carrer sale of pathogenic staphylococct in a hospital, which corresponds with scanty reports from other renters The mean rate of positre cultures in this fise and one-hall-month period wa, 19.5 percent. The throat proved to be less often positne for pathogenic staplıy. lococci than the nose. Antibiotic treatment of these patients did not alter the Baphy lococcal carrier rate, possibly because of the high resistance to drugs of the staphylococci present in the hospital. Tahle 20 show the sener- Ersity studies of the strains of staphy loccoccus uluch ne cultured in ·Metabolic Studies of Fibrocystic Disease of the Pancreas DR CBI 4RLES U. LOWE: We hase correlated and compared the data from 11 published metabolic study periods from parents with fibrocystic disease, with those of about 70 period from normal pa. tients in various age groups Statistical treaiment provided several conclusions, We could find no evidence that a high intake of ſat had any effect on absorption of nitrogen (figure 121 The attitude that it is 12 10 FAT INTAKE.am/ky/24 hrs o 20 30 40 50 GO 70 90 NITROGEN, 2 ABSORPTION Figure 12 Relation of nitrogen absorption in fat Intake in patients with biocu rase on normal diete harmful to feed a child with cystic fibrosis of the pancreas a high fat diet appears to lie unsupported by experimental endence. There was an excellent correlation between fat intake and fat absorption It hen the effects of -pecialized diets that included protein hidrolysates or pancreatin kere compared with the effects of normal diets, in children with filtex satse drea-e, there was a staffealls gnificant difference in the mitrogen retention berneen the two 73 ·drug.susceptible; these observations were made before antimicrobial drugs were available. Summary In hospital populations the use of antibiotic- cau change the rate of acquisition of drug-resistant strains of staphylococci. There is sound epidemiologic evidence that this is truly acquisition and not mutation within the patient. Acquisition may be modified ma. terially by choice of antibiotic drugs. Prolonged use of a drug such as tetracycline will certainly guarantee the development of druga resistant strains, at the same time probably gradually interſering with the therapeutic result. This poses enormous problems in therapy of staphylococcal infections because of their chronic nature. We must learn to alternate our drugs, and hope for new antihiotics to replace those to which the organism has become resistant. I prefer to give patients no more antimicrobial treatment than necessary Discussion Dk. PAUL R. PATTERSOX. Does Krught think it would be worth while to isolate patients who have a diagnosis of cystic fibrosis and have been on antibiotics for a period of time, or any patients who have been on antibiotics, for the protection of surgical pattents and children in the hospital? DR. KNIGHT: That question is coming up more and more, but it is not so much a matter of volation as it 19 of drug treatment Obviously, a patient under treatment with one of the major druga presents a difficult situation I do not know of any sers satisfactory isolation procedure. someone has to handle the patient Even in taking throat cultures strains may be transferred from person to person DR. DI SHT'AGNESE Some patients who are said to have re. sistant organisms on the ba-s of tents of penicillin or streptomycin sensitivity actualls improve clinicalls when large doses of the antı biotic are given DR. KIGHT: We must realize that te-ts in utro demonstrate only a limited facet of the characteristics of drug resistance, and there is no doubt that benefit can be gained by increasing dosage. in special situations seen, in face of resistance in titro.

·Respiratory Acidosis Basic Physiology DR. FRIK1 BRLCK: Re-piratory acidosis has been seen in some patients with cystic fibrosis of the pancreas It is due to the retention of metabolically evolved carbon dioxide, which disturbs the ratio between bicarbonale and carbonic acid in the blood and alters pH when the retention becomes acute in chronic cases, where the arte. ral retention of co, is high, retention of bicarbonate al o occurs, and the p!I may be close to normal Figure 13 is a diagram arranged so that on the horizontal axis the arterial co2 tension can be read in millimeters of mercury In CONTENT MARTIN OOO PLASNA COZ 20 yo 30 60 360 do vụ , co, LO O 78 76 74 72 TOH pH & 9 Figure 13 Data ultaned from patients with cutie hlusot showing the piesalene of some degree of reparatory autdusjs this diagram the points represent all the data obtained in our patients with cystic fibrosis of the pancreas, and indicate that most have come degree of respiratory acidosis Vlechanisms Table 21 lists the basic causes of respiratory acidosis. An increase or decrease in alveolar ventilation determines the oxygen 75 ·groups of patients. The predictability of nitrogen retention is the same, whether these children are given normal diets or specialized diets, with or without pancreatin, Caloric intake and nitrogen retention show a high degree of correlation in these patients. Nitrogen intake and nitrogen relention show a poor correlation. At constant caloric intake, there is no predictability of the rela. tion between nitrogen intake and nitrogen retention. If the nitrogen intake is kept constant, there is a high and significant degree of cor. relation between caloric intake and nitrogen retention. For comparison with normal patients, we lumped together all data from children with fibrocystic disease regardless of type of diet, but separated them according to age, using the years as the dividing point. Most of the balance material available on infants was for under one year of age, hut there were tuo published studies in children at 19 and 22 months of age. Te found that children with fibrocystic disease of the pancreas have the same degree of predictability of nitrogen retention with a gnen intake of calories or nitrogen as do normal children, with one important exception Plotting the predictability shous that as either caloric intake or nitrogen intake drops in these patients, they go into negative nitrogen balance at a higher intake than do normal children Comparison with studies on undernourished children showed that they behave much like children with fibrocystic disease of the pancreas, in that they have an auidity for nitrogen, Discussion Dr. ZEIZER Is there animal experimenial data that would ou a similar nutritional relationship between animals with liga. tion of the pancreatic ducts or other forms of pancreaue insufficiency, and normal animals? DK LOWE. Papers by Handelemah. Greenberg, Rouman and Wann have shown a similar correlation between caloric intake and nitrogen retention ·TI 111 17 .111 1 fusion The alveolar co, and arterial cop tension are usually not changed. In some cases there may be decreased arterial co, tension and respiratory alhalo is because, if there is sufficient anoxia to stimulate 11 whether the shunt occurs in the lungs or in the heart. When the distribution of gas is unes en and when diffusion is impaired, alveolar oxygen may be high, especially if there is hyperventilation. There is considerable difference between the alveolar oxygen tension and the arterial orygen tension, ie, the alveolararterial orygen gradient is increased. With hypoventilation, ie, decreased alseolar ventilation, the alveolar oxygen is low, and the gradient between alveolar and arte. rial oxygen should be normal. Hypoventilation is the outstanding mechanism in the production of re-piratory acidosis. Causes of hypoventilation I have listed in table 22 the causes of decreased alveolar venti. lation. Chef among these is depression of the respiratory center. Table 22 Causes of decreased alteolar 1 entilation 1 Depres-ed respiratory center 2 Paralysis of respiratory muscles 3 Decreased thoracic space or mobility Increased intrathoracic pressure S Decreased dr-tensibility of lung 6 Increa-ed respiratory dead space Oltruction + 7 Some respirator pluysiologists believe that there is no respiratory acidosis without depression of the respiratory center Increased intrathoracic pressure is also an important cause of hypoventilation, because air can not move against a pressure grad The outstanding example of this is found in teneion pneu. mothorax jent ·and con exchange. Afinule s entilation or tidal volume does not give a good indication of whether entilation is adequate. The effect of hypoventilation or decreased alveolar ventilation is more marked on co, tension than on that of oxygen. This has to do with the fact that oxygen diffuses in the gaseous medium of the airways more rapidly than cos then diffusion through the alveolar membrane is im paired, as in pneumonia or pulmonary emphysema, there is little effect on the arterial co, tension, hecause the diffusion coefficient Table 21 Effect of functional derangement on blood gas tension ABNORMALITY OF VENTILATIO Y Paco Pao. Alseolar ventilation serra ed Alveolar ventilation decreaned Diffusion impaired Distribution of gas urien of coz is about 20 to 25 times as high as that for oxygen, and the patients deselop marked decrease of the arterial oxygen tension long before there is any effect on CO2 Distribution of gas in the lungss an important factor there is onlt one lung with normal ventilation, and it is well per. Suced *th blood, the blood ossgen and cou will be normal However, if there are two lungs, only one of which is venulared as before, but both luny are perfused with blood. blood will return from the nonsentilated lung and mir in the pulmonary seins uith the blood from the normal part Theoreticall the origen satura Jion in the mixed Wood wall be los and the ci), fen-yon logh The co, in theme cd-es, houpier, in sual)s not affected because Woorl from the pulmonars vein stimulates the rewpurrators center causing lypenentilation of the well-senulated lung The co, tension in the blood coming from the good lung can thereby be dimisished to levels of 30 or 15 mm of mercury then mired >>h blood from the nonsenulated part of the lung. the co, tension in the mr. lure may be normal This cannot happen with oxigen because the hemoglobin in the blood perfusing the normal lung in saturated 10 almost in percent and cannot take up any more oxigen ·Cardiac and Pulmonary Complications in Fibrocystic Disease of the Pancreas Cardiac complications DR. STEPHEN 18. Royce: Our experience with cardiac complica. tions in fibrocystir disease is limited to the rather poorly understood phenomenon of cor pulmonale. Approximately one-third of the children who die with fibrocystic disease show, at some time, premortem evidence of clinical heart failure The lises of these children are usually terminated in an acute episode of irreversible heart failure The chest findings at postmortem are such that evidence of pul. monary heart disease might also be expected. Characteristically, there are an emphysemalous chest and hyperexpanded lungs that do not collapse when they are cut the diaphragm is low and flat. All these factors predispose to right heart strain the heart is usually small in bulk, but above normal in weight The anterior wall of the right ventricle is usually thicker than normal, a sign no longer considered a reliable index of right ventricular hypertrophy. Cardiac catheterization studies Since the physiologist regards a mean pulmonary artery pres. sure of 25 mm or more of mercury as the essential criterion for right heart strain, it was arranged to subject ll of our patients to cardiac catheterization tudies The patients ranged in age from 4 to 10 years All had varying degrees of pulmonary insufficiency as determined by ausculation, 10entgenographic study and exercise tolerance, and all had a chronic cough. The studies we performed were standard for cardiac patients, with the catheter introduced into the left arm, thence through the right heart and into the pulmonary artery. The correlation between high mean pulmonary artery pressure and low arterial orvgen saturation is shown in table 23. Pulmonary heart disease occurs in some children with fibro. cystic disease and, in general, the degree of right heart strain is in direct proportion to the degree of lung involvement. Whether right heart strain is a dominating factor in determining the prognosis in flrocystic disease is still unknown. 79 ·Obstruction is a possible cause of hypoventilation. However, there are practically no reports of respiratory acidosis described as caused by hypoventilation due to obstruction. Respiratory acid. osis is characteristically absent in asthma. aris. Respiratory acidosis in cystic fibrosis Since obstruction is described as the primary and most char. acteristic anatomie lesion in the lungs of patients with cystic fibrosis of the pancreas, it seemed worthwhile to compare the evidence for obstruction and hypoventilation in these patients, as measured by the pneumotachograph, with results of determination of gases in arterial blood. Six patients were studied and all had evidence of obstruction, by pneumotachographic analysis Determinations of gases in the blood show that slight or moderate obstruction does not produce sufficient hypoventilation 10 cause co2 retention, only three of the patients had respiratory acidosis, but all had reduced blood oxygen tension The lowering of oxygen tension in arterial blood is apparently not caused by hypoventilation. In fact, the alveolar-arterial oxygen gradient was increased in all these patients, probably because of diffusion dificulty or uneien distribution of gases. Since two patients had no pneumonia, exudate or bronchitis at the time they were studied, there was no reason to assume an comic barrier to diffusion in these cases. We can postulate, there. fore. that most of the patients had uneven distribution of gases, probably due to atelectalıc areas, or possibly due to collateral cara culation in the lungs, shunting blood anay from the right to the left side of the heart. It has been noted that adults with respiratory acidowe shou depression of the respiratory center and, therefore, breathe mainly under the stimulus of the reller renters which are stimulated by anoria If you remove the anoxic stimulus lm gning these patients oxygen the results may be disastrous We have done only tno preliminary studies on the effect of oxygen therapy in patients with fibrocytic disease Our findings corroborate those from studies on anults that re-puatory acidos can be a gre:ateri bu origen therapy because ventilation is de creased and co, tenstein increa-ed. Clinically, we have not seen adiere effects of administration of oxygen and } would not advice withholding oxygen from the parents although we must be aware of its potential danger. Adults with emphasema hale been treated with combined artificial respuration and origen ·ſound evidence that exercise tolerance is impaired, and some have clubbing of the fingers, even in the absence of any other signs of lung disease Our experience suggests that eventually they will find their lives dominated by the state of disease in their lungs. McIntosh, in studying a group of children over 10 years of age, found that all reported some coughing, even though their early childhood had been characterized primarily by gastrointestinal disturbance, with little coughing during the first year of life. Correlation of age with the onset of cough in some 300 cases is shown in figure 15. Most patients begin to cough in the period hefore six months of age LP Dead Living A 35 30 25 RUE 20RROR 15- mos 12 18 24 30 36 Figure 14 Length of life compared with mean pulmonary artery pressure No correlation 1 keen Before antibiotics were available, the survival of a child who roughed during the first weeks of life was generally measured in terms of months Of 38 infants seen with persistent cough before the age of two months who were treated with antibiotics, one-third were dead within six months. Despite antibiotics the early onset of cough continues to carry & poor prognosis, although survival time has been lengthened. Coughing suggests that the earliest problem is one of intra luminal obstruction Relief of obstruction is the objective of cough. ing as well as of the therapeutic regimen The effectiveness of therapy is often assessed by the degree that coughing is relieved ·Figure 14 shows a three-year followup of the original studies. The lack of correlation is apparent. The usefulness of cardiac cath. eterization as prognostic tool would appear to be limited. The studies showed, however, that there is laboratory evidence of right heart strain, which does correspond, roughly, to degree of pulmonary insufficiency. It does not have a consistent relationship with the shortened length of life. The pathogenesis of cardiac failure in cystic fibrosis disease makes it unlikely that any therapy would have more than limited effectiveness Digitalis and mercurials provide only temporary re. lief. Cardiac failure may be reversible, but only for a matter of weeks It is a sign that the lung field has lost its elasticity. To the Table 23 Relation of pulmonary artery pressure to systemic oxygen tension Arterial oxygen saturation Pulmonary artery prossur systolic moan 1B 30 20 40 25 33 26 3 28 10 31 +5 35 12 35 35 81 81 81 86 85 86 82 89 77 74 71 handicap of pulmonary hypertension is added the handicap of alveolar congestion with increased pulmonary rigidity. In our expe. rience there is no sign that has norse prognostic import than the onset of congestne failure Effect of pulmonary disease The general health of children with fibrocystic disease, their appetites, activities, and longesls are closels rorrelated with their degree of lung ini oliement Approximately fise percent of these children do not cough and do not hase chronic re-salon disease. Even among these we have ·The "Yentilation factor" is an average of the timed vital capacity, the maximum breathing capacity and the residual capacity expressed as percentages This roughly approximates the effectiveness of the lungs as a bellows mechanism and gives some measure of pulmonary dysfunction. The ventilation factor in this patient was 38 percent of normal, which means there is emphysema, with a tremendous amount of air trapping Table 24 Pulmonury function studies Predicted Observed | % Normal 3136 3136 99 99 1661 1292 298 51 53 41 30 52 Lung volume measurements Vital capacity, ml Timed vital capacity, 3 sec mol Max. breathing capacity 1/min Mar--after bronchodilator 1/min Residual capacity Residual air capacity, ml Total lung capacity, ml Residual capacity of total lung capacity, ml Ventilation factor, percentage 15 784 3920 20 100 2.3 1482 31-16 47 38 190 80 38 Evaluation of pulmonary disease The extent and severity of the pulmonary disease can be approximated using the ratings shown in table 25 This technique affords means of charting changes and making comparison of the patient with other children Principles of therapy Unless started before coughing begins, we should regard therapy in fibrocystic disease as active, not prophylactic treatment There are two prophylactic measures, both surgical, that may be indicated after onset of pulmonary symptoms. It has been shown that in heart failure, rales are often more pronounced on the right side of the chest than the left. Long before heart failure occurs, however, the child with fibrocystic disease is likely to show auscultatory evidence that the right side of his chest is more severely involved than the left. The syndrome of lobar atelectasis involves most frequently the right upper lobe, then the ·Once the cough becomes persistent, progressive involvement is the rule. Evidence that the child is losing the ability to ventilate his chest increases. Coughing spells are attended by flushing, then cyanosis, and, finally, by complete exhaustion. The chest movements become more rapid but also less eſficient. Then the chest becomes fixed in a position of full inspiration, and respiratory activity is limited. The terminal phase is that of severe emphysema, accompanied by a deep, wet, ineffective cough that exhausts the patient but does not produce sputum. The data in table 24, taken from observations on one child, are representative. The administration of a bronchodılating agent increased her ability to rentilate the chest to a significant degree. 100 No Cough 17 J. 50 6-12 mos / yr. mo mos Figure 15. Age of onset of cough in 300 patients with cystic fibrosis of The pancreas ·derosol therap? in advanced lung disease there is evidence that a barrier develops between the alieolar wall and the blood stream, and that antibiotics given by mouth or injection do not reach the site of infection. Here the use of aerosol therapy may be effective. Table 26 lists several aerosol mixtures. Zephiran®, in combination with streptomycin and penicillin, is particularly effective. We have limited experience with Tryptar. Our impression is that it is an extremely irritating material when given at the recommended rate. Of all the combinations used, that of the bronchodilator, Isuprel and Terramycin® is, perhaps, the most effective. Aerosols are most effective when the particle site is within a critical range of .05 to 3 millimicrons in diameter. The Bennet mask, the DeVilbiss No. 40 nebulizer and the Vaponefrin nebulizer have been used satisfactorily. The DeVilbiss 501 is a satisfactory compressor. Table 26 Aerosol agents used as mixtures Zephiran "Mistogen Solution" Tryptare Ephedrine Alkaloid Dornaset Propylene Glycol Allevaire (Triton A-20) Glycerine Isuprel Dishlled water ad < Antibiotic Combinations > 08 Cm 25 cc 25 cc 1000 cc We use aerosols, 15 ml of solution, for 20 minutes three times a day, and usually try to follow with a period of postural drainage This technique is practical for use with young children. As respiratory insoliement increases, the effectiveness of aerosol therapy decreases rapidly. In the fixed emphysematous state, aerosols probably penetrate to, but not beyond, the site of obstruction Poorly ventilated or nonsentilated areas of the long receive no therapeutic benefit to reach these areas, more compler equipment, such as the intermittent positive pressure breathing machine, must be used Physiologic evidence suggests that this type of therapy may possibly arrest the progression of emphysema in children Lacking knon ledge of the specific defect in the lungs, we are faced with the necessity of treating signs and symptoms as ther arise, rarely being able to anticipate or judge the effectiveness of our ·right middle lobe, and rarely if ever involves the left side of the chest. Since prognosis in this syndrome is poor, we might expect that resection of the lobe or part of the lobe would be followed by improvement. Eleven of 23 patients examined at more than 10 years of age showed an involvement of the paranasal sinuses. Polypectomy Table 25 Clinicul rating 5 4 3 Exercise tolerance Cough normal Jubsent Breathing euprea Clobbing absent 2. limited sey. limited home Izero (rare 11 session constant constant & daily mellectne hyperpnea dyspnea c dyspnea orthopnea exertion at rest angle lunoderate severe serere with reversed cyanoare coarse scattered general. sticky breath trales fized rales sounds Cheat clear Anscultation or other treatment of the nasal snuses might also be considered prophylactic therapy. Other therapeutic measures seem entirely active and should be planned for home care rather than hospital care Antibiotic therapy A difference in survival time following the use of penicillin in fibrocystic disease has been demonstrated. An even greater increase in survival time will follow the use of the nener antibiotics, prefer ably used in combination and alteration Dosage should he large 30 to 50 mg/kg per day Postural drainage With an efficient cough the patient removes debris consisting of bacteria, mucus and casts of the bronchioles Therapy must in crease the productiveness of the cough. Placing the patient is a jackknile position, with pillowe under his abdomen, and leaving him for twenty or thirty minutes and placing blocks under the foot of the child's bed to maintain bronchial drainage during nape and during sleep are effective measures for bringing up sputum ·Genetics DR. BARTON CHILDs: Considerable evidence has accumulated suggesting that cystic fibrosis of the pancreas is a genetically deter mined condition. Affirmation of a single gene hereditary characteristic in humans depends, in part, on agreement of observed distributions of affected and unaffected individuals within family groups of various sizes, with distributions expected according to specific hypotheses of modes of inheritance of genes. In most instances, the particular mode of inheritance will depend on the choice of criteria by which it is determined whether or not an individual is alfected. Thus, whether we consider a gene as dominant or recessive depends on whether we can detect both heterozygote and homozygote A dominant gene is one that will produce equal effects in both heterozygote and homozygote. It seems that many so-called dominant genes do not fulfill the definition of this term, because the manifestation in homozygotes is un. known. It is probable that every gene has a function that can be measured if detected, and we may escape the rigidity inherent in the use of the terms dominant and recessive if we consider what genes do in the homozygous and heterozygous states. When the genetics of cystic fibrosis of the pancreas was first studied, it seemed that the disease was due to a gene which expressed itself only in the homozygote, a perfect recessive. This assumed rather rigid criteria for diagnosis of the disease, including in most instances marked reduction or absence of pancreatic enzymes as well as pulmonary disease, or diagnosis at autopsy Assuming the proposed gene to behave as an autosomal recessive, statistical agree. ment has been demonstrated in several studies between the number of patients expected in families and the number observed. Table 27 shows the results of an analysis by the method of Hogben of data pooled from various investigations There is no significant departure from erpectation in the total values, nor is fare . fiftoronna hamnhn - L, - the distribution within families that he see in these data.

·therapy. It seems clear that therapy aimed at the relief of intraluminal obstruction must be given continually and for long periods of time, and must be planned as home therapy. Discussion DR. DI SANT'ACNESE. The cardinal and primary occurrence in the lung in fibrocystic disease is bronchial obstruction leading to generalized obstructive emphysema. With obstruction, secondary bronchopneumonia occurs. If the infection continues for months or years there may be permanent damage to the bronchi and progressive bronchopneumonia which is not checked by antibiotics This leads to pulmonary insufliciency and death, As long as there is no irreversible damage to the bronchi, antsbotic therapy is often successful in checking the process. DR ANDERSEN. We have been discouraged with the results of postural drainage. In contrast, we find there is some benefit in allowing the children to be up and around, at least for intervals during the day. It seems that in the latent stage of this disease, actuutty is more effective than postural drainage. Our experience with lobectomy includes seven children, two of whom were operated on at other hospitals We believe that there is a limited number of patients for whom this operation is appropriate Our criteria for lobectomy are as follows: 1) that the segmental atelectasis has been localized for at least six months; 2) it intohes not more than two lohes, usually the right upper and maddle, 3) the other lung must be free or nearly free of active infection following antibiotic therapy We have observed spontaneous re expansion of the atelectatic lobe following antibiotic therapy, some times several months after its discovery. The patient should therefore be given a trial of this therapy before surgery is seriously con sidered ·among individuals who could be heterozygotes. Since this time, parents and sibs have been found showing either high sweat electro lyte values alone, or in combination with obstructive emphysema and nuante Innans that antall indian that many of these show values of sweat electrolytes which are some what lower than those found in fully affected patients. Hypothetical mechanisms Though we assume the disease to be caused by a mutant gene or genes, which affect a wide variety of organs, we do not know the precise mechanism through which such a gene or genes act. Nor do we know the causes of variation in the expression of such genes in patients and their relatives. The question arises whether there might be more than one genetic entity, that is, several genes operating in different ways For instance, there might be two or more genes at different loci, not interacting, and whose modes of action are quite distinct, although producing certain phenotypic similarities. This might account for differences between families in sweat electrolytes and pulmonary disease among possible heter. ozygotes Alternatively there might be a system of multiple alleles, genes occupying the same locus, but whose actions are quantitatively dif. ferent Finally we must consider the effects of genotypic or environ. mental modification of the expression of such genes. It is well known that the phenotypic expression of single genes is subject to modification by other genes in the genotype, and that such expression depends upon the environments in which the gene acts These hy. potheses might account for some of the variability in the clinical manifestations among homozygotes, in which all the exocrine glands may be involved, or different ones in various degrees They might also account for the fact that some probable heterozygotes may show not only sweat electrolyte abnormalities, but also chronic pulmonary changes, while others exhibit only some alteration in the sweat Thus parent-child combinations showing partial forms of the disease in probable heterozygotes may be due to a gene which in such families could be called incompletely dominant This phenomenon is seen in patients with sickle cell disease who have thrombotic phenomena in the presence of electrophoretic patterns showing both normal and sickle cell hemoglobins. It is compatible with what is known of the action of natural selection, which works toward suppression of the effects of deleterious genes, causing them to evolve in the direction of recessiseness ·Other criteria must be fulfilled to provide support for this hypothesis. Males and females should be equally affected, and this was found to be so. No affected parents were noted. Of 38 step. children studied none were affected. If we assume the disease to be found among heterozygotes, approximately one-quarter of the step.children should be affected. if, on the other hand, the disease is found only among homozygotes, the probability of finding affected individuals among step-children would be one-quarter the probability of the common parent marrying another heterozygote Table 27 Siblings in family No. of families I 2 3 4 5 67 64 65 22 13 3 5 Total siblings 67 128 195 88 65 18 35 8 9 Correspondence of expected and observed distribution of affected individuals within families Observed | Expected Variance 67 67 0000 70 7311 7700 90 8431 17.095 36 32.18 9240 25 21.31 7.700 6 5.48 2.328 12 10.10 4 850 2.22 1 172 1 380 530 3.181 1 3 9 10 2 20 7 243 663 317 303 50 51 739 EV54.730 = 7 40 D = (317303 50) = 1350 D 13.30 7.40 There were also a fen affected collateral relatives, a phenomenon we would expect, since, of the disease appears only in homozygotes, the incidence among collateral relates should be several times thal among all the population, though still manyfold less than within immediate families. Statistical evaluation alone does not prove heredity, and the hypothesis would be founded upon firmer ground if we could find a characteristic among parents and other relatives which would satisfy mendelian requirements. Unul the discovers of the elesation in sueat electrolytes among some of the relaties of alleeted polenta, there was no published esidence of ant effect of the presumed gene ·among individuals who could be heterozygotes. Since this time, parents and sibs have been found showing either high sweat electro- İyte values alone, or in combination with obstructive emphysema and chronic bronchopneumonia. It appears that not all individuals who may be presumed to be heterozygotes show abnormality. In fact, the data of di Sant'Agnese indicate that the majority of such individuals are clustered within a few families. It may be significant that many of these show values of sweat electrolytes which are some what lower than those found in fully affected patients Hypothetical mechanisms Though we assume the disease to be caused by a mutant gene or genes, which affect a wide variety of organs, we do not know the precise mechanism through which such a gene or genes act Nor do we know the causes of variation in the expression of such genes in patients and their relatives. The question arises whether there might be more than one genetic entity, that is, several genes operating in different ways. For instance, there might be two or more genes at different loci, not interacting, and whose modes of action are quite distinct, although producing certain phenotypic similarities This might account for differences between families in sweat electrolytes and pulmonary disease among possible heterozygotes Alternatively there might be a system of multiple alleles, genes occupying the same locus, but whose actions are quantitatively different Finally we must consider the effects of genotypic or environ. mental modification of the expression of such genes It is well known that the phenotypic expression of single genes is subject to modification by other genes in the genotype, and that such expression depends upon the environments in which the gene acts. These hy. potheses might account for some of the variability in the clinical manifestations among homozygotes, in which all the exocrine glands may be involved, or different ones in various degrees. They might also account for the fact that some probable heterozygotes may show not only sweat electrolyte abnormalities, but also chronic pulmonary changes, while others exhibit only some alteration in the sweat Thus parent-child combinations showing partial forms of the disease 19 probable heterozygotes may be due to a gene which in such families could be called incompletely dominant. This phenomenon is seen 1 patients with sickle cell disease who have thrombotic phenomena in the presence of electrophoretic patterns showing both normal and sickle cell hemoglobins It is compatible with what is known of the action of natural selection, which works toward sup. pression of the effects of deleterjous genes, causing them to evolve in the direction of reressiveness.

·Other criteria must be fulfilled to provide support for this hypothesis. Males and females should be equally affected, and this was found to be so. No affected parents were noted. Of 38 stepchildren studied none were affected. If He assume the disease to be found among heterozygotes, approximately one-quarter of the step-children should be affected. If, on the other hand, the disease is found only among homozygotes, the probability of finding affected individuals among step-children would be one-quarter the probability of the common parent marrying another heterozygote 2 Table 27 Siblings in family 1 2 3 No of families 67 64 65 22 13 3 5 1 1 2 Total siblings 67 128 195 88 65 18 35 8 9 20 663 5 6 7 8 9 10 Correspondence of expected and observed distribution of affected individuals within families Observed Expected Variance 67 67 0 000 re 73.14 7790 90 8431 17 095 36 32 18 9 240 25 21 31 7.700 6 5 48 2 328 12 2010 4850 1 2 22 1.172 2 43 1.380 5.30 3.183 317 303,50 54 739 3 213 S.E. 151739 = 7.40 D = (3173113.50) 13 50 D 1&2 7 40 There were also a few affected collateral relatives, a phenomenon we would expect. since, if the disease appears only in homozygotes, the incidence among collateral relatives should be several times that among all the population, though ss!! manyfold less than within immediate families. Statistical evaluation alone does not prove heredity, and the Lunotheria would be founded upon firmer ground if we could find a HUL ILIEU u 4LUCU, sweat electrolytes among sume vi there was 110 published evidence of any effers of the presumed gene ·expression of the disease in homozygotes, but in only partial expres. sion in some heterozygotes. Thus we may say that in most families the recessive hypothesis is fulfilled, while in some the gene may be ralled partially recessite or partially dominant. Discussion DR. ROYCE: The statement has been made that this is the most common lethal gene in the population. Is that true? DR. CHILDs. It may be so The incidence of the disease has been estimated to be between one per 1,000 and one per 10,000. Assuming no selective advantage for the heterozygote, an assumption which may not be justified, the calculated mutation rate would be one per 1,000 to one per 10,000 mutations per locus per generation This is a situation quite outside our experience with any other human disease, and for this and other reasons this calculation must be considered with much reservation. Most mutation rates have frequencies less than one per 25,000 DR. DI SANT'AGNESE: Childs has appropriately emphasized the variability of expression of genes With regard to the sodium and chloride concentration in sweat we have taken 60 mEq/l of sodium or chloride as the upper limit of normal, because the concentrations in all our patients with known cystic fibrosis were above that There is no reason why heterozygotes should not have slightly lower concentrations However, electrolyte concentrations vary so much in apparently normal individuals that it would be hard to recognize the abnormal DR CHILDs. There is no reason at all why the sweat abnor. mality cannot be as variable as anything else. However, the closer we get to measuring the fundamental gene action, the less modifica tion or variability we find in the expression of that characteristic in those individuals who hold it. DR. SIIWACHMAN. I should like to bring up a point that has been raised in early investigations, that perhaps this disease is in some way related to one of the blood groups This is not the case We thought with Steinberg it would be worthwhile to investigate the possibility of a genetic linkage between the blood groups and this disease, because the hereduary pattern of cystic fibrosis is simple and known. Analysis of data from 70 families uncovered no relationship between cystic fibrosis and any of the following blood groups. ABO, Rh, Mys, Kell, Duffy, Lewis and r. and a few others that are not common ·The most likely hypothesis, then, is that which calle for a gene or genes which cause the disease in homozygotes and cause partial expression or no expression in heterozygotes. Though there is insufficient published data to favor strongly either the assumption of multiple alleles, or its alternative, genes at more than one locus, it should be possible to test these hypotheses by means of measurement of sweat electrolytes in parents, sibs, and col. lateral relatives In general, assuming genes at different loci, both parents of a particular family and some of their relaties should show the same expression in terms of sweat electrolytes. If, on the other hand, a system of multiple alleles is responsible, we should find not two, but three types of family; those in which both parents and their relatives show the same kind of expression, as well as some families in which one parent and some of his relatives show a mani festation which the other does not Genetic significance of abnormalities in sweal Speculations on the genetic aspects of this disease have been based on possible distributions within families of the one common characteristic which appears in nearly every patient who has this disease, and in some of their relatives, that is abnormalities in sweat electrolytes Sull, we find certain parents who, according to the genetic hypotheses we are considering, should be heterozygotes, but do not show the characteristics. Although the sweat electrolyte test will detect the homozygote in nearly every case, it performs this function for heterozygotes with distressing infrequency and of seems Iskely that this test, though the only one currently available, is not the final determinant for the presence of the gene in single dose. It should be possible, in theory at least, to find some quite con sitent characteristic of the gene action which would expose its presence wherever it exists. This speculation is based on the assumpmeasurasso n. the less modification there will be of that effect by the genotypic or external environment Such a determinant will be closely associated with whatever turns out to be responsible for alteration in function of such a wide vanety of organs in the body in this disease Summar It seems quite likely that cystic fibrosis of the pancreas results from the effens of gene mutation There might be either a system of alleles at one locus, or genes at different loci, which result in full

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·DA. HEKBERT J. BARTELSTONE: Aside from one short publıcation regarding pilocarpine and its effect of the pancreatic tissue. there has been no report of direct study of the relationship of autonomic regulation to the disease. An avenue of research might be opened up by exploring some of the effects of electrically induced or drug.induced stimulation to some of the exocrine glands whose function is influenced by the autonomic nervous system. There might be some alteration esther in the production of acetylcholine esterase or the permeability of receptor sites, which may be one of the underlying factors in the disease. Overstimulation of the parasympathetic nervous system or cholinergic fibers for a prolonged time might result in some of the findings that have been noted clinically, Conclusion DE FRAZIER. From the thoughts presented here we may con. clude that the genetic basis of this disease has been reafirmed. It has not been possible to discover, by either biochemical analysis or histo. chemical staining techniques, a qualitative abnormality of the mucopolysaccharides in those exocrine glands which secrete mucus A defect in production or availability of energy for elaboration of secretory products has been postulated on the basis of data oh. tained from study of the concentrations of electrolyte in sweat The clinical manifestations may be secondary to such a basic delect Application of basic physiologic principles to the discussion of the deranged functions of vanous organs has facılared a clear and lagical exposition of sound therapeutic proceri res

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