GASTROENTEROLOGY Copyright 1969 by The Williams & Wilkins Co. Vol. 56, No.4 Printed in U.S.A. EFFECT OF EXCLUSION, ACIDIFICATION, AND EXCISION OF THE DUODENUM ON GASTRIC ACID SECRETION AND THE PRODUCTION OF PENTAGASTRIN-INDUCED PEPTIC ULCERS IN CATS S. J. KONTUREK, M.D., J. DUBIEL, M.D., AND B. GABRYS, M.D. I Department of Medicine, Medical School, Krakow, Poland Gastric acid secretion and peptic ulcer production induced by pentagastrin infusion were studied in two separate series of cats after the following procedures. (A) Transplantation of the common biliary duct and pancreatic duct to the proximal jejunum. (B) Formation of the duodenal Thiry fistula open distally for perfusion with saline (Bl) or acid (B2). (C) Excision of the duodenal fistula. (D) Transplantation of the biliary and pancreatic ducts to the distal jejunum. Exclusion (Bl) and subsequent excision (C) of the duodenum were followed by an increase in basal acid secretion and in a shift to the left of the dose-response curve to pentagastrin. The maximal acid output was significantly higher only after the excision of the isolated duodenal loop (C). Basal achlorhydria and a decrease of the dose-response curve to pentagastrin with a shift to the right occurred after acidification of the duodenal loop (B2). The greatest production of peptic ulcers induced by 24-hr infusion of pentagastrin in the dose of 16 ~g per kg per hr was found in the group of duodenectomized animals with the biliary and pancreatic ducts transplanted to the distal jejunum (D). No statistically significant difference in the ulcer area was found between the group of cats with the duodenum left in the normal position and with the bile and pancreatic juices diverted to the upper jejunum (A), and the groups with excluded (Bl) or resected (C) duodenum, and with the alkaline juices diverted to the upper jejunum near the gastrojejunal junction. The acid perfusion of the duodenal loop (B2) completely prevented the production of peptic ulcerations. It is concluded that the inflow of bile and pancreatic juices plays an important role in the production of pentagastrin-induced peptic ulcers in cats. Diversion of bile and pancreatic juices from the duodenum to the jejunum has Received May 14, 1968. Accepted November 11, 1968. Address requests for reprints to: Dr. S. J. Konturek, I Department of Medicine, Medical School, Krakow, ul. Kopemika 17, Poland. This investigation was supported in part by Grant TW 00217 01 from the National Institutes of Health, United States Public Health Service. The authors are deeply grateful to Dr. J. D. Fitzgerald for supplying the pentagastrin used in these experiments (lei 50,123). 703 been shown to increase the rate of appearance and the area of pentagastrin-induced ulcers in the duodenum of cats.! The present report was designed to study the effect of exclusion of the duodenum as a Thiry loop, the effects of its acidification and subsequent excision on gastric acid secretion under basal conditions in response to graded doses of pentagastrin, and on the development of peptic ulcers produced by continuous intravenous infusion of a constant dose of pentagastrin.
704 KONTUREK ET AL. Vol. 56, No. 4 Materials and Methods Cats of both sexes, weighing between 1800 and 2300 g, were divided in two series, I and II. Series I, consisting of 10 cats provided with gastric fistulae according to the method previously described,3 was used to study secretory responses to pentagastrin. Series II, composed of five separate groups, each of 8 to 13 cats, was designed to study the appearance and the number of ulcers. Surgical Procedures Five surgical procedures were performed in both series of cats. (A) Transplantation of the pancreatic and common bile ducts with the surrounding piece of duodenum to the proximal jejunum just beyond the ligament of Treitz. (B) Formation of the duodenal Thiry fistula. The pylorus was resected and the proximal duodenum was closed. The distal end of the duodenum was then brought to the skin as a mucous fistula. A polyethylene tube (Intramedic PE 190, 2 mm external diameter) was introduced into the excluded duodenal loop and its tip, which was provided with small openings, was attached to the blind end of the loop and used to perfuse it either with saline (Bl) or with acid solution (B2). Gastrointestinal continuity was restored by end-to-end anastomosis between the stomach and the jejunum. (C) Excision of the duodenal fistula. (D) transplantation of the pancreatic and bile ducts with the surrounding piece of duodenum further down into the jejunum, about 25 cm distal from the gastrojejunostomy. Secretory Procedure The experiments, performed on conscious cats were started 2 to 3 weeks after each stage of the surgical procedure. Before the secretory test food was withheld from the animals for at ieast 18 hr. Throughout the experiment 0.15 M sodium chloride solution was continuously infused intravenously at a rate of 20 ml per kg per hr by a peristaltic pump (Unipan, Poland). Basal secretion was collected for two 15-min periods and pentagastrin (lci 50,123) was then added to the saline infusion, starting with the dose of 1 p.g per kg body weight per hr. The dose of pentagastrin was increased 2 to 4 times every 60 min during the 5-hr secretory tests, and ranged from 1 to 32 p.g per kg per hr. Gastric juice was collected every 15 min. The volume of the juice was measured and titratable acidity was determined using phenol red as an indicator. Acid output at a given dose level of pentagastrin infusion was calculated from the two highest consecutive 15-min periods of acid output multiplied by 2 and expressed as milliequivalents per hour. Throughout the experiments on cats with the duodenum excluded, the duodenal loop was perfused from a peristaltic pump at a constant rate of 20 ml per hr. The perfusion fluid was 0.15 M sodium chloride solution or 0.10 N hydrochloric acid solution. Production of Peptic Ulcers Peptic ulcers were produced in conscious cats fasted for 18 hr prior to the experiment. By random assignment some cats were infused continuously intravenously with 0.15 M sodium chloride solution and the others with pentagastrin dissolved in 0.15 M sodium chloride solution. The infusions were maintained at a constant rate of 10 ml per hr for 24 hr by a peristaltic pump. Pentagastrin, freshly dissolved every 12 hr, was infused in a dose of 16 p.g per kg per hr for 24 hr. During the infusion the animals were deprived of food and water and observed for any side effects. At the end of experiments the cats were killed by rapid intravenous injection of sodium pentobarbitol. The stomach and the intestines were inspected in situ and then removed for gross and histological examinations. The ulcer area was measured planimetrically as previously described.! In cats with the duodenum excluded, the duodenal loop was perfused by 0.15 M sodium chloride solution or by 0.10 N hydrochloric acid solution at a constant rate of 20 ml per hr throughout the experiment. The differences in gastric acid secretion and in the ulcer area between the different groups of cats were determined statistically using Student's t-test for unpaired data. 4 Results Secretory Studies Diversion of bile and pancreatic juice to jejunum (A). In all 10 cats with diversion of bile and pancreatic juices to the upper jejunum, basal acid secretion was negligible. Maximal acid response to pentagastrin was achieved in all animals at the dose of 16 p.g per kg per hr (figs. 1 and 2).
April1969 PENTAGASTRIN-INDUCED PEPTIC ULCERS 705 ~ Hel OtTrPUT ~ ULCER AREA P< 0.15 A 8& c o FIG. 1. Schematic presentation of different surgical procedures. Maximal acid response to pentagastrin and mean ulcer area in cats after the diversion of bile and pancreatic juice to upper jejunum (A), after exclusion of the duodenum and its perfusion with saline (Bll or acid solution (B2), after excision of the duodenum (C), and after the diversion of bile and pancreatic juice to distal jejunum (D). In this and figure 2, the vertical bars indicate standard error of the mean. Asterisk around the points indicates the statistically significant difference in acid output or in ulcer area between stage A and others. 5. 0. 0 o---<l A ~--() Sf 0. 0 Bs 0-. _.-0 C P< 0.05 ---- 1. 0 --.-------------1'-------- --- 8ASAL ~. PENTAGASTRIN 1~ ------ ----- L ------ ---- - r I. FIG. 2. Basal acid output and the dose-response curve in cats with gastric fistula after the following stages of surgical procedure: A, diversion of bile and pancreatic juice to proximal jejunum; B1, exclusion and saline perfusion of duodenum; B2, acidification of the duodenum; C, excision of the duodenum. Each line is a mean of 10 experiments in 10 cats.
706 KONTUREK ET AL. Vol. 56, No.4 Exclusion and saline perfusion of the duodenum (BJ). After the exclusion of the duodenum and irrigation of the duodenal loop with saline, basal acid secretion was significantly increased, whereas the maximal acid response to pentagastrin was almost unchanged. It was attained in 7 cats at the dose of 8 ~g per kg per hr and in 3 remaining animals at the dose of 16 ~g per kg per hr. With further increment in the dose of stimulant, the mean acid output remained at the same level. No statistically significant differences were found in the maximal acid output before and after exclusion of duodenum (table 1). Exclusion and acid perfusion of the duodenum (B2). The introduction of 0.10 N HCI into the excluded duodenal loop throughout the secretory test resulted in basal achlorhydria in all animals. In comparison with stages A and Bl, the dose-response curve to pentagastrin was significantly lowered at almost all dose levels except at the highest one and the peak of the curve was shifted to the right. The highest acid output occurred at the dose of 32 ~g of pentagastrin per kg per hr. Excision of the duodenum (C). After this surgical procedure, basal acid output remained at a level similar to that following duodenal exclusion. In comparison with stages A and Bl, the dose-response curve to pentagastrin was significantly higher at the levels of 4 and 8 ~g of pentagastrin per kg per hr. The maximal acid output was achieved in 7 cats at the dose of 4 ~g of pentagastrin and in the remaining 3 animals at the dose of 8 ~g per kg per hr. Doubling these doses of pentagastrin did not increase the acid output. Diversion of bile and pancreatic juice to the distal jejunum (D). After this surgical procedure, basal acid output and the dose-response curve to pentagastrin were similar to those observed after the previous stage (C). Production of Peptic Ulcers Diversion of bile and pancreatic ju ce to the jejunum (A). Infusion of pentagastrin in a dose of 16 ~g per kg per hr for 24 hr in 10 cats with the duodenum left in the normal position and the biliary and pancreatic ducts transplanted to the upper jejunum resulted in the development of duodenal ulcers and multiple erosions in all animals (fig. 3). Mean ulcer area was 154.8 ± 41 mm 2 The duodenal mucosa was edematous and hyperemic. The oxyntic gland area was normal in appearance. The pyloric gland area showed patchy hemorrhages, multiple erosions, and superficial ulcers. Histologically, the duodenal mucosa revealed TABLE 1. Summary of results from groups A to D indicating maximal dose of pentagastrin, maximalt'es ponse to pentagastrin, area of ulcers, and the. percentage of cats with ulcers...-- ---.-...---.---~::~-' -1-~~:::"". I I Procedure Maximal dose of I Maximal acid response Ulcer a rea Percentage of cats with ulcers mm2 % A. Diversion of bile and pancreatic I juices to proximal jejunum. 16 (lo)a B1. Exclusion and saline perfusion of the duodenum... 8 (10) B2. Exclusion and acid perfusion of the duodenum....... 32 (10) C. Excision of the duodenum... 4 (10) D. Diversion of bile and pancreatic juice to distal jejunum... 4 (10) 2.45 ± 0.25 2.75 ± 0.27 1.80 ± 0.45 3.22 ± 0.28 3.17 ± 0.22 154 ± 41 (10) 100 135 ± 56 (8) 100 19 ± 8 (6) 25 125 ± 72 (8) 100 384 ± 94 (8) 100 a Numbers in parent heses indicate the number of a nimals used in secretory study or peptic ulcer production. b Values are means ± standard error of t he mean.
April1969 PENTAGASTRIN-INDUCED PEPTIC ULCERS 707 3 4 5 FIG. 3 (left). Stomach and duodenum of a cat with duodenum left in normal position and with diversion of bile and pancreatic juice to jejunum. Pentagastrin in this and subsequent figures was infused in a constant dose of 16 p.g per kg per hr for 24 hr. There are multiple erosions and ulcer in upper duodenum. FIG. 4 (middle). Stomach and jejunum of a cat with duodenum resected and with diversion of bile and pancreatic juice to upper jejunum. There is an ulcer situated near the gastrojejunal junction. Below the ulcer there is a duodenal transplant with the entrance of bile and pancreatic duct. FIG. 5 (right). Stomach and jejunum of a cat with duodenum resected and diversion of bile and pancreatic juice to distal jejunum. There are multiple erosions and deep hemorrhagic ulcers. diffuse edema with mild leucocytic infiltration of the mucosa. Gastric and duodenal mucosa in three cats infused with saline for 24 hr appeared normal. Exclusion and saline perfusion of the duodenum (B1). In 8 cats with the duodenum excluded and perfused by 0.15 M sodium chloride solution, pentagastrin infusion caused the development in the jejunum of all animals of deep elongated or rounded ulcers penetrating the muscularis mucosa and generally situated just beyond the gastrojejunal anastomosis. Mean ulcer area was 135 ± 56 mm 2 Two ulcers were associated with hemorrhages. The jejunal mucosa showed edema and superficial erosions. Histologically, a mild degree of leucocytic infiltration in the jejunal mucosa was noted. The oxyntic gland area was normal. In the pyloric gland area multiple erosions were found. Three cats infused intravenously with saline alone did not show any change in gastric or jejunal mucosae. No statistically significant differences were found in the ulcer area between the cats with duodenum left in the normal position and the animals with duodenum excluded. Exclusion and acid perfusion of the duodenum (B2). In 6 cats with the duodenum excluded and perfused with 0.10 N Hel, pentagastrin infusion produced
708 KONTUREK ET AL. Vol. 56. No.4 only edema, redness, and superficial erosions in the upper jejunal mucosa. Histologically, leucocytic infiltration of the mucosa and focal mucosal hemorrhages were observed. Two cats showed small single ulcers involving only the mucosa. The mean ulcer area for the entire group was 19 ± 8 mm 2 Two cats with acid perfusion of the duodenal loop infused intravenously with saline showed normal gastric and jejunal mucosae. The mucosae of the duodenal loops infused with acid solution showed edema, superficial erosions, and patchy hemorrhages situated mostly near the opening of the polyethylene tube in all ca,ts. Excision of the duodenum (C). All 8 duodenectomized cats with biliary and pancreatic ducts transplanted to the upper jejunum and infused with pentagastrin exhibited deep jejunal ulcers and multiple erosions confined to the region just beyond the gastrojejunal junction (fig. 4). Mean ulcer area was 125 ± 72 mm 2 The upper jejunal mucosa showed edema and redness. No changes in gastric or jejunal mucosae were found in 2 cats infused intravenously with saline alone. Diversion of bile and pancreatic juice to the distal jejunum (D). In all 8 cats with the duodenum removed and the biliary and pancreatic ducts transplanted to the region about 25 cm distal from the gastrojejunal junction, pentagastrin infusion resulted in the appearance of deep, mostly hemorrhagic ulcers extending about 15 cm into the proximal jejunum (fig. 5). The mean ulcer area was 384 ± 94 mm 2 Three ulcers perforated the jejunal wall, producing diffuse peritonitis. The jejunal mucosa showed diffuse edema and redness. Histologically, there was heavy leucocytic infiltration involving the mucosa and submucosa, dilation of mucosal capillaries, and focal mucosal hemorrhages. Some cats vomited blood-stained acid juice (PH 1). Two cats infused intravenously with saline alone showed only edema and redness of the jejunal mucosa. Discussion These studies indicate that the duodenum excluded from the digestive stream but remaining in the body as an isolated loop is capable, after acid perfusion, of markedly inhibiting the production of pentagastrin-induced peptic ulcers. Since secretion has previously been shown to inhibit gastric acid secretion in dogs 5 and is endogenously released by acidification of the duodenal mucosa,6 it is tempting to assume that this hormone could be responsible for the observed effects of acid perfusion of the duodenal loop on gastric secretion and peptic ulcer production in cats. However, our further study7 on the effect of pure secretin on gastric acid response to pentagastrin and on pentagastrin-induced ulcers showed that this hormone only slightly inhibits gastric acid secretion in gastric fistula cats but is able to prevent completely the formation of peptic ulcers in these animals. Thus, in contradistinction to the effect of exogenous secretin, the acid perfusion of the duodenum results in the powerful inhibition of gastric acid secretion as well as in almost complete prevention of peptic ulcer formation by pentagastrin infusion. However, a climbing shape of the doseresponse curve to pentagastrin in animals with the duodenum perfused with acid solution implies that if a larger dose of this stimulant were administered the acid response to pentagastrin and the rate of peptic ulcer formation could reach the level obtained in cats with duodenum left intact or with the duodenum excluded and perfused with saline. Since the determination of blood ph was not made, the occurrence of systemic acidosis resulting from the absorption of acid introduced into the duodenal loop during the 24-hr period of peptic ulcer production is not excluded. However, the prolonged instillation of acid solution by gastrostomy in dogs was demonstrated 8 to result in the production of gastric and duodenal ulcers in dogs when systemic acidosis was not prevented. Therefore, if acidosis following acid perfusion of the duodenal loop played any role in ulcer formation in our experiments, an increase in the appearance of peptic ulcers should be expected. Alternatively, the prevention of penta-
April 1969 PENTAGASTRIN-INDUCED PEPTIC ULCERS 709 gastrin-produced ulcers by secretin administration or duodenal acidification could be explained by an increase in bile and pancreatic juice inflow to the intestines,9 with resulting protection of intestinal mucosae against the action of highly acid gastric juice stimulated by pentagastrin. The significance of neutralization is evidenced in the present study by the higher degree of ulcer development in the jejunum after the bile and pancreatic juices were diverted from the gastrojejunal junction to the distal jejunum. Since this procedure was not followed by any appreciable changes in gastric acid response to pentagastrin, it is apparent that the higher production of peptic ulcers in this instance is mostly due to the lack of appropriate neutralization of the proximal jejunal mucosa. It is of interest that the duodenum when excluded from the alimentary tract, even without acid perfusion, also seems to exert some inhibitory influence on gastric acid secretion. This is suggested by the significant rise in basal acid output and the tendency to a shift to the left of the dose-response curve to pentagastrin after the formation of the isolated duodenal loop. However, since the escape of some acid to the intestines in cats with the duodenum left intact (stage A) was not excluded and the effect of gastrojejunostomy (stage B) on the adequacy of gastric collection was not established, the difference in the dose-response curve to pentagastrin between stages A and B should be cautiously interpreted. The lack of any significant difference in the ulcer area between the cats with duodenum left in the normal position and those with the duodenum excluded might be caused by the diversion of bile and pancreatic juices to the jejunum in the former instance and by the inflow of alkaline juices in the region of the gastrojejunal junction in the latter. Slightly higher gastric acid secretion caused by the exclusion of the duodenum would be thus more extensively neutralized, resulting in better protection of the jejunal mucosa against digestion by gastric secretion. Stafford and Schnaufer 1u and Westerheide et al. ll observed in dogs in which the intestinal loops containing the duodenum were excluded from the alimentary stream a significant increase in acid secretion and a high percentage of fatal gastrojejunal ulcerations. Westerheide et al. ll suggested that these ulcers and acid hypersecretion were caused by the release of gastric secretagogue from the excluded intestinal loops, rather than by the elimination of the duodenal inhibitory mechanism. In the light of our present study on the resection of the isolated duodenal loop, which was followed by a further increase in pentagastrin-induced gastric secretion, this suggestion should be more critically considered. We reached similar conclusions previously in a study on dogs 12 when removal of successive parts of the duodenum was followed by an increase of basal and gastrin-stimulated gastric acid secretion. Excision of the entire duodenum abolished the inhibitory effect of endogenous intestinal acidification, suggesting that the acid-sensitive inhibitory mechanism is confined to the duodenum. Our present study on the exclusion, acidification, and excision of the duodenum confirms a number of previous reports ll - 16 on the existence of a duodenal inhibitory mechanism and emphasizes the importance of bile and pancreatic juices in the formation of peptic ulcers induced by pentagastrin infusion. REFERENCES 1. Konturek, S. J., and J. Dubiel. 1968. Effect of diversion of bile and pancreatic juice on pentagastrin produced duodenal ulcers in cats. Scand. J. Gastroent. In press. 2. Thomas, J. E. 1941. An improved cannula for gastric and intestinal fistulas. Proc. Soc. Exp. Bioi. Med. 46: 260-261. 3. Konturek, S. J., and W. Krill. 1966. The influence of gastrin-related peptides on HCI secretion in man and in cats with gastric fistulas and Heidenhain pouches. Gastroenterologia 106: 281-294. 4. Snedecor, G. W. 1946. Statistical methods, Ed. 4. The Iowa State College Press, Ames, Iowa. 5. Wormsley, K. G., and M. I. Grossman. 1964. Inhibition of gastric secretion by secretin and by endogenous acid in the duodenum. Gastroenterology 47: 72-81.
710 KONTUREK ET AL. Vol. 56, No.4 6. Preshaw, R. M., A. R. Cooke, and M. I. Grossman. 1966. Quantitive aspects of response of canine pancreas to duodenal acidification. Amer. J. Physiol. 210: 629-634. 7. Konturek, S. J. 1968. The effect of secretin on gastric acid secretion and peptic ulcers induced by pentagastrin in cats with intact or resected duodenum. Amer. J. Dig. Dis. 13: 874-881. 8. Cummins, G. M., M. I. Grossman, and A. C. Ivy. 1948. An experimental study of the acid factor in ulceration of the gastrointestinal tract in dogs. Gastroenterology 10: 714-726. 9. Vagne, M., F. G. Stening, F. P. Brooks, and M. I. Grossman. 1968. Synthetic secretin: comparison with natural secretin for potency and spectrum of physiological actions. Gastroenterology 55: 260-267. 10. Stafford, E. S., and L. Schnaufer. 1964. An investigation in dogs of gastrojejunal ulcerations subsequent to a new method of duodenal stimulation. Ann. Surg. 159: 802-815. 11. Westerheide, R. L., D. W. Elliot, and J. M. Hardacre. 1965. The potential of the upper small bowel in regulating acid secretion. Surgery 58: 73-81. 12. Konturek, S. J., and M. I. Grossman. 1965. Localization of the inhibition of gastric secretion by acid in the intestine. Gastroenterology 49: 74-78. 13. Sircus, W. 1958. Studies on the mechanisms in the duodenum inhibiting gastric secretion. Quart J. Exp. Physiol. 43: 114-133. 14. Quintana, R., S. Kohatzu, E. R. Woodward, and L. R. Dragstedt. 1964. Mechanism of duodenal inhibition of gastric secretion. Arch. Surg. 89: 585-591. 15. Andersson, S. 1960. Inhibitory effect of hydrochloric acid in duodenum on gastrin-stimulated gastric acid secretion in Heidenhain pouch dogs. Acta Physiol. Scand. 50: 105-112. 16. Konturek, S. J., and M. I. Grossman. 1965. Effect of perfusion of intestinal loops with acid, fat, or dextrose on gastric secretion. Gastroenterology 49: 481-489.