ACETYLSALICYLIC ACID AND IONIC FLUXES ACROSS THE GASTRIC MUCOSA OF MAN

GASTROENTEROLOGY Copyright 1968 by The Williams & Wilkins Co. Vol. 54, No.4, Part 1 of 2 Parts Printed in U.S.A. ACETYLSALICYLIC ACID AND IONIC FLUXES ACROSS THE GASTRIC MUCOSA OF MAN BERGEIN F. OVERHOLT, M.S., M.D., AND H. MARVIN POLLARD, M.D. Section of Gastroenterology, Department of Internal Medicine, University Hospital, University of Michigan, Ann Arbor, Michigan In his studies of the gastric mucosal barrier of the separated canine fundic pouch, Davenport has shown that acetylsalicylic acid damages the gastric mucosa. Mucosal injury was reflected by an increased loss of hydrogen ion from the luminal content together with increases in volume, sodium and potassium ion content, and occasionally with bleeding into the pouch.!. 2 The present study is an attempt to measure the effect of acid and acetylsalicylic acid on the human gastric mucosa. Increases in volume and net movement of sodium, potassium, and chloride ions into the lumen were found following irrigation of the stomach with aspirin. Methods Eleven subjects including 2 females and ranging from 22 to 56 years of age were studied. Seven subjects were asymptomatic and had normal acid output (11.4 to 29.6 meq per hr) following betazole hydrochloride stimulation, 1.5 mg per kg. One subject (D. S.) was in excellent health although he was markedly hypochlorhydric following betazole stimulation (0.24 meq per hr). Another subject (S. C., 36.0 meq per hr) had symptomatic and radiological evidence of a duodenal ulcer. One patient (J. H.) had gastroscopic evidence of gastric atrophy, while another (A. R., 18.2 meq per hr) had antral gastritis. The subjects were fasted 12 hr before the tests. A no. 16 Levin type tube was passed into the stomach and adjusted in position so Received June 30, 1967. Accepted December 13, 1967. Address requests for reprints to: Dr. Bergein F. Overholt, 525 E. 68th Street, New York, New York 10021. The authors are indebted to Horace W. Davenport for continuing interest and Miss Susan Heyliger for technical assistance. 538 that "free flow" was obtained; free flow prevailed when the stomach was rinsed with 50 ml of distilled water and 50 ml were easily returned. The tube was initially placed with fluoroscopy but the free flow adjustment allowed more complete collection of the instilled water. The location of the tube was then along the greater curvature 1 to 2 inches proximal to the antrum while the patient maintained the left lateral decubitus position. Saliva was expectorated during all studies. Following two 15-min basal collection periods, 110 ml of an acid solution (100 mm HCl, 15 mm N acl, 78 mm mannitol, Cr 51 C1 3 25 flc per liter) were instilled into the stomach. Ten milliliters were immediately withdrawn after gentle mixing and saved for analysis. At the end of 15 min the gastric contents were aspirated and the stomach was rinsed with 50 ml of distilled water. The rinse was collected and saved for Cr 51 counting to determine residual volumes. This procedure was then repeated for a total of four I5-min periods. On a separate day, usually following the acid instillation study, a second test was initiated as in the first study. During the second, third, and fourth periods, however, acetylsalicylic acid was added to the acid solution (20 mm acetylsalicylic acid, 100 mm HCI, 15 mm NaCl, 78 mm mannitol, Cr 51 C1 3 25 flc per liter). The acetylsalicylic acid was supplied by Bristol-Meyers Company, New York, N. Y. All samples were measured for volume to the nearest 0.1 ml and saved for estimations of sodium, potassium, chloride, and hydrogen ion concentrations, salicylate levels if appropriate, and Cr 51 counting. Gastric secretion and emptying were determined by the method of Hunt; using Cr 51 C1 3 as a nonabsorbable marker." 5 Ion fluxes (formula 1) were determined by taking the milliequivalent of each ion in the stomach at 15 min (meq,) plus the amount emptied

April 1968 GASTRIC MUCOSA OF MAN 539 (meqe) and subtracting the milliequivalent at the start (meqo). Xet ion flux = meqf + meq. - meqo (1) A positive flux indicates a net addition of ions into the lumen. A negative flux indicates the diffusion of an ion back into the mucosa. The ion fluxes during the first infusion period of each test were compared. Only if the hydrogen ion fluxes during the first period of the 2 procedures differed by less than 0.7 meq were the tests included in the study. This standard was established to exclude differences in mucosal permeability that were not related to salicylate infusion. The results of the last three infusion periods were used for analysis. Hydrogen ion content was determined by titration to ph 7.4 with 0.1 N NaOH using a glass electrode. Since acetylsalicylic acid was included in the hydrogen ion titrations in the aspirin studies, the concentration of the acetylsalicylic acid as separately determined was subtracted from the appropriate titration values. Sodium and potassium were determined with the internal standard flame photometer, chloride with the Aminco-Cotlove titrator, and Cr 51 by counting in a Nuclear-Chicago Autogamma. Salicylate levels were determined by the method of N atelson 6 and fluxes were calculated on a millimolar basis using formula 1. Statistical analysis was carried out using the t-test for the difference between the means of paired observations. 7 Ionic fluxes and volume changes for the first I5-min periods and also the final 45-min test periods were compared separately. Results The net movement of ions and the volumes secreted and emptied during the first I5-min period and during the 45-min periods of acid or acid-aspirin instillations are presented in tables 1 and 2. The data from 9 subjects are presented. Two patients were excluded as t he hydrogen ion fluxes during the first period did not fall within the limits previously defined. During the first infusion period of each of the two tests, no significant differences in the fluxes of hydrogen, sodium, potassium, or chloride ions or in the volume secreted were observed. There was, however, a significant and unexplained increase in the volume emptied through the pylorus during this period of the aspirinacid study ( P < 0.05). During the periods of aspirin-acid irrigation there were significant increases in the fluxes of sodium (P < 0.01), potassium TABLE 1. The results of instillation into the stomach of 100 ml of an acid solution" during the fi rst 15-min period of separate instillation studies (see text for details)b Subject and diagnosis H+ Na+ K+ CI- ASA ASA ASA Control! ASA.Control! Control I Control I Volume emptied through pylorus Volume "secreted" meq/15 min meq/ 15 min meq/15 min meq/15 min ml/15 min >n1/15 min G. F., normal. 2.92 3. 15 0.73 1.03 0.24 0.33 3.78 4.30 19.2 I 34.7 28.5 32.5 D. Ii., normal... 1.15 0.77 1.78 1. 72 0.62 0.48 3.24 2.70 29.0 50.9 38.7 37.1 J. W., normal. 0.91 0.21 1.73 2.10 0. 36 0.42 3. 01 1.79 6.0 12.2 26.7 30. 3 T. F., normal. 1.87 1. 36 4.74 2.47 0.48 0.42 7.55 4.60 35.2 27.4 61.6 36.3 E. U.,., normal. 1.89 2.52 0.32 2.48 0.11 0.32 2.24 5.42 10.5 43.0 15.9 50.6 J. H., gastric atrophy....., 0.91 0.64 1.89 1.91 0.26 0.07 2.97 2.45 33.1 29.3 29.3 33. 2 D. S., hypochlorhydria..... -0.71-0.61 1.02 1.59 0.15 0.38 0.65 1.59 7.0 24.0 14.2 26.0 S. C., duodenal ulcer......... 1. 29 1. 37 2.68 2.38 0.37 0.32 4.22 4.03 8.1 15.2 37.3 34. 5 A. Ii.,. antral gastritis...... 1.04 1.14 1.23 1.51 0.16 0.28 2.66 2.48 29.4 32. 1 19.2 23.7 a Solution contains 100 mm HCI, 15 mm NaCI, 78 mm mannitol, Cr 5l CI, 25 J.lC per liter. b Electrolytes are expressed as positive (diffusion into the lumen) or negative (diffusion into the mucosa) net movement during the 15-min period. c Female.

540 OVERHOLT AND POLLARD Vol. 54, No.4, Part 1 TABLE 2. The results of instillation into the stomach of 100 ml of a control acid solution" and an acetylsalicylic acid solution b during three 15-min periods on the same subjects meq/15 min meq/45 min meq/45 min meq/45 min meq/45 min ml/45 min ml/45 min H+ Na+ K+ Cl- Subject and Basal secre- Volume emptied Volume diagnosis tion through pylorus "secreted" Control! ASA Con-! ASA con-! ASA trol trol Control! ASA Absorption of acetylsalicylic acid --- mmoles/45 min G. F., nor- I mal.. 2.73 1.36 5.62 6.15 2.18 2.58 0.57 0.71 8.01 9.26 130.2 165.1 49.9 71.4 1.65 D. R., normal.... 1.13 0.67 0.53 1.34 4.71 7.16 1. 09 1.52 5.46 11.21 134.2 165.3 85.7 139.2 0.29 J. W., normal.... 0.34 0.10 1.47 1. 75 3.97 6.38 0.72 1.07 6.37 9.23 17.7 65.6 66.2 113.3 1.17 T. F., normal.... 1.38 0.89 2.25 1.58 7.14 8.11 0.68 0.95 11.02 11.15 98.7 67.7 93.4 105.5 2.97 E. U. d, normal.... 0.31 1.26 3.59 4.74 2.21 5.63 0.35 0.79 5.13 10.00 67.5 84.2 52.9 109.8 1.44 J. H., gastric atrophy... 1.63 0.70-0.78-0.51 4.88 7.00 0.62 0.31 5.65 6.22 90.7 73.0 56.4 80.3 2.34 D. S., hypochlorhydria. 0.00 0.20-2.09-2.12 4.44 2.99 0.39 0.88 2.68 2.90 65.1 16.3 68.1 67.8 1.26 S. C., duodenal ulcer.... 1.67 1.49 2.94 3.48 6.94 10.16 0.66 1. 64 9.64 16.36 17.6 100.8. 87.2 144.9 0.72 A. R. d, antral gastritis.. 1.02 0.51 0.45-0.22 3.86 4.10 0.26 0.33 5.17 5.17 119.2 119.6 45.7 55.1 1.15 Means.... 1.32 0.80 1.55 1.80 4.48 6.01 0.59 0.91 6.57 9.06 82.3 95.3 67.3 98.6 1.44 ± SD..... 0.74 0.50 2.35 2.63 1. 75 2.47 0.03 0.46 2.56 3.94 44.1 48.6 17.8 31.9 0.76 a Solution contains 100 mm HCl, 15 mm NaCl, 78 IDM mannitol, CrSlCb25!,c per liter. b Solution contains 20 IDM acetylsalicylic acid, 100 mm HCl, 15 mm NaCl, 78 IDM mannitol, Cr"C!' 25!,c per liter. C Electrolytes are expressed as positive (diffusion into the lumen) or negative (diffusion into the mucosa) net movement during the 45-min period. Volumes are expressed as milliliters per 45-min period and acetylsalicylic acid absorbed as millimoles per 45 min. dfemale. (P < 0.05), and chloride (P < 0.05) ions, but no significant change in hydrogen ion flux was detected. In the different subjects studied, the degree of mucosal barrier changes reflected by the increase in ion flux was not related to the amount of aspirin absorbed or to the amount of acid backdiffusion measured during the control period. A significant increase in volume (P < 0.01) was observed during aspirin instillations. There were no differences in gastric emptying during the two tests and there was no bleeding into the gastric lumen. Discussion Whether aspirin is deleterious to the gastrointestinal tract-particularly the stomach-has been debated in the literature for years. Davenport 1, 2 studied the effect of salicylates on the mucosa of the canine fundic pouch and showed that gastric mucosal damage by this and other organic acids is accompanied by a loss of hydrogen ion from the gastric lumen and an increase in the volume and in the sodium and potassium content in the lumen with occasional bleeding. These changes reflect an altered permeability of the mucosa. Davenport's studies indicate that, although aspirin alone may damage the canine gastric mucosa, this damage is accentuated by hydrochloric acid in proportion to the acid concentration. 2 In the present study, a physiological concentration of Hel (100 mm) and an aspirin concentration of 20 mm, equivalent to 2 aspirin tablets dissolved in 180 ml of gastric juice, was tested on the human stomach. The validity of the present study depends on the use of a nonabsorbable reference marker and on the reliable recovery of gas-

April 1968 GASTRIC MUCOSA OF MAN 541 tric contents. In our laboratory the immediate removal of the instilled acid Cr 51 Cl 3 solution from the stomach provided 99.4% recovery if one rinse was included. "When an aspirin-acid solution containing Cr 51 Cl 3 was instilled into the stomach of a pylorusligated dog, 97.8% of the Cr 51 was recovered from the gastric lumen at the end of 30 min when one 50-ml wash was included. This result corresponds to those of Bloom et a1. 5 who performed recovery experiments using a Cr 51 Clg-acid solution in Heidenhain pouch dogs. In the present study, significant luminal increases of sodium, potassium, and chloride ions were detected during the instillation of the aspirin-acid solution. The sodium and potassium changes reflect an increased permeability of the barrier to ion movement and are perhaps associated with damage to the mucosa. 1, 2 The chloride increase could be associated with the movement of sodium and potassium ions that occurs during damage or it could be associated with the increase in secretion that might accompany damage to the stomach. 8 In contrast to Davenport's studies on the dog, a significant increase in hydrogen ion back diffusion was not detected during aspirin instillations. Even in subjects J. H. and D. S., who had back diffusion during their control test and should therefore be more susceptible to acid injury to the stomach,9 increased acid back diffusion during the aspirin-acid test was not found. However, small changes in hydrogen ion absorption may not have been detected either because of the mathematical limitations of Hunt's technique for measuring gastric secretion and emptying or because of the possible increase in acid secretion stimulated by the release of mast cell histamine into the interstitial fluid during salicylate injury.lo The volumes found in the stomach during the aspirin-acid instillations were significantly increased over the control acid test. Whether a modified ultrafiltrate of plasma or an increase in secretion that might occur during mucosal damages accounts for the fluid increase following aspirin irrigation of the human stomach has not been determined. There was no correlation between the amount of aspirin absorbed and abnormal ion movement or volume increases. Davenport's criteria of injury to the gastric mucosa have in part been met by this study. We have shown that a volume of fluid containing sodium, potassium, and chloride passes into the gastric lumen during instillation of acid solutions and to a significantly greater extent during aspirinacid solutions. We were unable to detect back diffusion of hydrogen ions during the aspirin-acid irrigation. Summary The effect of a 100 mm HCI solution on the movement of hydrogen, sodium, potassium, and chloride ions across the human gastric mucosa and on volumes s e ~ creted by the stomach and emptied through the pylorus was determined and compared with a 100 mm HCI-20 mm acetylsalicylic acid solution. The results of 45-min irrigations on 9 subjects are reported. During irrigation with the aspirin-acid solution, movement of sodium, potassium, and chloride ions into the lumen was significantly increased, but significant changes in hydrogen ion movement were not detected. Volumes recovered from the stomach during the aspirin study were significantly increased. Neither the presence of a pre-existing abnormal mucosal barrier permeability nor the quantity of aspirin absorbed had any apparent effect on ion movement. ';V e conclude that, under the conditions of this study, irrigation of the human stomach with the aspirin-acid solution lowered the barrier of the gastric mucosa to ion movement but did not produce unequivocal evidence of chemical damage to the mucosa. REFERENCES 1. Davenport, H. W. 1964. Gastric mucosal injury by fatty and acetylsalicylic acids. Gas troenterology 46: 245-253. 2. Davenport, H. W. 1965. Damage to the gastric mucosa: effect of salicyla tes and stimulation. Gastroenterology 49: 189-196. 3. Hunt, J. N. 1959. Gastric emptying and secretion in man. Physiol. Rev. 39: 491-533.

542 OVERHOLT AND POLLARD Vol. 54, No.4, Part 1 4. Donaldson, R. M., Jr., and R. F. Barreras. 1966. Intestinal absorption of trace quantities of chromium. J. Lab. Clin. Med. 68: 484-493. 5. Bloom, D. S., E. D. Jacobson, and M. I. Grossman. 1967. Validation of dilution indicators in the stomach. Gastroenterology 52: 205-210. 6. Natelson, S. 1957. Microtechniques of clinical chemistry for the routine laboratory, pp. 332-334. Charles C Thomas, Publisher, Springfield, Ill. 7. Steel, R. G. D., and J. H. Torrie. 1960. Principles and procedures of statistics. Mc Graw-Hill Book Company, Inc., New York. 8. Davenport, H. W. 1966. Fluid produced by the gastric mucosa during damage by acetic and salicylic acids. Gastroenterology 50: 487-499. 9. Overholt, B. F., and H. M. Pollard. 1968. Acid diffusion into the human gastric mucosa. Gastroenterology 54: 182-189. 10. Johnson, L. R., and B. F. Overholt. 1967. Release of histamine into gastric venous blood following injury by acetic or salicylic acid. Gastroenterology 52: 505-509.