Changes in Pancreatic and Intestinal Enzyme Activities following Chenodeoxycholic Acid Feeding M. D. O'DONNELL, 0. FITZGERALD & K. F. McGEENEY Dept. of Medicine and Therapeutics, University College, Dublin, Ireland O'Donnell, M. D.. Fitzgerald. 0. & McCeeney, K. F. Changes in pancreatic and intestinal enzyme activities following chenodeoxycholicacid feeding. Scund. J. GusScand J Gastroenterol Downloaded from informahealthcare.com by Nyu Medical Center on 01/06/15 For personal use only.

troent. 1977. 12, 683-687.

The effect of CDCA feeding on pancreatic and intestinal enzymes was studied. Mice were fed 0.5% w/w chenodeoxycholic acid in a normal diet. Pancreatic lipase concentration was significantly increased after 3 days on the CDCA diet, while amylase and trypsin concentrations were significantly higher at 23 days when compared with the controls. At 70 days there was a significant increase in the concentrations of amylase, trypsin, and lipase. Protein concentrations paralleled the rise in enzyme levels. Amylase and lipase, when measured as specific activities. were still higher than the controls at 70 days. Intestinal amylase levels did not change during the experiments, but intestinal rglucosidase activity increased significantly in the CDCA-treated animals. The results are discussed in terms of their similarity with those reported to occur after feeding soybean trypsin inhibitor. Key-words: Chenodeoxycholic acid diet: mouse; pancreatic and intestinal enzymes Mary D . O'Donnell, Ph.D., Dept. of Medicine and Therapeutics, University College, ' Woodview', Stillorgan Road, Dublin 4. Ireland

The effectiveness of chenodeoxycholic acid (CDCA) therapy in the dissolution of cholesterol gallstones has been recorded ( I , 4, 1 I , 23). Some concern has been expressed over the possible sideeffects of CDCA and its metabolites (5, 16, 22). CDCA is a primary dihydroxy bile acid which undergoes 7--CLdehydroxylation in the colon with the formation of lithocholic acid. We have demonstrated that CDCA and lithocholic acid are inhibitors of %-amylase (EC 3.2.1.1) in vitro (19). Since the inhibition of %-amylase by CDCA was abolished in the presence of excess albumin, it is probable that CDCA is not a specific inhibitor of 3 amylase but a compound that can bind to many enzymes and proteins. The ingestion of naturally occurring trypsin inhibitors by some rodent and avian species can cause enlargement of the pancreas with an increase in its digestive enzyme content (2, 3). Hypertrophy and hyperplasia of the rat pancreas have been demonstrated by Melmed et al. (16) by orally adminis-

tered soybean trypsin inhibitor. In the light ofthese reports and in view of the inhibitory effect of CDCA on a-amylase it seemed appropriate to study the effects of orally administered CDCA on enzyme levels of the pancreas and intestine.

MATERIALS AND METHODS Six-week-old male mice (inbred bca strain) weighing 1 4 - 18 g were purchased from Roebuck Farm. Welwyn, UK. They were randomly allocated into groups, each comprising 8 animals and kept in individual cages throughout the experiment. The animals were fed on laboratory animal food (ground pellets from Bolands Ltd., Dublin) for one week prior to the start of the experiment. Groups A, B, and C of eight mice each, were fed this diet containing added 0.5% CDCA (Weddel Pharmaceuticals, UK). Corresponding groups (A'. B', and C ')of eight animals served as controls and received the food without added CDCA. Groups A and A'

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M . D . O'Donnell. 0. FitzGerald & K . F. McGeeney

were killed three days after initiation of treatment. The paired groups B and B I , and C and C I were killed after 23 and 70 days respectively. The mice had free access to water and food and were weighed at the start of the experiment and then at 1&20 day intervals. Animals were killed by ether anaesthesia after a 16-18 hour fast. The pancreas was rapidly removed, rinsed in saline, and kept at 4 "C, blotted, weighed, and homogenized in 4ml of ice-cold 0.1 M NaCl containing 5 mM CaCI, using a Mickle homogeniser (Gomshall, Surrey, UK). A portion of upper intestine (- 12cm in length) was removed at a point 3-4cm distal to the pylorus in each animal. The intestine was cut open and rinsed (10 sec) in the above saline solution. Any adjoining tissue was removed prior to weighing and homogenization as above. All tissues were centrifuged (5000g. 4 "C. 15 min) and supernatants kept at -20°C prior to analysis. Tissue analyses Amylase activity was measured by the Phadebas blue starch method (Pharmacia, London) as modified (20). Trypsinogen was activated by enterokinase (Miles laboratories, Inc., Kankakee, III.)Trypsin was measured by its action on benzoylarginine-p-nitroanilide as substrate (6). Free fatty acids. released by lipase action on emulsified olive oil. were measured by Novak's method (18). Glucosidase activity was measured with maltose as substrate at pH 6.1 in the presence of 2mM EDTA as described (2 1). Protein concentration was determined according to the method of Lowry et al.

(14) using bovine albumin (Fraction V, Sigma Chemical Co., London. U K ) as standard. Histology. A small portion of both tissues (-50mg pancreas and -1cm intestine) from 6 animals of the C and C 1 group, was preserved in formal saline. Calculations. Results of enzymic analysis have been expressed as (a) activity per gram of tissue and (b) activity per milligram of protein. Data have been evaluated statistically by the use of Student's 't' test. RESULTS Weight gain during ingestion of CDCA.The control mice gained weight at a rate significantly higher than the CDCA-fed mice. as shown in Fig. 1. Protein content. The concentration of protein per gram pancreas and intestine of the animals fed control and C D C A diets is shown in Table I. The CDCA-treated animals showed a significant increase in pancreatic protein concentration after 3 days (p < 0.005) and 70 days (p < 0.0 1). At day 23 the difference was not significant (0.05 < p < 0.1). The intestinal protein concentration was significantly higher in the CDCA-fed animals after 70 days (p < 0.05). N o significant difference was shown after 3 or 23 days (Table I).

Table. 1. Effect of CDCA and a control diet on pancreatic and intestinal protein levels in the mouse Protein concentration (mg/g)

z C

CONTROLS

601

D A Y S ON DIET

Fig. 1. Change in weight of mice, measured as percentage of their initial weights, during ingestion of CDCA. Weights of the control mice were significantly higher after 10 days (p < 0.0005), 3 0 days (p < 0.0005). 50 days (p

Changes in pancreatic and intestinal enzyme activities following chenodeoxycholic acid feeding.

Changes in Pancreatic and Intestinal Enzyme Activities following Chenodeoxycholic Acid Feeding M. D. O'DONNELL, 0. FITZGERALD & K. F. McGEENEY Dept. o...
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