Activity distribution of seven digestive enzymes along small intestine in calves during development and weaning

Ten groups of calves were used to study the changes in activity levels and distribution of seven hydrolases in the intestinal mucosa during development and weaning. The calves in the first group were sacrificed at birth while those in the remaining nine groups were either milk-fed until slaughter on days 2, 7, 28, 56, 70, and 119; or weaned between days 28 and 56 and then slaughtered on days 56, 70, and 119, respectively. The small intestine was immediately cut off and divided into five segments, ie, duodenum, proximal jejunum, median jejunum, distal jejunum, and ileum. In the milk-fed animals, the activity levels of aminopeptidases A and N, alkaline phosphatase, lactase, and isomaltase were maximum at 2 days of age, and then declined sharply between days 2 and 7 but did not change significantly thereafter. By contrast, the maltase activity increased between days 7 and 119, while no sucrase activity was detected. Weaning resulted in a decrease in the activity of lactase and an increase in that of aminopeptidase N, maltase, and isomaltase. The distribution of all these enzymes along the small intestine was slightly influenced by age but not at all by weaning.     

Intestinal disaccharidase activity following pancreatic duct occlusion in the rat

The influence of pancreatic secretions on growth and brush-border enzyme activity, throughout the entire small intestine, was examined in the rat. Pancreatic secretions were excluded from the gut lumen by stapling the pancreatic ducts, without interruption of bile flow. The entire small intestine was studied as four segments; the duodenum and three distal segments of equal length. Weight of intestine and mucosa, and mucosal sucrase, isomaltase, lactase, and alkaline phosphatase activity were measured 10-15 days following pancreatic duct occlusion, or sham-operation. The duodenum of pancreatic duct-occluded animals exhibited significant hypertrophy. In general, specific and total disaccharidase activities were greater in duct-occluded animals than in controls throughout the intestine. The increase was more pronounced in distal than in proximal segments. The sucrase/isomaltase ratio was significantly greater in pancreatic duct-occluded animals than in controls in the two distal segments. Alkaline phosphatase activity was not affected by pancreatic duct occlusion. The greater relative increase of disaccharidase activities and sucrase/isomaltase activity ratios in the distal segments of duct-occluded animals, indicates a more important regulatory role of pancreatic enzymes in the distal small intestine. It is concluded that regulation of intestinal brush-border enzyme activity by pancreatic secretion is selective for enzyme and site as follows: disaccharidases, but not alkaline phosphatase, are regulated; the sucrase subunit of the sucrase/isomaltase complex is most sensitive to regulation, while lactase is least sensitive; and the regulatory effect on disaccharidases is greater in distal than in proximal intestine.     

Effects of hydrocortisone and pentagastrin on the activity of intestinal disaccharidases and alkaline phosphatase in weanling rats

Effect of bilateral adrenalectomy and subsequent injections of hydrocortisone and pentagastrin on the activity of different intestinal digestive enzymes were measured in 20-day-old rats. Eleven days after adrenalectomy the activity of lactase, sucrase, and alkaline phosphatase, but not maltase, was significantly decreased when compared with sham-operated rats. In adrenalectomized rats, repeated injections of hydrocortisone (50 mg/kg) significantly increased the activity of lactase, sucrase, maltase, and alkaline phosphatase by 15%, 49%, 32% and 121%, respectively, over the corresponding adrenalectomized control. Pentagastrin (500 microgram/kg) injections to adrenalectomized rats produced significant 41% and 58% increments in lactase and alkaline phosphatase activities, compared with the adrenalectomized control. Sucrase activity was unaffected by pentagastrin, but maltase showed a non-significant 34% higher activity than in the adrenalectomized control. Adrenalectomy by itself lowered the Km and Vmax of alkaline phosphatase by 33% and 66%, respectively, which were increased to 95% and 70% of the corresponding sham-operated level by either hydrocortisone or pentagastrin treatment. When intestinal homogenates from salinetreated adrenalectomized rats were mixed in equal proportion with homogenates from sham-operated or hydrocortisone- or pentagastrin-treated animals, Km values for alkaline phosphatase were found to be similar to those observed for sham-operated or hormone-treated groups alone. However, in the same mixed preparations Vmax values were found to be additive.     

Small-intestinal and colonic changes after biliopancreatic bypass for morbid obesity

Morphologic and functional adaptations of the functioning intestine were evaluated in 41 patients before and after biliopancreatic bypass for morbid obesity. This surgical procedure diverts pancreatobiliary secretions via the duodenum and the jejunum into the colon, the remaining small intestine being anastomosed to the stomach after antrectomy. In the proximal ileum there was an 80% increase of the height of villi; the specific activities of maltase, sucrase, and aminopeptidase in brush border membranes remained unaffected, and that of lactase tended to decrease. In the distal ileum villi heights increased only by 58%, and disaccharidase activities (except for maltase) were slightly enhanced. In the colon the mucosa displayed, in some patients, focal appearance of true villi, and brush border enzyme activities increased concomitantly. We conclude that biliopancreatic bypass induces an adaptation of all intestinal segments of the functioning intestine; this adaptation tends to compensate for the shortening of the gut continuity.     

Consequences of adrenalectomy on small intestine trophic parameters in aged and young rats: evidence of defective adaptation by aging and lack of corticoids

Previous study pointed to an important role of adrenals and glucocorticoids in the trophic status of the adult small intestine mucosa, with possible implications during stress events. Small intestine morphological and biochemical consequences of 10-day bilateral adrenalectomy and also sham-related laparotomy were determined in 23-month-old Sprague-Dawley rats. As described in young rats, adrenalectomy in old rats leads to partial atrophy and disorganization of the proximal small intestine epithelium, with an increase in the number of Paneth cells and reduced crypt cell proliferation. We also observed a decrease of goblet cell number and a reduction of all enzyme activities including disaccharidases, in contrast with the specific induced response shown in young rats. A number of marked biochemical effects have also been noted in aged rats subjected to solely laparotomy, suggesting age-related adaptation impairments. In conclusion, adrenalectomy modified the differentiation processes of the small intestinal mucosa in both young and aged rats, and some parameters underlined that the lack of corticoid-mediated adaptive process are exacerbated by cumulative surgical stress (event) and aging.     

Thyroid hormone differentially regulates rat intestinal brush border enzyme gene expression.

Thyroid hormone [triiodothyronine (T3)] has been shown to play a critical role in the growth and maturation of the mammalian small intestine, but its mechanism of action has not been well studied. In the current study, an animal model of hypothyroidism and hyperthyroidism was used to study the effects of T3 on the small intestine. Adult rats were treated with propylthiouracil for a 6-week period and then given injections of either saline (hypothyroid) or 30 micrograms/100 g body wt of T3 (hyperthyroid). Northern blot analyses showed marked differential regulation of brush border enzyme gene expression. Lactase messenger RNA (mRNA) levels decreased approximately 75% along the length of the small intestine, whereas sucrase levels were unchanged. The intestinal alkaline phosphatase mRNA species were upregulated by T3, especially the 3-kilobase band, which increased most dramatically in jejunum. Further experiments showed significant levels of both the alpha-1 and beta-1 T3 receptor mRNAs within the small intestinal mucosa. Histological examination showed that T3 treatment causes marked villus hyperplasia throughout the length of the small intestine. These results provide insight into the mechanism by which T3 exerts its influence on the growth and differentiation of the intestinal epithelium.     

Effect of Chronic Ethanol Consumption on the Activities of Residual Small Bowel Brush-Border Enzymes after Proximal Jejunum Resection in the Rat

Ethanol consumption has a toxic effect on the epithelium of the small bowel, but enterocyte maturity is very difficult to measure under these circumstances. However, when ethanol intake is combined with enterectomy, enterocyte immaturity is greater, permitting an easier separation of these two effects. In a group of rats (13 male Wistar rats weighing ∼220 g) fed a liquid diet containing 35% ethanol for 4 weeks after resection of the proximal jejunum, the residual small intestine brush border maltase, sucrase, and lactase activities were similar to those of a pair-fed control group (13 animals). However, alkaline phosphatase activity was decreased in the mucosa and in the enterocyte brush border, probably because of the lower activity of this enzyme in the jejunum-ileum remnant of the alcoholic group.     

Increase of intestinal brush border hydrolases in mucosa of streptozotocin-diabetic rats

Summary Experimental diabetes mellitus in rats was induced by streptozotocin. Five days after administration of streptozotocin intestinal brush border hydrolases (maltase, sucrase, trehalase, lactase) and alkaline phosphatase were markedly elevated at all levels of the small intestine as measured in the total homogenate and in the isolated brush border preparation. Insulin treatment beginning 15 h after administration of streptozotocin was able to decrease the increased disaccharidase activity due to streptozotocin diabetes. In experimental diabetes mellitus of rats trransport as well as digestive functions of the intestinal mucosa are stimulated.This work has been presented at the meeting of the European Society for Clinical Investigation, Scheveningen, The Netherlands, April 1972 and has appeared in abstract form (1a)     

Hepatotoxic Agent Thioacetamide Induces Biochemical and Histological Alterations in Rat Small Intestine

We have assessed the effect of the oral ingestion of thioacetamide on small intestine structure and function. Thioacetamide-treated rats showed diminished mucosa weight; protein, DNA, and RNA content; and leucine aminopeptidase activity as compared to controls in both jejunum and ileum. In the jejunum, there was a reduction in the activities of alkaline phosphatase, ATPase, glucose-6-phosphatase, and myeloperoxidase, whereas in the ileum, maltase, lactase, and -glutamyltranspeptidase were reduced. In both jejunum and ileum we found enlarged intercellular spaces, dark epithelial enterocytes, and lymphocyte infiltration. Enterocytes showed lobulated nuclei, deranged mitochondria with loss of their cristae, dilated rough endoplasmic reticulum containing dense material, and vesiculation of the smooth endoplasmic reticulum and the Golgi apparatus. Smooth muscle cells of the intestine exhibited ultrastructural alterations. These findings indicate that chronic oral intake of thioacetamide mimics not only hepatic alterations but also small intestine alterations normally associated with human cirrhosis.     

Combined impact of mucosal damage and of cystic fibrosis on the small intestinal brush border enzyme activities

In 61 cystic fibrosis (CF) patients, the small intestinal mucosa was studied at the time of diagnosis before starting therapy. In 19 out of 61 patients, partial villous atrophy on light microscopy and shortened villi on stereomicroscopic examination were seen. On the biopsy specimens, maltase, sucrase, lactase and alkaline phosphatase activities were studied. Comparison of the enzymatic activities in CF patients having damaged mucosa and a group of patients having similar mucosal lesions of unspecified origin (UTID), reveals a significantly more pronounced decrease of the alkaline phosphatase activity (p < 0.005) in the CF patients. This is in agreement with previous reported results in CF patients with normal mucosa. The abnormal mucosal findings could be due to the decreased neutralization of the gastric content delivered into the duodenum, the early inflammatory reaction present in the CF mucosa and/or to the impaired synthesis of membrane glycoproteins and enzymes secondary to the CFTR mutation.     

Inhibitory effect of somatostatin on cholera toxin-induced diarrhea and glycoenzyme secretion in rat intestine

The effects of somatostatin on cholera toxin-induced secretory diarrhea and the appearance of glycoenzymes in the intestinal lumen and intestinal lymph were investigated in rat small intestine. After exposure to cholera toxin, marked fluid accumulation in the small intestinal tract and elevation of the jejunal mucosal cyclic AMP (cAMP) concentration were observed. The activity of alkaline phosphatase, aminopeptidase and sucrase increased in the intestinal lumen after toxin exposure. In intestinal lymph, alkaline phosphatase activity was increased after cholera toxin administration, while aminopeptidase activity remained unchanged. Somatostatin suppressed cholera toxin-induced secretory diarrhea, but it did not affect the elevated mucosal cAMP concentration. This peptide also inhibited the appearance of glycoenzymes in the intestinal lumen and lymph induced by cholera toxin administration. These results suggest that somatostatin exerts its inhibitory effects on cholera toxin-induced secretory diarrhea and on the appearance of glycoenzymes in the intestinal lumen and lymph by affecting processes beyond cAMP formation.     

Effect of various doses of medroxyprogesterone acetate on intestinal functions in rats

The dose relationship between medroxyprogesterone acetate (MPA), a long acting contraceptive, and rat intestinal digestive and absorptive functions has been investigated. The study revealed that the activities of brush border sucrase, lactase and leucine aminopeptidase were stimulated only at high doses, viz 70 mg/kg (180 mumol/kg) body weight and above, whereas the activity of alkaline phosphate was depressed at comparatively low dose (17.5 mg/kg; 45 mumol/kg body weight). This decrease was found to be significant (p less than 0.001) at all the doses tested. The inhibition in the intestinal uptake of calcium paralleled the decrease in alkaline phosphatase activity. Relatively high amount of MPA (140 mg/kg; 360 mumol/kg) was required to augment the uptake of glucose and amino acid. The results obtained do not indicate a close relationship between the dose of the drug and the extent of alteration in the rat intestinal digestive and absorptive functions. The study appears to confirm the association between brush border enzymes activities and uptake of nutrients in rat intestine.     

Decreased lysophospholipase and increased phospholipase A2 activity in ileal mucosa from patients with Crohn's disease

Lysophospholipase (EC 3.1.1.5) and phospholipase A2 (EC 3.1.1.4) were determined in ileal mucosa from patients with Crohn's disease (CD) and non-inflammatory bowel diseases ( NIBD ). In addition, the activities of alkaline phosphatase, sucrase, maltase, and lactase were determined. The lysophospholipase activity, like that of alkaline phosphatase, sucrase and maltase, was decreased in affected areas of CD, whereas the phospholipase A2 activity was rather increased. Lysophospholipase and phospholipase A2 activities in apparently unaffected mucosa from CD patients were in between those in healthy mucosa from NIBD patients and those in affected mucosa from CD patients. These findings point to the possibility that the mucosal activity of lysophospholipase, like that of other brush border enzymes, is decreased in CD. This may render the mucosa less capable to handle lysolecithin, a potentially harmful agent formed in the intestine and known to induce inflammation in a number of experimental systems.     

Explant culture of human fetal small intestine

Human fetal intestine (10-14 wk gestation) has been cultured as explants in a serum-free Leibovitz L-15 medium for periods up to 9 days. As determined by light microscopy, the overall architecture of the intestinal explant was maintained throughout the culture period. At the ultrastructural level the villus absorptive cells remained tall with well-defined brush border, apical tubular system, and supranuclear and infranuclear accumulations of glycogen. All other epithelial cell types were also preserved. The incorporation of [3H]thymidine and [3H]leucine continued during the culture period, reflecting a sustained synthesis of deoxyribonucleic acid and proteins. The hydrolytic activities of the brush border membrane were established based on data obtained throughout the course of the culture of a large number of intestinal specimens. Sucrase, maltase, glucoamylase, trehalase, lactase, alkaline phosphatase, and gamma-glutamyl transpeptidase activities increased during the 9 days of culture even though different patterns were recorded. These observations clearly established that human fetal small intestine can be maintained in organ culture for at least 9 days in a serum-free medium.     

Exogenous leptin controls the development of the small intestine in neonatal piglets

Leptin, a hormone produced and secreted by adipose tIssue, muscles and stomach, is involved in the regulation of adipose tIssue mass, food intake and body weight in neonatal animals. It is also produced in the mammary glands and secreted into the colostrum and milk. Since leptin receptors are widely distributed in the small intestine mucosa, the aim of the present study was to investigate the effect of exogenous leptin on the development of the small intestine in neonatal piglets. Male neonatal piglets were fed with sow's milk or artificial milk formula. Every 8 h the latter received either vehicle or leptin (2 or 10 microg/kg body weight). The animals were either killed after 6 days of treatment and the small intestine sampled for histology and brush border enzyme activities or were tested for marker molecule (Na-fluorescein and BSA) absorption in vivo. Feeding milk formula slowed the maturation of small intestinal mucosa compared with feeding sow's milk. However, after leptin treatment the length of the small intestine was increased, and intestinal villi length, but not crypt size, was reduced compared with controls. The mitotic index was increased and the percentage of vacuolated enterocytes was reduced in the entire small intestine. Enterocyte brush border protease and lactase activities were reduced in the jejunum. Na-fluorescein marker molecule absorption did not change but that of BSA was reduced 3.8-fold. In conclusion, exogenous leptin administered in physiological doses reversed the maturation of the small intestinal mucosa to the range found in sow-reared piglets.     

The mechanism of elevated alkaline phosphatase activity after bile duct ligation in the rat

Alkaline phosphatase activity in the liver and intestine increases after bile duct ligation, reportedly by increased enzyme synthesis. To ascertain the mechanism of this increased synthesis in the absence of a cDNA clone encoding the enzyme, we have estimated the concentration of liver and intestinal alkaline phosphatase mRNA by translational analysis. Monospecific antiserum to rat placental alkaline phosphatase was raised. The resulting antiserum precipitated two peptides of 53 and 56 kd after translation of liver poly(A) + RNA. The precipitation of both peptides was blocked by the single 64 kd placental alkaline phosphatase. Processing of the cell-free products by microsomal membranes produced peptides of 62 and 64 kd. Antiserum to rat intestinal alkaline phosphatase also identified two peptides as products of intestinal RNA translation. After bile duct ligation, we confirmed a transient 2-fold increase in alkaline phosphatase activity in the intestine and a more constant 7-fold increase in the liver. However, the alkaline phosphatase mRNA concentration remained unchanged in both organs. We conclude that increased alkaline phosphatase synthesis after bile duct ligation results from an enhanced rate of translation of mRNA.     

The effect of methotrexate on rabbit intestinal mucosal enzymes and flora

Twelve rabbits were given methotrexate (5 mg/kg) IV for 4 days and compared to a total of 23 control animals for changes in light and dissecting microscope appearance, mucosal lactase, maltase, sucrase, alkaline phosphatase, and succinic dehydrogenase activity, and changes in intraluminal flora. There were no changes in flora or histology, but statistically significant depression of lactase, maltase, and alkaline phosphatase occurred. Sucrase and succinic dehydrogenase depression was not significant. Lactase activity, in contrast to that of the other enzymes, was depressed by methotrexate, 5 mg/kg for 2 days. It is concluded that mild derangement of metabolic integrity of intestinal mucosa, as induced by small doses of methotrexate, does not result in qualitative or quantitative alteration of recoverable bacteria from the proximal intestinal lumen in rabbits.Supported in part by Grant AM 0816401 from the US Public Health Service.     

Effect of Stasis on Intestinal Enzyme Activities

Summary: Activities of the small intestinal mucosal enzymes lactase, sucrase, maltase, alkaline phosphatase and N-acetyl-β-glucosaminidase were studied in rats with surgically-induced upper intestinal stasis and in control animals. The first four are brush border enzymes, the latter a lysosomal enzyme. There was reduction in the activities of all enzymes in the operated animals. This change was significant and most marked in mucosa lining the blind loop and gut distal to it; areas in which there is gross bacterial overgrowth and excessive levels of intraluminal deconjugated bile salts. The significance of these findings in relation to malabsorption consequent on bacterial contamination of the upper gut is uncertain and requires further study.     

Sex variation in the activities of mucosal hydrolytic enzymes in the small intestine of the rat.

Mucosal response of alkaline phosphatase, ATPase and disaccharidase (lactase, maltase and trehalase) activities to sex hormones were studied by comparing male and female rats and castrated males and by injecting testosterone into castrated males. Alkaline phosphatase showed a very steep gradient in the small intestine from the oral to the aboral end, whereas ATPase activity in the ileum was still about 50% of that in the duodenum. Both enzymes showed only minor sex variations and weal response to castration. Lactase and maltase had peak activities in the jejunum, but trehalase activity was nearly equally high in the duodenal mucosa as in the jejunum. Jejunal lactase activity was about 50% lower in female than in male rats and castration decreased activity in males to the same low level as found in females. The administration of testosterone to castrated male rats did not enhance activity. Maltase activity showed similar sex variation, although castration was not able to decrease activity during the test period. Trehalase activity was lower in female than in male rats. The administration of testosterone enhance activity in castrated males.     

Prenatal Ethanol Exposure Alters the Expression of Intestinal Hydrolase mRNAs in Newborn Rats

To gain insight into the postnatal growth delay induced by ethanol in utero, we characterized functional impairments of the small intestine of neonatal rats prenatally exposed to ethanol using a well-described model of gestational alcoholism (25% ethanol w/v in the drinking water). Expression of the intestinal enzymes–lactase-phlorizin hydrolase (LPH) and intestinal alkaline phosphatase (IAP)-that are critical for enteral nutrition of neonates was studied. Characteristic patterns of LPH and IAP expression along the proximal-distal (horizontal) and crypt-villus (vertical) axes of the small intestine, as well as the intracellular localization of LPH and IAP mRNAs and immunoreactive proteins within absorptive enterocytes, were not altered by prenatal exposure to ethanol. However, a 10- to 15-fold increase in the number of LPH and IAP mRNA molecules per absorptive enterocyte was found throughout the intestine of ethanol-exposed neonates, compared with controls, whereas lectese and alkaline phosphatase activities per enterocyte remained unchanged. These findings suggest that ethanol in utero alters the mRNA abundance of epithelial enzymes in newborn rat small intestine. Changes in mRNA abundance could be an important aspect of enterocyte adaptation to high ethanol concentrations in gastrointestinal amniotic fluid of ethanol-exposed fetuses.