Characteristics of proteolysis in the gastrointestinal tract in the early postnatal period

The activity of proteinases was increased, when we used experimental casein for artificial feeding of newborn rats. Control casein contained in milk substitutes delayed protein evacuation from the stomach. The activity of intestine pancreatic proteinases was increased when we used experimental casein and decreased, for control casein. When the activity of pancreatic proteinases increased, lysosomal intestine activity decreased.     

Protein and peptide degradation in the intestine of the common brushtail possum (Trichosurus vulpecula)

A protein (bovine serum albumin: BSA) and a peptide (luteinizing hormone releasing hormone: LHRH) were used to evaluate proteolytic activity in the intestine of common brushtail possums (Marsupiala, Trichosurus vulpecula). Luminal and mucosal extracts were isolated from the duodenum, jejunum, ileum, caecum, proximal colon and distal colon, their protein content assessed and specific activities in metabolising LHRH and BSA determined in vitro. The degradation of LHRH by luminal extracts was compared with that by the pancreatic enzymes, chymotrypsin, trypsin, and elastase. The protein concentration (microg x mg-1) of mucosal extract in the duodenum was higher ( P<0.05) than in the proximal colon, but that of luminal extracts did not differ significantly between regions. Proteolytic activity of luminal extracts was greater ( P<0.01) in the jejunum and ileum than in the hindgut. In the small intestine, proteolytic activity of luminal enzymes far exceeded that of mucosal enzymes ( P<0.05). All three pancreatic enzymes hydrolysed LHRH, but chymotrypsin had the greatest activity. This study has demonstrated that, in possums, proteolysis occurs primarily in the small intestine through luminal enzymes, with chymotrypsin playing a major role. The possum hindgut contributes little to the metabolism of peptides and proteins, identifying it as a potential site to target for their absorption following oral delivery.     

Effect of cortisone on the developmental pattern of the neutral and the acid β-galactosidases of the small intestine of the rat

1. The developmental pattern and effect of cortisone on acid beta-galactosidase and neutral beta-galactosidase were studied in postnatal rats by a recently proposed method for their independent determination. 2. After birth the acid beta-galactosidase activity increases in the ileum, whereas it decreases slightly in the jejunum. On day 16 after birth the activity in the ileum decreases and in 20-day-old rats activity in both parts of the intestine decreases to adult values. In suckling animals the activity in the ileum exceeds the jejunal activity severalfold and in adult animals the activity in the jejunum is slightly higher than that in the ileum. 3. Neutral beta-galactosidase activity is high after birth and decreases in both jejunum and ileum after day 20 after birth. In 12-20-day-old rats activity in both parts is essentially the same, but in adult animals jejunal activity exceeds ileal activity four-to five-fold. 4. Cortisone (0.5, 2.0 or 5.0mg/100g body wt. daily for 4 days) does not influence the activity of either enzyme in 60-day-old rats. Acid beta-galactosidase activity is decreased after cortisone treatment in 8-, 12-, 16-and 18-day-old rats, with sensitivity to cortisone increasing with the approach of weaning. No effect of cortisone on acid beta-galactosidase is seen in 8-day-old rats. Neutral beta-galactosidase activity is increased in the ileum of 8-, 12-, 16- and 18-day old rats, but only in the jejunum of 8-and 12-day-old rats.     

Effect of protein deficiency in the female rat diet during pregnancy and lactation on the activity of the membrane and soluble forms of digestive enzymes in the offspring small intestine

Protein deficiency in female rats diet during pregnancy and lactation resulted in deceleration of induction of sucrase both forms in the jejunum and ileum; in acceleration of induction of the maltase membrane from in the jejunum; and in suppression of the lactase membrane form in the ileum; in earlier forming of the adult-type distribution of activity of the membrane form of intestinal alkaline phosphatase and in a decrease in activity of the enzyme soluble form. The findings are corroborated by a suppression of activities of the membrane and soluble forms of the small intestine digestive enzymes in 30-day old rat pups fed with a control (adequate) ration starting 21 days after the birth.     

Characteristics and postnatal development of the acid lipase activity of the rat small intestine.

Acid lipase was identified in the rat small intestine by using esters of 4-methylumbelliferone as substrates. Maximum activity towards the oleate ester was found at pH 4.0. In adult animals, the activity of acid lipase exhibited both latency and sedimentability, indicating a lyosomal localization. The activity of acid lipase was practically the same along the height of the villus, thus paralleling the distribution of acid beta-galactosidase. In adult rats, the activity of acid lipase in proximal (jejunum) and middle (mid-jejunum) sections of the small intestine was practically the same and exceeded the activity in the distal (ileum) section by a factor of 2. In suckling rats, the activity of the enzyme in the mid-jejunum exceeded that in the jejunum and ileum by 2.5- and 1.5-fold respectively. During postnatal development, the acid lipase activity of the mid-jejunum showed a peak between days 10 and 15, at which time it exceeded the adult mid-jejunum activity by 5--6-fold.     

Differential sensitivity of intestinal brush border enzymes to pancreatic and lysosomal proteases.

Isolated human intestinal brush border membranes were used as sources of enzyme to study their degradation by proteolytic enzymes. Human intestinal brush border hydrolases undergo degradation by two separate proteolytic systems. Sucrase and alkaline phosphatase are degraded by pancreatic proteases (e.g. chymotrypsin) at neutral pH, whereas trehalase is degraded by lysosomal extracts at acid pH. Both the membrane bound and membrane free isolated enzymes had similar sensitivity to proteolytic enzymes. Thus, initial removal from the membrane is not essential as a prerequisite to proteolysis. It is postulated that the brush border membrane of the intestine is subject to proteolysis by pancreatic enzymes from the external cell surface and by lysosomal proteases within the cell.     

Effect of dexamethasone on activity of glycosidases and proreinases of intestine of sterlet <Emphasis Type="Italic">Acipenser ruthenus</Emphasis>

Effect of dexamethasone (2.0 mg/kg) on activity of enzymes hydrolyzing proteinaceous and carbohydrate food components in the intestine of the starlet Acipenser ruthenus is studied. Dexamethasone modifies the activity of proteinases more than of glycosidases. As a rule, the hormone significantly decreases the level of proteolytic and general amylolytic activity of the intestinal mucosa and chyme in comparison with intact fish on the first day into the experiment and increases it on the 7th or 14th day. The dynamics of activity was different in enzymes of different chains (glycosidase and proteinase) and preparations (the mucosa and chyme).     

Differentiation of endocrinocytes in the small-intestinal epithelium of albino rats in ontogenesis]

By means of the light microscopy method differentiation and dynamics on contents of endocrinocytes have been studied in epithelium of the initial part of the jejunum, of the caudal part of the ileum and of the middle part between them during pre- and postnatal periods of development. Formation of the endocrine apparatus of the mucous membrane epithelium in the small intestine takes place simultaneously with tissue differentiation of the epithelial layer. The population density of endotheliocytes of the mucous membrane is maximal in the initial part of the jejunum and decreases gradually in the caudal direction. By the 15th day of the postnatal development endocrinocytes in composition of the epithelium in the small intestine reach a high level of the specific differentiation and their amount increases by 1.3 times in comparison with that in newborn animals.     

Effect of pH on the proteinase activities in the intestine mucosa,chyme, and enteral microbiota in the piscivorous fish differing in their ecological traits

The effect of pH on the activity of proteinases of intestine mucosa, chyme, and enteral microbiota was studied in three piscivorous fish species of the Rybinsk Reservoir differing in their ecological traits: pike Esox lucius, burbot Lota lota, and zander Sander lucioperca. In all fish species, studied pH of ten is optimal for the functioning of proteinases of intestine mucosa; in chyme proteinases, optimal pH values vary from six to ten. Optimal pH for functioning of proteinases of enteral microbiota is seven for zander and pike; pH of nine, for burbot. High activity of chyme and microbiota proteinases within the diapason of pH values of six to nine is a characteristic of burbot. Relative activity of proteinases of intestine mucosa in all fish species is not more than 15% of the maximal activity; that of chyme and enteral microbiota is lower than 40% in zander, close to 50% in burbot, and 80 and 50%, respectively, in pike. It is suggested that diversity of the patterns of pH-dependence of enteral microbiota proteinases relates to the specificity of microbiota in various ecological zones of the reservoir (littoral, sublittoral, and bathyal).     

The dynamics of proteinases activity in fish chyme and intestine mucosa upon exposure to fresh and brackish water in vitro

The dynamics of activities of proteinases in roach and perch chyme and in bream intestine mucosa upon exposure to water with various salinities for 96-216 h were studied. Irrespective of the water salinity and source of the enzymes, the proteolytic activities in chyme and intestine mucosa may rise considerably during exposure. This rise is presumably provided by the intestine microflora. This noted phenomenon is least typical for the ichthyophage-facultative benthophage perch when compared to typical benthophages. This is most likely determined by the lower species diversity of the enteral microbiota in perch.     

Activity of Peptide Hydrolases in Epithelial and Subepithelial Layers of Small Intestine of Rats of Different Age after Protein Deprivation

A significant decrease of protein content in epithelial, stromal, and muscular-serosal layers of jejunum and ileum, especially in aged animals, is revealed in rats of different age groups (young, mature, aged) after 10-day-long protein deprivation. The responses of peptide hydrolases (aminopeptidase M and glycylleucine dipeptidase) were different. There were, as a rule, no changes of these enzyme activities in small intestine of young rats, except for decreased activity of aminopeptidase M in stromal layer of jejunum and increased activity of both peptide hydrolases in epithelial layer of ileum. In mature animals, increased activities of these enzymes also was observed in the epithelial layer of jejunum and ileum and in the stromal layer of ileum in comparison with the rats receiving a full-value nutrition. In aged rats, a much more pronounced rise of these enzyme activities in the layers of small intestine was revealed after protein starvation in comparison with young and mature rats. Probably, such increase of the peptide hydrolase activities in the layers of small intestine after protein deprivation can be considered as an adaptive-protective reaction of the organism in response to formation of significant amounts of low-molecular peptides as a result of protein catabolism     

Interrelationship of glutathione–cystine transhydrogenase and glutathione reductase in developing rat intestine

1. Glutathione reductase and glutathione-cystine transhydrogenase activity in supernatant fractions of whole homogenates and homogenates of mucosal and muscular layers were determined in developing rat intestine after determination of the optimum conditions for assay of the two enzymes. In jejunum from adult rat, the K(m) values for GSSG reductase and GSH-cystine transhydrogenase activities were 0.25mm-GSSG and 0.23mm-cystine respectively. 2. The two activities could be differentiated by stability studies since GSSG reductase was stable at 60 degrees C for 10min and could be stored at 4 degrees C for 24h without loss of activity. GSH-cystine transhydrogenase, on the other hand, was denatured at 60 degrees C and completely inactive after 24h storage at 4 degrees C. 3. Based on calculations of total activities, both enzymes increased from the eighteenth day until the animals were young adults. 4. Total GSSG reductase activity increased at a greater rate with age than total GSH-cystine transhydrogenase activity as evidenced by activity ratios for GSH-cystine transhydrogenase/GSSG reductase of 0.44 and 0.12 in ileum from suckling and adult rats respectively, and 0.31 and 0.24 in jejunum from suckling and adult rats respectively. 5. In mucosa from adult rats GSSG reductase was more active in the ileum than in the jejunum, whereas GSH-cystine transhydrogenase activity was higher in the jejunum. 6. GSH-cystine transhydrogenase was active only in the muscle cells of the ileum of 7-day-old rats but became localized primarily in the mucosal layer in the adult rat. However, GSSG reductase activity was distributed evenly between the two layers throughout the intestine.     

Phosphatidylcholine synthesis in the developing small intestine.

1. Phosphatidylcholine synthesis in the foetal, newborn and adult small intestine of rats was studied by determination of cytidine diphosphocholine-1,2-diacylglycerocholine phosphotransferase (cholinephosphotransferase) and acyl-CoA-1-acyl-sn-glycerol-3-phosphocholine acyltransferase (lysophosphatidylcholine acyltransferase) activities and the incorporation of [1-14C]oleic acid into phosphatidylcholine. 2. Cholinephosphotransferase activity was low in foetal jejunum and ileum, increased 3-4 fold in the ileum by 6 days of age and by 12 days in the jejunum. Jejunal activity remained constant throughout weaning; ileal activity gradually decreased to values 25% of that of the jejunum. 3. Lysophosphatidylcholine acyltransferase activity was high in foetal jejunum and ileum, decreased 70% immediately after birth in the jejunum and increased to adult values by 12 days of age. Ileal activity decreased by 20% after birth, but decreased more rapidly at weaning to 30% of the activity in jejunum. 4. Initial rates and steady-state incorporation of [1-14C]oleic acid into phosphatidylcholine by jejunal rings of 10 day-old rats exceeded that observed in jejunal rings from adult rats by 2-4-fold. 5. In the postnatal jejunum, neither cholinephosphotransferase and lysophosphatidylcholine acyltransferase activities nor oleic acid incorporation were stimulated by cortisone administration in vivo.     

大鵟消化系统蛋白水解酶种类和活性分析

采用蛋白水解酶复性电泳(G-PAGE)技术对大(Buteo hemilasius)消化系统5种器官腺胃、胰脏、十二指肠、空肠、大肠蛋白水解酶的种类和性质进行了研究,以期为研究野生鸟类的分类地位、系统演化提供基础资料,结果表明,①受pH值的影响和制约,大消化系统蛋白水解酶的活性在碱性、中性与酸性条件下递减;②在酸性条件下,45 ku蛋白水解酶存在于除腺胃外的各受检器官;③pH 7.0时,腺胃、胰脏酶谱相似,均含有683、5、342、0 ku的蛋白水解酶;④pH 8.0时,空肠和十二指肠的蛋白水解酶种类最多、活性最强,分别检出8种和7种蛋白水解酶。总之,pH值对蛋白水解酶的活性有明显的制约作用,46、41ku蛋白水解酶随着pH值的增高而失去活性,为酸性蛋白水解酶,250、2064、5 ku蛋白水解酶随着pH值的增高活性逐渐增强,为碱性蛋白水解酶。十二指肠和空肠的蛋白水解酶种类多、活性强,可能为蛋白质消化的主要场所。     

Lysosomal proteinase activity and the the lysosomes of the small intestine in postnatal ontogeny

It has been shown that in postnatal animals, the activity of lysosomal proteinases in the distal part of the small intestine was considerably higher than in the proximal one. With transition from milk to definitive nutrition, the activity of lysosomal proteinases gets equalized in the proximal and distal parts of the small intestine. The data concerning the proximo-distal gradient of distribution of the activity of lysosomal proteinases have been supported. Transition from milk to definitive nutrition has given rise to ultrastructural alterations in the lysosomal apparatus.     

Experimental data proving the presence of inhibitors of amylase activity in biliary and pancreatic juice

Two parallel studies on albino male rats are performed. In the first study, there is a group which underwent resection of the proximal third of the small intestine. While the other group despite resection of the same segment also has a ligated common biliary and pancreatic duct. In the second study, one group of the experimental animals is only with ligated pancreatic duct and in the other group the same duct is implanted in the initial part of the ileum. On the 15th day after the surgical interventions the amylase activity and the absorption of glucose in the small intestine are studied by the method of turned sacs "in vitro". It is established that the glucose transport does not change after the four surgical interventions. However, the amylase activity increases about twice times after resection of the upper third of the small intestine and more than 4 times after resection of the same segment with simultaneous ligature of the common biliary and pancreas duct. Only at ligating the duct, the amylase activity is decreased in the jejunum and is significantly increased in the ileum, while its implantation in the initial part of the ileum does not change its activity in both studied segments of the small intestine. It is concluded that there are unknown inhibitors for the amylase activity in the biliary and pancreatic juice. The discussed issue is why they inhibit only the enzymatic compensatory processes without influencing the transport systems of the small intestine.     

Diversity of digestive proteinases in Tenebrio molitor (Coleoptera: Tenebrionidae) larvae

The spectrum of Tenebrio molitor larval digestive proteinases was studied in the context of the spatial organization of protein digestion in the midgut. The pH of midgut contents increased from 5.2-5.6 to 7.8-8.2 from the anterior to the posterior. This pH gradient was reflected in the pH optima of the total proteolytic activity, 5.2 in the anterior and 9.0 in the posterior midgut. When measured at the pH and reducing conditions characteristic of each midgut section, 64% of the total proteolytic activity was in the anterior and 36% in the posterior midgut. In the anterior midgut, two-thirds of the total activity was due to cysteine proteinases, whereas the rest was from serine proteinases. In contrast, most (76%) of the proteolytic activity in the posterior midgut was from serine proteinases. Cysteine proteinases from the anterior were represented by a group of anionic fractions with similar electrophoretic mobility. Trypsin-like activity was predominant in the posterior midgut and was due to one cationic and three anionic proteinases. Chymotrypsin-like proteinases also were prominent in the posterior midgut and consisted of one cationic and four anionic proteinases, four with an extended binding site. Latent proteinase activity was detected in each midgut section. These data support a complex system of protein digestion, and the correlation of proteinase activity and pH indicates a physiological mechanism of enzyme regulation in the gut.     

ABCG2 expression and uric acid metabolism of the intestine in hyperuricemia model rat

Abstract

To elucidate roles of the intestine in uric acid (UA) metabolism, we examined ABCG2 expression, tissue UA content and xanthine oxidoreductase (XOR) activity in different intestinal segments. Male SD rats were assigned to control group or oxonic acid-induced hyperuricemia (HUA) group. In control rats, ABCG2 was present both in villi and crypts in each segment. Tissue UA content and XOR activity were relatively high in duodenum and jejunum. However, in HUA rats, tissue UA content was significantly elevated in the ileum, whereas it remained unaltered in other segments. Moreover, ABCG2 expression in the HUA group was upregulated both in the villi and crypts of the ileum. These data indicate that the ileum may play an important role in the extra-renal UA excretion.     

Effect of diversion of bile-pancreatic juice to the ileum on pancreatic secretion and adaptation in the rat

Cannulas were implanted to collect bile and pancreatic juice, and the collected secretions were pumped back into the intestine at the level of the duodenum or the proximal ileum. The effect of 6 days of such treatment on pancreatic secretion and on pancreatic growth was determined. The effect on pancreatic secretion was studied by measuring the pancreatic secretory response to a stimulus, provided by acute diversion of bile-pancreatic juice from the proximal intestine. Trophic effects were studied in a separate group of rats by measuring pancreatic weight, protein content, and chymotrypsin activity after an overnight fast. Stimulated pancreatic secretion was 2.1 times greater for protein output and 3.4 times greater for fluid output in rats with chronic diversion of bile-pancreatic juice to the ileum. Pancreatic weight, protein content, and chymotrypsin activity were increased 2.6, 2.9, and 4.8 times, respectively, by chronic diversion of bile-pancreatic juice to the ileum. These results indicate that pancreatic hypertrophy and hyperplasia reported in rats with bile-pancreatic duct transposition to the ileum are the result of loss of feed-back inhibition from bile-pancreatic juice in the proximal intestine.     

The effect of starvation on the control of phosphofructokinase activity in the epithelial cells of the rat small intestine.

1. The effect of depriving rats of food for 48 h on the specific activity of phosphofructokinase in the epithelial cells of the small intestine and on the regulatory properties of the enzyme displayed in crude (particle-free) mucosal extracts was studied. 2. The specific activity of phosphofructokinase, measured under optimal conditions at pH8, in the mucosa of fed rats showed a negative aboral gradient along the intestine, decreasing from 15.2 +/- 1.2 units (mumol/min)/g wet wt. in the proximal jejunum to 4.6 +/- 1.2 units/g wet wt. in the terminal ileum. 3. After starvation, the gradient was diminished, but not abolished; the diminution in gradient was due almost exclusively to a decrease in the specific activity of phosphofructokinase in the proximal jejunum by about 30%, there being no change in the terminal ileum. 4. In fed rats, the susceptibility of phosphofructokinase to inhibition by ATP, when assayed in crude mucosal extracts under suboptimal conditions, was independent of length along the small intestine; the ratio of the activity observed at pH 7.0 in the presence of 0.5 mM-fructose 6-phosphate and 2.5 mM-ATP to the optimal activity at pH 8, v0.5/V, was 0.36 +/- 0.05 in the proximal jejunum and 0.42 +/- 0.07 in the terminal ileum. 5. After starvation, the susceptibility of phosphofructokinase to inhibition by ATP was increased and was again found to be independent of length along the small intestine: after starvation, v0.5/V was 0.19 +/- 0.04 and 0.20 +/- 0.07 for the proximal jejunum and the terminal ileum respectively. 6. Re-feeding of previously starved rats on a high-carbohydrate diet overnight for 16 h restored both the specific activities of phosphofructokinase and its susceptibility to inhibition by ATP to normal values for fed rats. 7. The data support the idea that the specific activities and the regulatory properties of phosphofructokinase in the epithelial cells of rat small intestine are mediated by distinct humoral factors. 8. The changes in glucose utilization rate of the jejunum when rats are starved can in principle be accounted for by a combination of changes in the specific activity and in the regulatory properties of mucosal phosphofructokinase.