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.     

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 effect of temperature on chyme proteinase activities in perch Perca fluviatilis L. and roach Rutilus rutilus (L.)

The effect of temperature on the chyme proteinase activities of the perch Perca fluviatilis L. and the roach Rutilus rutilus (L.) was investigated. The two fish species showed different characters of temperature dependence and different values of activation energy for caseinolytic and hemoglobinolytic chyme proteinases. The perch was characterized by a higher level of relative proteinase activities and lower activation energy for protein hydrolysis at a temperature of 0–10°C than the roach.     

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).     

The influence of testosterone on activity of glycosidases and proteinases in the intestine of the sterlet (Acipenser ruthenus)

Observation of the influence of testosterone (0.7 mg/kg) on the activity of glycosidases and proteinases, which function in the chyme and the mucosa of the sterlet intestine (Acipenser ruthenus), revealed a decrease in enzyme activity of both chains as against that one typical for intact individuals. The activity of the investigated enzymes changes in both the experimental and control groups of fish. Nevertheless, they have unlike dynamics of this activity. In a number of cases, we indicated a significant increase in enzyme activity of both chains in comparison to the control group. We also observed an increase in activity of chyme glycosidases, which was significant on the 21st day as compared to intact fish.     

Milk protein digestion and lysosomal proteinases of the ileal mucosa in young rats]

The proteolytic activity of lysosomal and pancreatic proteinases was studied in the chyme and tissue homogenates of the jejunum and ileum of 12- and 30-day rats in order to elucidate whether lysosomal proteinases of the mucous membrane of the ileum participate in cavitary digestion. During milk feeding the proteolytic activity of acid (lysosomal) proteinases in the ileum was 3 times greater than in the jejunum, which has been demonstrated both for the mucous membrane and the contents of these parts of the small intestine. In rats on definitive nutrition the activity of acid proteinases from the jejunum and ileum remained almost unchanged both in the mucous membrane and in the contents. The data obtained also indicate that pancreatic proteinase absorbed from the chyme of the small intestine may participate in parietal digestion.     

Effect of temperature on proteinase activities in intestinal chyme and mucosa of fish of different ecological groups

Effect of temperature on activities of proteinases in intestinal chyme and mucosa was studied in three fish species (pike-perch, zope, roach) belonging to different ecological groups by their nutrition type. There was revealed a significant difference of dependence of enzyme activities in chyme on temperature in the benthophage, roach (a higher level of relative activity in the range of lower temperatures and a wider zone of temperature optimum) as well as of values of apparent energy of activation of the protein hydrolysis process as compared with that in planktoand ichtyophages, zope and pike-perch, which indicates a significant effect of the enteral microbiota proteinases and of nutrition objects on characteristics of hydrolases functioning in fish intestine.     

Effect of temperature on proteinase activities in the intestinal chyme and the mucosa of various ecological fish groups

Effect of temperature on activities of proteinases in intestinal chyme and mucosa was studied in three fish species (pike-perch, zope, roach) belonging to different ecological groups by their type of feeding. There was revealed a significant difference of dependence of enzyme activities in chyme on temperature in the benthophage roach (a higher level of relative activity in the zone of lower temperatures and a larger zone of temperature optimum) as well as of values of apparent energy of activation of the protein hydrolysis process as compared with that in plankto- and ichthyophages--zope and pike-perch, which indicates a significant effect of the enteral microbiota proteinases and of nutrition objects on characteristics of hydrolases functioning in fish intestine.     

Effect of pH on the activity of proteinases in intestinal mucosa,chyme, and microbiota of fish from the Cuciurgan reservoir

Proteolytic activities of the intestinal mucosa, chyme, and enteral microbiota have been studied in a wide range of pH values in five fish species from the Cuciurgan reservoir (Moldova). Differences in pH dependence of the intestinal proteinase activity of fish are determined by their feeding type. The maximum activity of proteinases is found in the pumpkinseed Lepomis gibbosus. The minimum activity of proteinases has been demonstrated by the zander Zander lucioperca. The pH optimum of the mucosa and chyme in all fish species (except for the European perch Perca fluviatilis) is 10. The pH optimum of the intestinal microflora varies from 6.0 (in the common carp Cyprinus carpio) to 10 (in the crucian carp Carassius carassius), whereas that in the perch from the Cuciurgan and Rybinsk reservoirs is 7. The majority of fish species, mostly Zander lucioperca and Lepomis gibbosus, are characterized by high proteniase activity of the microbiota, in the pH ranging from 6 to 9. It is assumed that proteinases in the enteral microbiota of fish are able to make up for the relatively low activity of those synthesized by their digestive system in the range of low pH values.     

Impact of polychlorinated biphenyls on the activity of intestinal proteinases and carbohydrases in juvenile roach Rutilus rutilus (L.)

The chronic effects that polychlorinated biphenyls (PCBs) have on the activities of proteinases and carbohydrates in intestinal mucosa and chyme in juvenile roach Rutilus rutilus (L.) have been studied for the first time. Upon consuming food with PCB 50.8 ng/g wet weight for 218 days, the exposition of fingerlings in aquariums with dirt bottoms (contents PCB 425.6 ng/g dry weight) reduces the proteolytic activity of mucosa to a greater degree than amylolytic activity; the activity of sucrase changes in different directions. In 2-year-old fish receiving PCB only with food (50.8 ng/g wet weight bottom), proteolytic and amylolytic activities of the mucosa are reduced 18 and 35% in comparison with the control (52 days); the sucrose activity is reduced 13% (52 days) and 22% (169 days). In most cases the relation of amylolytic activity to proteolytic activity in fish of the experimental groups is less than in the control. The enzyme activities in chyme change in different directions, depending on the terms and conditions of the experiment.     

Generalized and selective feedback inhibition of secretion of pancreatic enzymes

Selective inhibition of secretion of proteinases, amylase and lipase by the pancreas, was revealed. The inhibition is directly dependent on the dose of intraluminal enzymes. The latter's increase transforms the selective inhibition into a generalised one. Possible mechanisms of selective and generalised feedback inhibition of the exocrine pancreatic secretion is discussed, the secretion playing an important role in urgent adaptation of the pancreatic enzymes secretion to the duodenal chyme properties.     

Activity of tissue cathepsin-L-like proteinases of women with womb body oncopathology

Activity and optimal pH of cathepsin-L-like proteinases was studied in benign and malignant tumours of the womb body. In the benign tumors activity of cathepsin-L-like proteinases changes depending on the expansion and depth of extension benign tumour and is defined by proliferative potential of tumour cells of myometrium and endometrium. Activity of cathepsin-L-like proteinases in malignant epithelial tumour of endometrium--adenocarcinoma is inversely proportional to the level of differentiation of the tumour cells.     

Effect of low-temperature hardening on activities of proteolytic enzymes and their inhibitors in the leaves of wheat and cucumber seedlings

On seedlings of winter wheat (Triticum aestivum L.) and cucumber (Cucumis sativus L.), the dynamics of cysteine and serine trypsin-like proteinases and also trypsin inhibitors at cold hardening (5°C for wheat and 10°C for cucumber) was studied. Activation of proteinases and inhibitors coincided in time or preceded an increased tolerance in wheat and cucumber seedlings in the early period of their hardening. After attaining the highest wheat tolerance, activity amidases reduced, whereas the increased activity levels of cysteine proteinases and trypsin inhibitors was maintained during the entire period of hardening. In cucumber, in these period activities of amidases and trypsin inhibitors reduced, whereas the activity of cysteine proteinases was maintained at the level close to the initial one. It is suggested that cysteine proteinases, amidases, and trypsin inhibitors are involved in plant adaptation to cold.     

Effect of thyroid hormones on activity dynamics of enzymes of intestinal mucosa of juvenile roach <Emphasis Type="Italic">Rutilus rutilus</Emphasis>

The effect of thyroid hormones on activity dynamics of enzymes (proteinases and glycosidases) of intestinal mucosa of juvenile roach Rutilius rutilus was investigated. Application of substances increasing and decreasing the level of thyroid hormones in blood plasma significantly influences the growth rate and the activity of proteinases and glycosidases functioning in the intestinal mucosa. In most cases, the activity level of trypsin-like proteinases and the activity of glycosidases in the fish exposed to triiodothyronine were significantly higher than in the control. The activity level of chymotrypsin-like proteinases in fish form the group with exposure of exogenous triiodothyronine only in the end of the experiment surpassed the values of this parameter in the control fish. In the fish developing at deficiency of thyroid hormones, the growth rate and proteinases activity were significantly lower in comparison with the control.     

Hydrolytic activity of Virgibacillus sp. SK37, a starter culture of fish sauce fermentation, and its cell-bound proteinases

Fish sauce production relies on a natural fermentation process requiring 12-18 months for process completion. Virgibacillus sp. SK37 has been shown to be a potential strain for fish sauce acceleration. However, hydrolytic activity of proteinases bound at cell surface of this strain has not been well elucidated. Addition of 0.2 % CaCl(2) (w/w) in conjunction with starter cultures of Virgibacillus sp. SK 37 increased protein hydrolysis as measured by α-amino group content throughout fermentation (P < 0.05). Cell-bound proteinases from Virgibacillus sp. SK 37 were extracted into a free form by incubating the washed cells in Ca(2+)-free buffer at 37 °C for 2 h. Cell-bound proteinases revealed molecular mass of 19, 20, 22, 32, 34, and 44 kDa based on a synthetic peptide zymogram. The proteinases showed subtilisin-like serine characteristics with the highest activity at 50 °C and pH 8 and 11. Activity of the extracted proteinases increased ~4 times at ≥100 mM CaCl(2). In addition, CaCl(2) enhanced thermal stability of the extracted proteinases. Enzymes showed proteolytic activity in either the absence or presence of 10 and 25 % NaCl toward fish muscle, soy protein isolate, and casein substrates. Cell-bound proteinases were likely to play an important role in protein hydrolysis during fish sauce fermentation.     

Purification and partial characteristic of a major gliadin-degrading cysteine endopeptidase from germinating triticale seeds

The endopeptidase of the highest electrophoretic mobility was the main endopeptidase hydrolyzing gliadin in the endosperm of germinated triticale (X Triticosecale Wittm.) grains after three days of imbibition. Activity of this endopeptidase, named EP8 starts to be detectable after two days of imbibition. The appearance of its activity in the endosperm on a second day of imbibition may suggest that EP8 is synthesized in aleurone during germination and/or secreted into the starchy endosperm as an inactive polypeptide during grains development and then activated. EP8 was isolated from the endosperm of germinating triticale seeds and purified 257-fold using ammonium sulphate, ion exchange chromatography on DEAE Sepharose CL-6B and gel filtration on Sephadex G-100. The enzyme was totally inhibited by E-64—class-specific cysteine proteinases inhibitor and activated by thiol compounds. Molecular weight estimated by SDS-PAGE was 39.5 kDa. The optimum pH for the hydrolysis of gliadin was 4.2 and for hemoglobin 5.2. High activity of EP8 against wheat gliadin in vitro suggests that this cysteine endopeptidase plays a major role in the mobilization of storage proteins in the endosperm of germinating triticale grains.     

Effect of macrophage differentiation and exposure to mildly oxidized LDL on the proteolytic repertoire of THP-1 monocytes

Lipid-laden monocyte/macrophages in atherosclerotic plaques can produce a range of proteinases capable of degrading components of the plaque extracellular matrix, an event that may weaken plaques, rendering them vulnerable to rupture. The effects of differentiation from monocytes to macrophages and exposure to mildly oxidized LDL (Ox-LDL) on the expression of a range of proteinases and their inhibitors were assessed in the human THP-1 cell line. Of 56 proteinases/inhibitors investigated, 17 were upregulated during macrophage differentiation, including several matrix metalloproteinases (MMPs) and cathepsins along with their native inhibitors. Similarly, expression of matrix-degrading proteinases was also increased during differentiation of human primary macrophages. In conjunction, the proteolytic capacity of the cells increased, as assessed by substrate zymography. Subsequent exposure of differentiated THP-1 cells to mildly Ox-LDL increased the expression of a control gene (adipocyte lipid binding protein) and increased the activity of nuclear factor-kappaB and activator protein-1 in serum-free conditions but did not significantly affect the expression of any of the proteinases or inhibitors investigated. These results indicate that in this model macrophage differentiation, rather than exposure to Ox-LDL, has a more important effect on the expression of genes involved in extracellular matrix remodeling.     

Activity and respiration in the anemone, Metridium senile (L.) exposed to salinity fluctuations

Activity and respiration in the anemone, Metridium senile (L.), were monitored under both constant and fluctuating salinity conditions. During constant exposure to 50% sea water it was found that the animals retracted the tentacles and that the rate of oxygen consumption decreased by ≈50%. The same response was elicited from animals in 100% sea water in a contracted state. Animals exposed to continually fluctuating salinities were found to retract the tentacles, contract the body wall, and produce amounts of mucus during periods of decreasing salinities. These reactions were reversed during exposure to increasing salinity. Oxygen consumption never ceased entirely in animals exposed to dilute sea water and it was found that during declining oxygen tension M. senile regulated its oxygen consumption until the environmental oxygen tension fell to ≈30% saturation.     

The content of sex hormones in the enteric medium of female ruminants

It has been established that in the chyme of females of the seep Ovis ammon and the reindeer Rangifer tarandus with an actively functioning corpus luteum of the estrous cycle or in pregnancy, the total amounts and concentrations of progesterone and estradiol are higher, while the progesterone/estradiol ratio, lower, than in the blood serum. The conclusion has been made that the transport of these hormones across the rumen wall occurs from the blood into the chyme, while across the cecal wall, from the chyme into the blood.     

Transport of fat through jejunal absorptive epithelium in vitro

The mucosa of excised pieces of jejunum of fasting rats was exposed for 10 min to fresh chyme obtained from other rats which had been digesting either buttered bread or bread alone. The mucosa was then processed for electron microscopy of lipid material. After exposure to fatty chyme, fat globules were scattered throughout the cytoplasm of absorptive epithelial cells, and concentrated in both supranuclear and infranuclear components of the Golgi apparatus. They were also present within lymphatics of the villus. After exposure to both types of chyme, the basal parts of the cells were much elongated through collection of a finely granular deposit of non-lipid material. This collection was probably due to failure of clearance of absorbed material from the cells in the absence of blood flow through the villus.