Biomedical subjects

V Keim

Publications and source records attributed to V Keim.

89 records · Page 5Linked to original sources

PubMed1988

Identification of rat pancreatic secretory proteins after separation by high-performance liquid chromatography.

In the present study an improved method of reversed-phase high-performance liquid chromatography (HPLC) for separation of rat pancreatic juice proteins is introduced. Aliquots of pancreatic juice were saved from conscious rats during basal secretion. The secretory proteins were separated on a wide-pore silica column by use of a multistep acetonitrile/water gradient. Up to 14 individual peaks could be separated by one run. Molecular weight analysis by sodium dodecyl sulfate (SDS)-gels allowed identification of peaks representing amylase, lipase, procarboxypeptidases, proelastase, chymotrypsinogen, and trypsinogen. Injection of pure rat amylase increased one specific peak which was assumed to represent amylase in the juice profile. Small amounts of residual enzymatic activities were measured for amylase, trypsin, and chymotrypsin in material of certain peaks. Activities of lipase, ribonuclease, and carboxypeptidases were not found, which reflected degradation of these enzymes by the separation procedure. High activities of phospholipase A2 were detected in one specific, early-eluting peak. Reversed-phase HPLC offers precise, reproducible, and rapid separation of the major proteins of rat pancreatic juice.

Animals↗

PubMed1988

Changes in pattern of enzyme secretion by rat pancreas during repeated trypsin inhibitor treatment.

In conscious rats with cannulated pancreatic ducts, the synthetic trypsin inhibitor camostat (400 mg/kg) was instilled into the stomach once daily on 5 consecutive days. Each camostat administration stimulated the secretion of juice, bicarbonate, and protein. From day 2 to day 5, basal protein secretion measured before the daily instillation of camostat was increased, whereas basal secretion of volume and bicarbonate remained unchanged. The secretory responses to camostat increased up to day 3 of treatment without further change on days 4 and 5. In response to acute camostat instillation, parallel secretion of enzymes was found. Within 1 day, however, a change in the composition of secreted enzymes was observed. After 3 days, the percentage of amylase in pancreatic juice decreased by approximately 50%, whereas the relative amount of proteinases and the specific activity of phospholipase A2 doubled. The alteration of protein patterns in pancreatic juice parallels the effect on enzymes stored in the gland. The results demonstrate that adaptation of enzyme secretion in the exocrine pancreas occurs after a short period of treatment with camostat. Because this compound stimulated protein, volume, and bicarbonate secretion, it is suggested that other factors in addition to cholecystokinin may be involved in this adaptation process.

Amylases↗

PubMed1988

Influence of experimental hyperthyroidism on the adult rat pancreas, small intestine, and blood gastrin levels.

Thyroxine modulates the ontogenic changes of animal tissues. In this study, the effects of experimental hyperthyroidism on the adult rat pancreas, small intestine, and serum gastrin were evaluated. Hyperthyroidism was induced by oral feeding of thyroxine (T4) in increasing dosages (150-450 micrograms/kg body weight; 3 weeks) and controlled by measurements of the circulating hormones. The increase of thyroid hormones in blood (T4 ng/dl: thyroxine-treated rats 10.8 vs. controls 3.3; p less than 0.01; given are means) was accompanied by hypergastrinemia (IR-gastrin pg/ml: T4-treated rats 169 vs. controls 25; p less than 0.05). The T4-treated animals consumed more food but lost about 20 g of their initial body weight. Pancreatic wet weight (g: T4-treatment 1.72 vs. controls 1.42; p less than 0.05), DNA (micrograms/g body weight: T4-treatment 2.42 vs. controls 1.5; p less than 0.05), and protein (micrograms/g body weight: T4-treatment 131.7 vs. controls 63.5; p less than 0.05) were increased, whereas no pronounced influence on pancreatic amylase, trypsin, and chymotrypsin was found. The gut wet weight after thyroxine administration (18.1 g vs. 15.4 g of controls; p less than 0.05) was elevated, but length, DNA, protein, and brush border enzyme activities remained unaltered. Our data demonstrate in adult rats a small but significant trophic response of pancreas and gut to repeated oral thyroxine administration.

Animals↗

PubMed1987

Evidence in vivo of asynchronous intracellular transport of rat pancreatic secretory proteins.

Labeled proteins which appeared in pancreatic juice after the intravenous (i.v.) injection of [35S]methionine into conscious rats with chronic pancreatic duct fistulae were separated by gel electrophoresis and measured by determination of the radioactivity of each of the separated bands. Radioactivity appeared in the secreted proteins 20 min after injection of the label. In the subsequent 10 min, 6.44% of the radioactivity was found in trypsinogen, whereas 100 min later only 3.4% of the radioactivity was associated with this enzyme. The values at 10 and 100 min for amylase were 10.85% and 21%, respectively, showing an earlier appearance of labeled trypsinogen than of amylase. Chymotrypsinogen behaved similarly to trypsinogen. Early secretion of labeled proteases was also demonstrated by separation of pancreatic proteins by two-dimensional gel electrophoresis followed by fluorography. In pancreatic duct cannulated rats, zymogen granules were prepared 30 and 60 min after injection of the labeled methionine. Determination of the radioactivity of the individual proteins demonstrated a similar time course of the labeling pattern in the zymogen granule fraction to that in pancreatic juice. The results of the experiments suggest an asynchronous secretion of newly synthesized rat pancreatic proteins.

Amylases↗

PubMed1987

Influence of secretagogues on asynchronous secretion of newly synthesized pancreatic proteins in the conscious rat.

The secretion of newly synthesized pancreatic enzymes was studied in pancreatic duct cannulated rats after intravenous injection of 100 microCi of [35S]methionine. Secretion rate was stimulated by intravenous infusion of either cerulein (0.2 microgram/kg h) or carbachol (10 nmol/kg h) starting simultaneously with or 180 min before the injection of the labeled methionine. Secretory proteins were analyzed by sodium dodecyl sulfate (SDS) gel electrophoresis or by nondenaturing gel electrophoresis followed by determination of the radioactivity associated with the individual proteins. Similar to unstimulated controls in all experiments, an early secretion of newly synthesized trypsinogen and chymotrypsinogen was found, whereas amylase and lipase were secreted only after a certain lag period. The results suggest that the intracellular transit of endoproteases is faster than that of other enzymes, irrespective of whether or not secretagogues were applied.

Amylases↗

PubMed1987

Tryptophan rich diet as a new approach to study the serotoninergic enteropancreatic axis.

The influence of a tryptophan enriched diet (L-tryptophan added as 1% of total diet), fed over 10 days, on the rat duodenum and pancreas was studied by immunohistology, measurements of serotonin and tryptophan tissue concentrations by HPLC, and incubations of pancreatic lobules. Ingestion of a tryptophan enriched diet resulted in increased contents of tryptophan and serotonin in the duodenum that was not accompanied by a significant change of the serotonin cell density. Neither basal nor CCK-stimulated amylase release from isolated pancreatic lobules was altered after tryptophan enriched food. Although serotonin could be extracted from the pancreas, no increase in serotonin concentration was detected after ingestion of the tryptophan diet. A 'serotonin loading' diet may be a useful tool to study the significance of amines produced by gut endocrine cells in respects to enteropancreatic connections.

Animals↗

PubMed1986

Pancreatic secretion in the rat influenced by the low molecular weight serine proteinase inhibitor Gabexate mesilate.

The effect of intravenous or intragastric administration of the synthetic proteinase inhibitor Gabexate mesilate (GM) on the pancreas of rats was investigated. Infused intravenously at 4 mg kg-1 h-1, GM inhibited both basal or cerulein (0.2 microgram kg-1 h-1)-stimulated pancreatic protein secretion. Intracellular transport and secretion of newly synthesized pancreatic enzymes was not influenced by intravenous infusion of GM. Intragastric administration of GM (400 mg kg-1) on four consecutive days increased pancreatic wet weight, protein and enzyme content of the gland. A preferential increase of proteinases above glucosidases was observed. Pancreatic lobules from inhibitor-treated rats released 30% less amylase in response to cerulein or carbachol when the rate of discharge was expressed in percent of initial content. Expressed in ku amylase/microgram DNA secretion rate was two-fold higher than in controls. In pancreatic duct cannulated rats GM (400 mg kg-1 h-1), introduced intragastrically on five consecutive days, stimulated volume-bicarbonate and protein secretion rate, the secretory response on the fifth day being significantly higher than on the first day. Enzyme pattern in pancreatic juice changed characteristically: mainly the amount of acidic proteinases increased, whereas the amount of the basic isoforms was altered only slightly.

Animals↗

PubMed1986

Rapid adaptation of pancreatic enzyme secretion in the conscious rat. I. Influence of endogenous and exogenous stimulation.

Pancreatic protein composition in basal and stimulated pancreatic juice was studied in conscious rats with cannulated pancreatic ducts. The animals were fed with 3 different diets for 14 days. Secretion rate was stimulated by acute derivation of the juice, intravenous infusion of cerulein and intraduodenal infusions of either amino acids, fat or carbohydrate. In the juice the amount of secretory proteins was measured. The various secretory stimuli did not lead to any rapid change of composition of the secreted enzymes. A so-called 'nonparallel secretion' of pancreatic enzymes could not be found in the conscious rat.

Amylases↗

PubMed1986

Rapid adaptation of pancreatic enzyme secretion in the conscious rat. II. Effects of fasting and dietary modulation.

The influence of fasting and dietary modulation on basal and stimulated pancreatic enzyme secretion was studied in conscious rats with cannulated pancreatic ducts. Two days' fasting decreased pancreatic secretion by about 30%. After stimulation of the secretion rate by acute derivation of pancreatic juice or by intravenous infusion of cerulein (0.2 micrograms/kg/h) no rapid change of enzyme composition was found. In further experiments, two groups of rats were fed with carbohydrate-rich and carbohydrate-poor diets, respectively. After pancreatic duct cannulation and a postsurgical recovery period, the diets were interchanged. An adaptation of pancreatic enzyme secretion to the new diet was observed in pancreatic juice. Stimulation of the secretion rate during the adaptation process by acute derivation of the juice or by intravenous infusion of either cerulein, secretin, carbachol or cerulein plus secretin did not further modify the composition of pancreatic proteins. 'Nonparallel' secretion of enzymes was never found when sufficient time for recovery from pancreatic duct surgery was allowed.

Animals↗

PubMed1986

Asynchronous secretion of newly synthesized pancreatic proteins in the rat.

The secretion of newly synthesized pancreatic proteins was studied in conscious rats with cannulated pancreatic ducts. Labeled amino acids (3H-leucine and 14C-amino acid mixture) were injected intravenously. The proteins of the pancreatic juice were separated by gel electrophoresis, and the radioactivity in each band was determined. An early secretion of labeled trypsinogen and chymotrypsinogen was found, whereas amylase and lipase were secreted after a certain lag period. In vitro pulse chase experiments showed that amylase was synthesized in the acinar cell but did not move with the same efficacy to the zymogen granules and into the pancreatic juice as proteases.

Amino Acids↗

PubMed1986

Pancreatitis-associated protein in bile acid-induced pancreatitis of the rat.

Experimental pancreatitis was induced in rats by intraductal injection of 0-4% Na-taurocholate. After 0-21 days the glands were removed and amylase, trypsin and chymotrypsin were measured in pancreatic homogenates. The enzyme activities decreased within 48 h to 5-20% of the control values. When 0-1% Na-taurocholate was used, the values tended to increase after 10-21 days. In pancreatic tissue, 12 h after induction of pancreatitis a 'pancreatitis-associated protein' (PAP) was found, which persisted for 4-7 days. A correlation could be found between the severity of pancreatitis and the amount of PAP in pancreatic homogenates.

Amylases↗

PubMed1986

Prevention of experimental pancreatitis by somatostatins.

Somatostatin is known to exert a beneficial cytoprotective effect in acute experimental pancreatitis, although it is ineffective regarding mortality in human pancreatitis. The somatostatin analog 008 was found to be more efficient in taurocholate-induced pancreatitis in rats and in ceruletid-induced pancreatitis.

Amylases↗

PubMed1985

Failure of secretin to prevent or ameliorate cerulein-induced pancreatitis in the rat.

Infusion of supramaximal concentrations of the synthetic pancreozymin analog cerulein induces acute edematous pancreatitis in the rat. Vacuolization and necrosis of acinar cells is paralleled by an almost complete reduction of pancreatic secretion from the cannulated duct. Preinfusion or coinfusion of synthetic secretin slightly increases pancreatic volume and protein secretion. This effect is only transient, and is always overcome by the actions of cerulein. Secretin does not prevent or improve the cellular destruction of the acinar cells. The results suggest that secretin has no beneficial effect on hormone-induced pancreatitis.

Amylases↗

PubMed1984

An additional secretory protein in the rat pancreas.

An additional protein in rat pancreatic juice has been observed, which is present in healthy rats after pancreatic duct cannulation or in rats in which experimental pancreatitis has been induced by either cerulein or taurocholate. The protein appears 1/2-1 day after surgery or onset of pancreatitis, is present for the following 3-4 days and disappears afterwards. It is found in pancreatic homogenate or in zymogen granules from rats with pancreatitis, but not in normal rats. It does not seem to be related to the recently described 'pancreatic stone protein'. We would like to refer to this protein as 'pancreatitis-associated protein'.

Animals↗

PubMed1982

Physiological studies of exocrine pancreatic secretion in conscious rats. 5. Communication: circadian rhythms of flow rate and composition of pancreatic juice.

1. In conscious rats provided with indwelling pancreatic and biliary catheters, the circadian variations of flow rate, electrolyte- and protein concentrations are followed. 2. Individual enzymes are separated by Polyacrylamid-Gelelectrophoresis and quantitatively determined by photometry after evaluation of the coloured bands. 3.) Volume flow rate increases towards the evening and reaches maximal values in the early morning hours (2-4 a. m.). 4. A similar nocturnal maximum can be observed for the concentrations and elimination rates of total protein, potassium and bicarbonate, while the concentration of sodium is fairly constant and only slightly decreasing in very high flow rates. 5. Typical circadian variations can be observed for the individual enzyme fractions: at night, the part of amylase is higher and the amount of chymotrypsinogen is lower, together with the increased nocturnal protein elimination. At day time these relations are inversed. 6. This rhythm is also existing in fasting animals, although with reduced basal values and smaller amplitude of variation. 7. In fasting animals the relative part of amylase is smaller compared with that of chymotrypsinogen, probably in relation to the reduced elimination of total protein. 8. The circadian variations of exocrine pancreatic secretion may be primarily inaugurated by hormonal action of hormones, whose release is controlled by nervous centers in the brain.

Animals↗

PubMed1981

Physiological studies of exocrine pancreatic secretion in conscious rats. 3. Communication. Separation and determination of enzymes by gel-electrophoresis.

1. A method is described for the electrophoretic separation and following quantitative determination of the different protein fractions of pancreatic juice. 2. A polyacrylamide (PAA) gel-electrophoresis is performed in two systems: one anionic at pH = 9.0 with 7.5% PAA and a cationic one at pH = 5.0 with 10% PAA. 3. After about two hours of separation using a current of 3 and 4 mA respectively, the gels are stained with Amido-black 10B after fixation of the proteins with acetic acid. 4. The colour in excess is removed with acetic acid. 5. The coloured bands are isolated by cutting. 6. Evaluation of the colour by NaOH and determination of the colour intensity by photometry. 7. Evaluation of the colour extinction as relative percentages of the different protein and enzyme fractions. (Extinction of total dye = 100%) 8. There are at least 13 bands in the cationic system and about 7 additional ones in the anionic system distinctly separated, 12 of them could be identified using enzymatic methods. 9. Evidence is given that the colour intensity has a linear relationship to enzyme activity. 10. The validity of the presented method is discussed and compared with other electrophoretic methods.

Amylases↗

PubMedDate not supplied

PAP I interacts with itself, PAP II, PAP III, and lithostathine/regIalpha.

PAP I, PAP II, PAP III, and lithostathine/regIalpha are members of a multigenic family of proteins expressed in several tissues. PAP I was shown to be antiapoptotic, mitogenic, and anti-inflammatory and can promote cell adhesion to the extracellular matrix. Lithostathine/regIalpha can be mitogenic. Because polymerization might regulate activity, we examined the ability of rat PAP I to interact with itself (homodimerization), PAP II, PAP III, and lithostathine/regIalpha (heterodimerization) by the yeast two-hybrid system, affinity experiments, and crosslinking. PAP I interacted significantly with all members of the PAP protein family, homodimerization showing the strongest interaction as judged by the beta-galactosidase test. This was confirmed by showing specific affinity between a MBP-rPAP I fusion protein and the native rPAP I. Finally, crosslinking experiments showed that rPAP I formed dimers in solution. These findings should be taken into account in functional studies involving PAP I and PAP-related proteins.

Acute-Phase Proteins↗