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Journal of Lipid Research May 1979With the limited stirring procedure used in the present work, substrate and enzyme together form a segregated and well-defined system on the surface. The lipase...
With the limited stirring procedure used in the present work, substrate and enzyme together form a segregated and well-defined system on the surface. The lipase molecules responsible for the lipolysis are only those that are adsorbed on the glyceride monolayer. After a study of the stirring procedure, two series of systematic experiments were done: a) the bulk concentration of the enzyme was varied with different constant surface concentrations of the substrate, and b) the surface concentration of the substrate was varied with different constant bulk concentrations of the enzyme. The influence of the surface concentration of substrate on a) the rate of lipolysis, V,; b) the enzyme activity, a,; and c) the enzyme adsorption, Ze, were each determined by a different procedure. The values obtained verify the enzymic activity equation (a = V/Ze). The roles of other factors (Ca2+ ions, and pH) which govern the adsorption of the enzyme and its specific activity were also studied in preliminary experiments.
Topics: Adsorption; Animals; Diglycerides; Glycerides; Kinetics; Lipase; Mathematics; Membranes, Artificial; Pancreas; Protein Binding; Surface Properties; Swine
PubMed: 458267
DOI: No ID Found -
The Journal of Cell Biology Oct 1970The origin, morphogenesis, and biochemical differentiation of the dorsal and ventral pancreas of the rat embryo have been investigated in order to ascertain the...
The origin, morphogenesis, and biochemical differentiation of the dorsal and ventral pancreas of the rat embryo have been investigated in order to ascertain the similarities and dissimilarities between the two lobes. We have utilized a culture system in which the primitive gut gives rise to a number of differentiated organs, including the dorsal and ventral pancreas. The two pancreases do not undergo fusion in these cultures, thus allowing independent analyses of the two lobes for comparison with in vivo results. The dorsal pancreas first appeared at the 23-25 somite stage while the ventral pancreas appeared approximately 12 hr later at the 29-30 somite stage. Guts from embryos as young as 12 somites were capable of developing both pancreases in vitro. In spite of the 12 hr difference between the times of their appearance, the dorsal and ventral pancreases exhibited identical patterns of morphological and biochemical differentiation. The two lobes contained the same exocrine enzymes and hormones, at similar levels, differing only in their glucagon content, the dorsal pancreas possessing a fivefold higher glucagon specific activity. The implications of these results are discussed.
Topics: Amylases; Animals; Carboxypeptidases; Cell Differentiation; Chymotrypsin; Culture Media; Culture Techniques; Endoderm; Enzyme Precursors; Female; Glucagon; Histocytochemistry; Insulin; Male; Microscopy, Electron; Morphogenesis; Pancreas; Rats; Ribonucleases; Time Factors
PubMed: 5513553
DOI: 10.1083/jcb.47.1.235 -
Cell Transplantation 2009The available information concerning the characteristics and composition of collagenase batches, which are effective in the digestion of human pancreas for islet...
The available information concerning the characteristics and composition of collagenase batches, which are effective in the digestion of human pancreas for islet transplants, is scarce and incomplete. A large inter- and intrabatched variability in activity and efficiency of blend enzymes available for isolation has been observed. The aim of this study was to characterize enzyme blend components. Liberase batches were characterized by SDS-PAGE analyses, microelectrophoresis, and then by MALDI-TOF MS analysis. Three main bands were detected by SDS-PAGE analysis and submitted to MALDI-TOF MS analysis. Two bands were found to correspond to class I (isoform beta and another of 106 kDa) and one to class II (isoform delta) collagenase. These results represent an important step towards a complete characterization of enzymes, with the final aim of identifying key components for a standardized product.
Topics: Collagenases; Electrophoresis, Polyacrylamide Gel; Humans; Isoenzymes; Pancreas; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Thermolysin
PubMed: 19499708
DOI: 10.3727/096368909788341270 -
Shock (Augusta, Ga.) Aug 2012In bowel ischemia, impaired mucosal integrity may allow intestinal pancreatic enzyme products to become systemic and precipitate irreversible shock and death. This can...
In bowel ischemia, impaired mucosal integrity may allow intestinal pancreatic enzyme products to become systemic and precipitate irreversible shock and death. This can be attenuated by pancreatic enzyme inhibition in the small-bowel lumen. It is unresolved, however, whether ischemically mediated mucosal disruption is the key event allowing pancreatic enzyme products systemic access and whether intestinal digestive enzyme activity in concert with increased mucosal permeability leads to shock in the absence of ischemia. To test this possibility, the small intestinal lumen of nonischemic rats was perfused for 2 h with either digestive enzymes, a mucin disruption strategy (i.e., mucolytics) designed to increase mucosal permeability, or both, and animals were observed for shock. Digestive enzymes perfused included trypsin, chymotrypsin, elastase, amylase, and lipase. Control (n = 6) and experimental animals perfused with pancreatic enzymes only (n = 6) or single enzymes (n = 3 for each of the five enzyme groups) maintained stable hemodynamics. After mucin disruption using a combination of enteral N-acetylcysteine, atropine, and increased flow rates, rats (n = 6) developed mild hypotension (P < 0.001 compared with groups perfused with pancreatic enzymes only after 90 min) and increased intestinal permeability to intralumenally perfused fluorescein isothiocyanate-dextran 20 kd (P < 0.05) compared with control and enzyme-only groups, but there were no deaths. All animals perfused with both digestive enzymes and subjected to mucin disruption (n = 6) developed hypotension and increased intestinal permeability (P < 0.001 after 90 min). Pancreatic enzymes were measured in the intestinal wall of both groups subjected to mucin disruption, but not in the enzyme-only or control groups. Depletion of plasma protease inhibitors was found only in animals perfused with pancreatic enzymes plus mucin disruption, implicating increased permeability and intralumenal pancreatic enzyme egress in this group. These experiments demonstrate that increased bowel permeability via mucin disruption in the presence of pancreatic enzymes can induce shock and increase systemic protease activation in the absence of ischemia, implicating bowel mucin disruption as a key event in early ischemia. Digestive enzymes and their products, if allowed to penetrate the gut wall, may trigger multiorgan failure and death.
Topics: Animals; Blotting, Western; Enzymes; Expectorants; Hemodynamics; Immunohistochemistry; Intestine, Small; Male; Pancreas; Pancreatic Elastase; Peptide Hydrolases; Permeability; Rats; Rats, Wistar; Shock; Trypsin
PubMed: 22576000
DOI: 10.1097/SHK.0b013e31825b1717 -
JOP : Journal of the Pancreas Sep 2012Porcine pancreatic enzymes (PPE) extracted from glandular stomach has been used for the treatment of pancreatic cancer patients. Unfortunately, no information is...
CONTEXT
Porcine pancreatic enzymes (PPE) extracted from glandular stomach has been used for the treatment of pancreatic cancer patients. Unfortunately, no information is available on the in vitro and in vivo effect on the pancreas and other tissues.
OBJECTIVE
We used Syrian Golden hamsters, a unique pancreatic cancer model, to obtain basic information on PPE for its eventual use for the treatment of pancreatic cancer.
DESIGN
PPE was used in different concentrations in vitro and in vivo. The stability of the enzyme in the water solution was investigated. It was given to the hamsters by gavage in concentrations of 1g/kg and 400 mg/kg for short periods and in aqueous solution for 65 days. Plasma enzyme and insulin, the size of islets and the number of the insulin cells per islet were examined.
RESULTS
The enzyme activity of PPE was maintained in water solution for at least 24 hours. Due to its content of calcium chloride it showed a high toxicity to normal and malignant hamster pancreatic cancer cells and human pancreatic cancer cell lines in vitro. PPE did not alter the plasma pancreatic enzyme levels regardless of the dose, duration and application route. On the contrary, PPE reduced their levels significantly. Remarkably, it also reduced the level of insulin, the size of the islets and the number of insulin cells in the islets significantly.
CONCLUSION
The results imply that PPE does not enter the blood circulation but it appears to slow down the function of both the exocrine and endocrine pancreas.
Topics: Amylases; Animals; Calcium Chloride; Cell Count; Cell Line; Cell Line, Tumor; Cricetinae; Dose-Response Relationship, Drug; Female; Glucagon; Humans; Immunoassay; Immunohistochemistry; Insulin; Insulin Secretion; Islets of Langerhans; Lipase; Male; Mesocricetus; Necrosis; Pancreas; Pancreatic Neoplasms; Pancreatin; Swine; Trypsin
PubMed: 22964953
DOI: 10.6092/1590-8577/522 -
International Journal of Pancreatology... Apr 1998Adaptive changes in exocrine and endocrine pancreatic function, as well as changes in pancreas size and morphology, were not observed after 4-wk of oral pancreatic... (Clinical Trial)
Clinical Trial Randomized Controlled Trial
CONCLUSIONS
Adaptive changes in exocrine and endocrine pancreatic function, as well as changes in pancreas size and morphology, were not observed after 4-wk of oral pancreatic enzyme application. These findings suggest that the normal pancreas does not significantly adapt--either morphologically or functionally--to a 4-wk oral application of high-dose pancreatic enzymes.
BACKGROUND
The control of exocrine pancreatic enzyme secretion is not completely understood. Although it has been established that exocrine pancreatic secretion is mainly regulated in the short-term by the amount of pancreatic enzymes in the proximal small intestine, it is not known whether long-term application of pancreatic enzymes causes changes in exocrine pancreatic secretion in humans.
METHODS
Twelve healthy male volunteers (median age 27 yr) participated in a prospective, randomized, placebo-controlled, double-blind study. Six were placed in the treatment group and six in the placebo group. Over a 4-wk period, the six subjects in the treatment group took 18 capsules of Panzytrat (20,000 units of lipase, 18,000 units of amylase, and 1000 units of protease per capsule) daily. Before (wk 0), 4 wk following pancreatic enzyme application and 2 wk afterward, a secretin-cerulein test was carried out in all subjects to study exocrine pancreatic function (trypsin, chymotrypsin, bicarbonate content, and total pancreatic fluid secretion in the duodenum). One day following the secretin-cerulein test, a standard test meal was given to all subjects to analyze endocrine pancreatic function. Additionally, before starting the treatment, once per week during treatment and 2 wk afterward, an ultrasound examination of the pancreas was carried out to see whether there was any change in pancreas size and morphology.
RESULTS
Trypsin content in the duodenal aspirates following simultaneous stimulation with secretin and cerulein after 4 wk of high-dose pancreatic enzyme application was 92% in the treatment group and 82% in the placebo group compared with the wk 0 test results (100%). Two weeks after enzyme application, the secretin/cerulein-stimulated trypsin content was 88% in the treatment group and 107% in the placebo group. None of these changes was statistically significant. The same results were seen for chymotrypsin content, amylase, and bicarbonate content as well as for total pancreatic fluid secretion. Additionally, no change in the endocrine pancreatic function could be observed after 4 wk of pancreatic enzyme treatment. Pancreas ultrasonography revealed no alteration in pancreas size or parenchymal structure during the 4 wk of treatment and the following 2 wk.
Topics: Adaptation, Physiological; Adult; Amylases; Dose-Response Relationship, Drug; Double-Blind Method; Drug Combinations; Endopeptidases; Gastrointestinal Agents; Humans; Islets of Langerhans; Male; Pancreas; Pancreatin; Prospective Studies; Reference Values
PubMed: 9629509
DOI: 10.1385/IJGC:23:2:115 -
Pharmacology & Toxicology Dec 2002Cholecystokinin (CCK) acting through its G protein-coupled receptor is now known to activate a variety of intracellular signaling mechanisms and thereby regulate a... (Review)
Review
Cholecystokinin (CCK) acting through its G protein-coupled receptor is now known to activate a variety of intracellular signaling mechanisms and thereby regulate a complex array of cellular functions in pancreatic acinar cells. The best studied mechanism is the coupling through heterotrimeric G proteins of the Gq family to activate a phospholipase C leading to an increase in inositol trisphosphate and release of intracellular Ca2+. This pathway along with protein kinase C activation in response to the increase in diacylglycerol stimulates the secretion of digestive enzymes by the process of exocytosis. CCK also activates signaling pathways in acini more related to other processes. The three mitogen activated protein kinase cascades leading to ERKs, JNKs and p38 MAPK are all activated by CCK. CCK activates the ERK cascade by PKC activation of Raf which in turn activates MEK and ERKs. JNKs are activated by a distinct mechanism which requires higher concentrations of CCK. Both ERKs and JNKs are presumed to regulate gene expression. CCK activation of p38 MAPK also plays a role in regulating the actin cytoskeleton through phosphorylation of the small heat shock protein HSP27. The PI3K-PKB-mTOR pathway is activated by CCK and plays a major role in regulating protein synthesis at the translational level. This includes both activation of p70 S6K leading to phosphorylation of ribosomal protein S6 and the phosphorylation of the binding protein for initiation factor 4E leading to formation of the mRNA cap binding complex. Other signaling pathways activated by CCK receptors include NF-kappaB and a variety of tyrosine kinases. Further work is needed to understand how CCK receptors activate most of the above pathways and to better understand the biological events regulated by these diverse signaling pathways.
Topics: Animals; Calcium; Cholecystokinin; GTP-Binding Proteins; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Pancreas; Receptors, Cholecystokinin; Rodentia; Signal Transduction
PubMed: 12688372
DOI: 10.1034/j.1600-0773.2002.910606.x -
The Yale Journal of Biology and Medicine 1992This paper reviews the relationships between the effects of glucocorticoids on rat pancreatic acinar AR42J cell polyamine levels and cellular growth and differentiation.... (Review)
Review
This paper reviews the relationships between the effects of glucocorticoids on rat pancreatic acinar AR42J cell polyamine levels and cellular growth and differentiation. Glucocorticoids inhibit the growth of AR42J cells. Glucocorticoids either stimulate or inhibit the formation of polyamines in a variety of cell types. Cells require polyamines for normal growth. Therefore, we tested the hypothesis that polyamines mediate the effects of glucocorticoids on AR42J cells. First, to confirm that AR42J cells required polyamines for growth we examined the effects of inhibiting ornithine decarboxylase (ODC). ODC is the most important and generally rate-limiting enzyme in the synthesis of the polyamines. As expected, the ODC inhibitor difluoromethylornithine (DFMO) inhibited AR42J cell DNA synthesis, and the addition of exogenous putrescine reversed this effect. The levels of growth inhibition by glucocorticoids and DFMO treatment were similar. Second, we examined the effects of glucocorticoids on ODC. Surprisingly, glucocorticoids increased levels of AR42J cell ODC mRNA, ODC activity, and putrescine. Glucocorticoids increased these parameters over a similar time-course as they decreased DNA synthesis. Analog specificity studies indicated that a glucocorticoid receptor mediated both the growth inhibitory and ODC stimulatory effects. Dose-response studies indicated, however, that growth inhibition was more sensitive to dexamethasone (DEX) than were ODC levels. Therefore, polyamines do not account for the effects of glucocorticoids on AR42J cell growth. In these cells, glucocorticoids have opposite and independent effects on ODC and growth.
Topics: Animals; Cell Division; Dexamethasone; Ornithine Decarboxylase; Pancreas; Rats; Tumor Cells, Cultured
PubMed: 1340062
DOI: No ID Found -
The Journal of Biological Chemistry Oct 1978When pancreatic DNase I is used as a specific biochemical reagent in the preparation of nuclear ribonucleic acids or nuclear proteins, freedom from contaminating...
When pancreatic DNase I is used as a specific biochemical reagent in the preparation of nuclear ribonucleic acids or nuclear proteins, freedom from contaminating ribonucleases or proteases is an important property of the enzyme preparation. A simple one-step procedure has been developed to effect complete removal of trypsin, chymotrypsin, and chymotrypsinogen by a combination of affinity chromatography and salting-out adsorption on lima bean protease inhibitor coupled to Sepharose (a column (0.9 X 60 cm) operated in series with a regeneratable 1-ml bed). Commercial preparations of DNase (about 10 mg) give a quantitative yield of the enzyme that is protease-free as evidenced by full stability for more than 10 days at pH 8 and 37 degrees C even in the absence of the protecting action of Ca2+. Removal of the last traces of RNase has been accomplished by affinity chromatography on a column (0.4 X 72 cm) of 5'-(4-aminophenyl-phosphoryl)-uridine 2'(3')-phosphate-Sepharose; the product is a highly active DNase that gives no detectable hydrolysis of RNA by assay on radioactive substrates.
Topics: Chromatography, Affinity; Deoxyribonucleases; Pancreas; Peptide Hydrolases; Ribonucleases
PubMed: 701244
DOI: No ID Found -
BMB Reports Oct 2009Porcine pancreas development is not well studied at the molecular level despite being a therapeutic resource for diabetic patients. In this study, we investigated...
Porcine pancreas development is not well studied at the molecular level despite being a therapeutic resource for diabetic patients. In this study, we investigated expression of lineage markers and performed proteomic analysis. Expression of the early lineage markers Pdx1 and Ptf1a was developmentally conserved between mice and pigs, whereas expression of the islet differentiation marker Pax4 was delayed in porcine compared with murine pancreas development. Proteomic analysis found that expression levels of chymotrypsinogen were downregulated during porcine pancreas development while those of digestive enzymes like lipases, elastase and serine protease were up-regulated. In addition, specific isoforms of protein folding assistants such as protein disulfide isomerase and prefoldin were expressed at specific stages during the maturation of digestive enzymes. Taken together, these results show that development of the porcine pancreas is regulated by a concerted interplay of pancreas lineage marker proteins and other specified proteins, resulting in a functional endocrine and exocrine organ.
Topics: Animals; Biomarkers; Cell Lineage; Electrophoresis, Gel, Two-Dimensional; Enzyme Inhibitors; Enzyme Precursors; Pancreas; Protein Folding; Proteins; Proteomics; Sus scrofa
PubMed: 19874711
DOI: 10.5483/bmbrep.2009.42.10.661