Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Retinal pigment epithelium plasma membranes have been isolated by differential and density gradient centrifugation of glass-bead-bound, collagenase-treated cells. Electron microscopic evidence indicates that the glass-bead-bound cells were devoid of red blood cells, rod outer segments and other ocular cell contaminants. The plasma membranes were recovered in 4-6 micrograms/eye yields and purified 10-fold by 5'-nucleotidase and alkaline phosphodiesterase I, and 6.5-fold by (Na+ + K+)-ATPase. Plasma membrane purity as measured by covalent labeling of the epithelial cell plasma membrane proteins with p-(diazonium) benzene[32S]sulfonic acid was 8-19-fold. In purified plasma membranes contamination by mitochondria was undetectable and lysosomal contamination reduced 100-fold, while endoplasmic reticulum was 2-fold enriched. SDS-polyacrylamide gel electrophoresis of the plasma membrane proteins revealed 23-26 major bands by Coomassie blue staining and 12-16 major bands by radioactive labeling. The plasma membranes exhibited a 3-fold lower concentration of docosahexaenoic acid, a 3-fold higher cholesterol/phosphate ratio, and were 10-fold enriched in cholesterol per micrograms protein when compared to the whole cell fraction. Retinal epithelial plasma membranes contain an average of 1 mol cholesterol per mol of lipid phosphorus, a high palmitic acid concentration (39 mol%) and a low concentration of docosahexaenoic acid (2 mol%). The lipid profile of the retinal pigment epithelial plasma membranes indicates that they are typical of plasma membranes from many other cell types and that they appear to be less fluid than total rod outer segment membranes.
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PMID:Isolation of plasma membranes from the bovine retinal pigment epithelium. 298 2

Ionic fluoride is taken up by rabbit synovial fibroblasts in culture. The uptake was accompanied by an increased production of latent collagenase, and decreased activity of 5'-nucleotidase.
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PMID:Fluoride ions increase collagenase production by rabbit synovial fibroblasts. 298 24

Cholesteryl esterase activities were determined in homogenates of rat heart (ventricles), isolated, calcium-tolerant, cardiac myocytes and aortic tissue and were compared with acid and neutral triglyceride lipase activities in these fractions. Using cholesteryl oleate/phosphatidylcholine/taurocholate emulsions and digitonin pretreatment of the enzyme fractions, acid and neutral cholesteryl esterase activities were measured in all tissue preparations. In contrast to the acid and neutral triglyceridase and acid cholesteryl esterase activity, the neutral cholesteryl esterase activity was subject to substrate inhibition. Upon isolation of cardiac myocytes, and in contrast with the recovery of neutral triglyceride lipase activity, only a small portion of the neutral cholesteryl esterase (6%) was recovered, suggesting that nonmyocyte neutral cholesteryl esterase activity markedly contributes to the relatively high activity detectable in whole ventricular homogenates. The recovery of large amounts of neutral cholesteryl esterase activity in the supernatant of collagenase-digested heart tissue, obtained during the isolation of myocytes, which is also markedly enriched in activities of two endothelial marker enzymes (5'-nucleotidase and angiotensine-converting enzyme) may indicate the predominant contribution of neutral cholesteryl esterase activity from coronary endothelial cells to this activity detectable in ventricular homogenates. Relative to the activity in ventricular and myocyte homogenates, aorta homogenates possessed the highest specific neutral cholesteryl esterase activity. We propose that in addition to coronary endothelium, smooth muscle cells also contribute to the neutral cholesteryl esterase activity in ventricular homogenates.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Cholesteryl esterase activities in ventricles, isolated heart cells and aorta of the rat. 303 10

Sperm maturation and storage occur in a unique milieu created in large part by the epididymal epithelium. To learn more about the interaction of the epididymal epithelial cell with both luminal and systemic environments, we now report on the preparation and characterization of epididymal epithelial cell plasma membranes. A preparation enriched for epididymal epithelial cell plasma membranes was isolated from collagenase-digested epididymal tubule fragments by hand-Dounce homogenization, differential centrifugation, and sucrose gradient centrifugation. The final membrane fraction was enriched 11-fold for the plasma membrane marker 5'-nucleotidase; 2.6-fold for the lysosomal marker acid phosphatase, and 3-fold for the Golgi marker thiamine pyrophosphatase. No enrichment was observed for mitochondrial or endoplasmic reticulum enzyme markers. Specific and saturable transferrin-binding activity was also detected in the final preparation. Electron microscopy revealed the presence of vesicles and sheets of membranes as well as an occasional Golgi apparatus. The plasma membrane fraction was used to generate monoclonal antibodies. Of 102 wells exhibiting growth, 12 were positive by immunofluorescent staining of frozen sections. Ten of these recognized determinants in epithelial cells, and 2 stained peritubular smooth muscle cells. Most of the epithelial cell-specific antibodies stained brush border alone or in combination with the basolateral plasma membrane. Three antibodies stained the Golgi apparatus. Most antibodies were specific for particular epididymal regions, 3 also recognized determinants in the kidney, and 1 stained residual bodies in the testis.
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PMID:Isolation and characterization of epididymal epithelial cell plasma membranes. 336 69

1. A collagenase digest of rat kidney cortex was separated into four bands by zonal centrifugation. 2. Two of these bands were shown by light-microscopy to contain glomeruli and tubular fragments, which were free from each other and well separated from other renal material. 3. Protein, N-acetyl-beta-glucosaminidase, 5'-nucleotidase, l-leucine beta-naphthylamidase, leucine aminopeptidase, acid phosphatase and alkaline phosphatase were assayed across the gradient. 4. The greater proportion of these enzyme activities was recovered in the tubular fragments and acid phosphatase was the only enzyme detected in significant amounts in the glomeruli. 5. Tubular fragments and glomeruli were sedimented and multiple forms of beta-naphthylamidase, N-acetyl-beta-glucosaminidase, acid phosphatase and alkaline phosphatase were investigated by starch-gel electrophoresis.
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PMID:The distribution of some hydrolases in glomeruli and tubular fragments prepared from rat kidney by zonal centrifugation. 433 86

Smooth muscle cells were dissociated from normal rabbit aorta by incubating the tissue in Hanks' solution containing elastase, collagenase, and hyaluronidase. The isolated cells contained significant amounts of the following acid hydrolases: N-acetyl-beta-glucosaminidase, N-acetyl-beta-galactosaminidase, beta-galactosidase, beta-glucuronidase, alpha-mannosidase, beta-glucosidase, acid phosphatase, and cathepsins C and D. The cells were disrupted and fractionated by isopycnic centrifugation on sucrose density gradients in the Beaufay automatic zonal rotor. Lysosomes with a modal density of 1.16 were identified by the distribution of these acid hydrolases and by the latency of N-acetyl-beta-glucosaminidase and beta-galactosidase. Other particulate enzymes studied in these sucrose gradients included cytochrome oxidase and monoamine oxidase (mitochondria), 5'-nucleotidase and leucyl-beta-naphthylamidase (plasma membrane), and catalase (? peroxisome). This microanalytical subcellular fractionation technique is applicable to the study of milligram quantities of many other tissues, both normal and pathological.
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PMID:Lysosomes of the arterial wall. I. Isolation and subcellular fractionation of cells from normal rabbit aorta. 434 42

An attempt was made to concentrate plasma membranes of homogenized chondrocytes isolated by collagenase digestion of rachitic rat epiphyseal growth plate cartilage. This study reports the characterization of enzymes in the plasma membrane of isolated chondrocytes and their comparison with extracellular matrix vesicle components. The plasma membrane-enriched fractions that were obtained showed a sevenfold increase in 5'-nucleotidase and a 15-fold increase in alkaline phosphatase, both of which are regarded as plasma membrane markers. SDS-polyacrylamide gel electrophoretic profiles of proteins extracted from membrane fractions contained several major protein bands also seen in isolated matrix vesicles. These studies indicate the usefulness of concentrating plasma membrane components from isolated chondrocytes, after the chondrocytes have been enzymatically freed from investing matrix and other stromal components by collagenase.
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PMID:Isolation of a plasma membrane-enriched fraction from collagenase-suspended rachitic rat growth plate chondrocytes. 609 Jun 23

Microvessels were isolated from rat brain using a double collagenase treatment which removed the endothelial basement membranes. The isolate was characterized by intact luminal and abluminal membranes and an absence of pericytes and astrocyte membranes. Minimal contamination by 5'-nucleotidase, an enzyme believed exclusively localized within the plasma membranes of neuroglia, established the purity of the isolated microvessels. Enrichment of alkaline phosphatase and gamma-glutamyl transpeptidase activity in microvessel preparations supports the endothelial localization of these enzymes.
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PMID:Isolation and characterization of brain endothelial cells: morphology and enzyme activity. 610 94

Rat adipose tissue was digested with collagenase and separated into adipocytes and stromal-vascular cells. The adipocytes accounted for 40% of the total adipose tissue adenosine deaminase activity, 32% of 5'-nucleotidase activity and 87% of adenosine kinase activity. This distribution suggests that adipocyte are the major cell type involved in adenosine utilization in adipose tissue. Furthermore, it suggests that the high sensitivity of isolated adipocytes to adenosine is representative of their sensitivity of isolated adipocytes to adenosine is representative of their sensitivity in vivo.
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PMID:Distribution of adenosine metabolising enzymes between adipocyte and stromal-vascular cells of adipose tissue. 626 98

Liver parenchymal cells from adult rats were isolated by treatment with collagenase and cultured as monolayers in Williams medium E with 10% fetal or calf serum. The additions of dexamethasone and insulin to the medium were essential for maintaining liver functions of the cells. These cells synthesized and secreted various serum proteins into the medium. Gluconeogenesis and glycogenolysis were enhanced by glucagon, and lipogenesis was stimulated by insulin. Many enzymes were also induced by various hormones. These activities were very low in freshly isolated cells, but were restored when the cells were cultured for a few days. Markers of plasma membranes, such as 5'-nucleotidase and insulin receptors, were reduced to half the normal levels on freshly isolated cells, but they were restored to the normal levels during culture of the cells without added hormones. Analysis of the profile of amino acids in the medium showed that freshly isolated cells were in a catabolic state of protein turnover and released branched chain amino acids into the medium, but that cultured cells consumed amino acids, not only for protein synthesis, but also for other metabolic processes, such as gluconeogenesis. These findings show that freshly isolated cells have impaired functions and are unsuitable for use in studies of liver metabolism, but that after culture for a few days the cells regain the activity of normal liver and hance become useful for studies of liver functions. Studies with these cells are simpler and give clear results than studies in vivo.
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PMID:Biochemical functions of adult rat hepatocytes in primary culture. 693 75


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