Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Rat calvaria bone cells isolated by collagenase digestion form a bone-like matrix which mineralizes in vitro in the presence of beta-glycerophosphate, in less than 2 weeks. The purpose of this work was to investigate, in this mineralizing rat osteoblastic cell culture, the synthesis of collagen, osteocalcin, and bone alkaline phosphatase (ALP). The results obtained indicate (1) After 15 days in culture, the extracellular-matrix contains collagen type I, V, and to some extent type III. Metabolic labeling at day 14, during the phase of nodules mineralization as well as new nodules formation, shows that collagen types I and type V are synthesized; (2) During the phase of cell growth, no osteocalcin could be detected in the medium, however, at the point of nodule formation, the osteocalcin level reached values of 3.55 +/- 1.39 ng/ml, followed by a 30-fold increase after nodules became mineralized. At day 14, after metabolic labeling, de novo synthesized osteocalcin was chromatographed on an immunoadsorbing column. With urea-SDS PAGE the apparent molecular weight was determined to be 9,000 daltons. (3) Specific activity of ALP was found to be 10 nmol/min/mg of proteins at cell confluence. At day 15, when nodules are mineralized, this activity was increased by 40-fold. The Michaelis constant was 1.58 10(-3) M/L. ALP was inhibited by L-homoarginine and levamisole but not by L-phenylalanine. ALP was shown to be heat sensitive at 56 degrees C with two slopes of inhibition.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Expression of collagen, osteocalcin, and bone alkaline phosphatase in a mineralizing rat osteoblastic cell culture. 137 88

To determine whether immune complex-like material is incorporated into the extracellular matrix (ECM) of proliferated RA synovium, cell-free matrices were isolated from pannus removed at joint replacement surgery, and were subjected to differential extraction. When the IgG and albumin concentrations in the ECM extracts were compared to those in simultaneously obtained synovial fluids, the IgG was found to be enriched 8.8-fold. Approximately 95% of the IgG was extractable with 6M Guanidine-HCl and 8 M Urea-B-ME. Further extraction with collagenase and low-pH buffers did not result in any additional recovery of IgG. Matrix-associated IgG demonstrated a restricted mobility on IEF with a pI of 4.8. The extracellular matrix of RA pannus is enriched in an acidic IgG species. Incorporation of IgG appears to be secondary to non-covalent interactions and may represent an additional reservoir of immune complex material in the rheumatoid joint.
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PMID:Synovial extracellular matrix II. Specific incorporation of immunoglobulin into the cell-free matrix of pannus. 139 21

Hydrolysis of arginine into urea and ornithine (Orn) was observed to take place in several segments of the rat nephron including cortical and medullary pars recta of the proximal tubule (PST) and collecting duct (CD). This work was now extended to the adult mouse and rabbit. Representative nephron segments, obtained by microdissection of collagenase-treated kidneys, were incubated with L-[guanido-14C]arginine (216 microM). Addition of urease produced 14CO2 + 2 NH3 from the newly formed urea released in the incubate. 14CO2 was trapped in KOH and counted. In both species, as well as in the rat, the PST was the site of the highest urea + Orn production, with an intensity increasing from cortex to medulla. For other nephron segments, the pattern was not similar in all species. Significant production of urea + Orn was observed in the proximal convoluted tubule and the medullary thick ascending limb in the rabbit, but not in the CD of either the rabbit or the mouse. The functional significance of this urea + Orn production remains unclear. The total amount of urea generated intrarenally by this reaction does not seem sufficient to play a significant role in the urinary concentrating mechanism. It may be assumed that Orn could be further metabolized to polyamines and play a role in maintaining cell integrity and function in the PST, especially in its medullary part, exposed to hypertonicity and poor oxygen supply.
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PMID:Localization of urea and ornithine production along mouse and rabbit nephrons: functional significance. 144 76

In HeLa cells transcription of the c-jun gene is activated strongly and rapidly by ultraviolet (UV) irradiation and, to a somewhat lesser extent, by treatment with phorbol ester tumor promoters. In the same cells UV and phorbol esters only marginally enhance the abundance of RNA transcribed from the jun D gene and from the gene coding for the serum response factor (which in turn acts on the UV and phorbol ester response element of the c-fos gene). In contrast to c-jun, jun B transcription is induced more efficiently by phorbol ester than by UV irradiation, suggesting that the members of the jun family are differently regulated. The promoter of c-jun carries two enhancer elements resembling AP-1 binding sites: the jun1 UV response element (URE-71 TGACATCA -64) and the jun2 URE (-190 TTACCTCA-183). These elements act independently in the UV induced expression of c-jun. In the context of the complete c-jun promoter they seem not to be required for c-jun induction by phorbol esters. When fused to the Herpes simplex thymidine kinase promoter, however, the isolated elements mediate induction by both UV and phorbol esters. UV and phorbol ester treatment of cells increases the binding of transcription factors to both elements. Both elements bind factors different in modification or/and constitution from AP-1, the heterodimeric transcription factor composed of c-Fos and c-Jun that controls the activity of the UV and phorbol ester response element (-72 TGAGTCA-66) of the human collagenase gene.
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PMID:Ultraviolet-radiation induced c-jun gene transcription: two AP-1 like binding sites mediate the response. 156 Dec 39

Periportal (pp) or perivenous (pv) liver parenchymal cells from female adult Uje: WIST rats were isolated after retro- or antegrade digitonin infusion followed by collagenase perfusion in the opposite direction. The morphological results revealed a distinct acinar-related destruction of the pv- or pp-zone by digitonin. The remaining cells of the respective other zone showed a good structural maintenance. After subsequent conventional collagenase perfusion the yield, viability and structural integrity of the isolated hepatocytes were high. The zonal cell separation was indicated by significant differences in the pp marker glucose-6-phosphatase and the pv marker glutamine synthetase found in the isolated pp or pv cell populations. Under our experimental conditions including the use of female rats, the alanine aminotransferase and glutamate dehydrogenase as well as ethylmorphine N-demethylase and ethoxycoumarin O-deethylase activities were evenly distributed in both preparations. Under stimulating conditions the capacity for urea synthesis was similar in both pv and pp cells.
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PMID:Biochemical and morphological studies on perivenous and periportal liver parenchymal cells from female rats isolated by digitonin-collagenase method. 168 Jul 46

The stimulation of alpha-1 adrenergic receptors in the mammalian nephron increases sodium reabsorption. In this study, alpha-1 adrenergic receptors in the inner medullary collecting duct (IMCD) cells were examined by radioligand binding technique. The IMCD cells were prepared from the rabbit kidney by incubating the inner medullary slices with collagenase and treating the isolated cells with hypotonic solution to lyse cells other than IMCD cells. The equilibrium binding of [3H]prazosin to IMCD cell homogenate was measured after incubation for 30 min at 25 degrees C in the absence (total binding) and the presence (nonspecific binding) of 100 microM phentolamine. The specific binding (the difference between total and nonspecific binding) of [3H]prazosin was saturable with a Bmax of 30 fmol/mg of protein and Kd of 0.9 nM. The displacement of [3H]prazosin binding to IMCD cells by adrenergic antagonists and agonists displayed the order of potency: beta-4-hydroxyphenyl-ethyl-amino-tetralone greater than phentolamine greater than naphazoline greater than epinephrine greater than yohimbine greater than norepinephrine greater than phenylephrine greater than propranolol. Because IMCD cells in the kidney have a hypertonic environment, the specific binding of [3H] prazosin to IMCD cells was also measured in a buffer that was made hypertonic (1200 mOsmol/kg of water) with NaCl and urea, the major solutes of the renal medulla. The hyperosmolality increased the Kd of [3H]prazosin to 5.2 mM without a change in its Bmax.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Alpha-1 adrenergic receptors in renal medullary collecting duct cells. 168 13

We investigated the steroid biosynthetic capabilities of ovarian granulosa and thecal elements of the viviparous dogfish, Squalus acanthias. In this report we present evidence that granulosa cells secrete quantitatively important amounts of progesterone (P), testosterone (T), and estradiol-17 beta (E), while theca has a more limited capacity to synthesize T and E. Ovarian granulosa cells were obtained from animals at each stage of gestation. After collagenase dispersion, an aliquot of 250,000 cells was incubated at 18 degrees C in basal medium, containing Eagle's salts, glutamine, penicillin, streptomycin and adjusted with 136 mM sodium chloride and 350 mM urea. After a 4 hour incubation, the content of P, T, and E in medium was determined by radioimmunoassay. P was not detectable at any time, while E was present throughout the cycle, being maximal when gestation is three quarters complete (Stage C). T gradually increased from Stage B toward late pregnancy. In Stage C granulosa cells, E production increased in the presence of graded doses of T substrate. Also, a homologous pituitary extract (1/25 equivalents) and the calcium ionophore A23187 stimulated production of all 3 steroids. Using radioisotopes, granulosa cells showed a wide range of synthetic capacities. In Stage C thecal tissue, E production also increased in the presence of graded doses of T substrate, while pituitary extract only increased T. When granulosa and theca were recombined, in the presence of pituitary extract, P levels decreased with a corresponding increase in T, when compared to granulosa alone. These data suggest a possible interaction between granulosa and theca for steroid biosynthesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Regulation of ovarian steroidogenesis in vitro in the viviparous shark, Squalus acanthias. 172 88

The Xenopus oocyte was evaluated as an mRNA expression system for water and urea transporters. Osmotic water permeability (Pf) was measured from the time course of oocyte volume in response to osmotic gradients using a real-time imaging method. Diffusional water permeability (Pd) was measured by 3H2O efflux. In mature oocytes treated with collagenase to remove the follicular cell layer, Pf was 8.6 +/- 0.6 x 10(-4) (SD) cm/s (n = 32) at 25 degrees C and independent of the time after oocyte removal (0-8 days). The activation energy (Ea) for Pf was 10.2 kcal/mol (10-32 degrees C). Pf was independent of osmotic gradient size (50-200 mosmol) in swelling experiments but decreased in an unpredictable manner in shrinking experiments. Pf was not altered by removal of the vitelline membrane but was decreased by 75% when the follicular cell layer was intact. In collagenase-treated oocytes, amphotericin (0-500 micrograms/ml) increased Pf from 8 x 10(-4) to 84 x 10(-4) cm/s in a dose-dependent manner. Pd was 3.4 +/- 0.2 x 10(-4) (SE) cm/s at 25 degrees C, 1.5 +/- 0.2 x 10(-4) cm/s at 4 degrees C, and 5.1 +/- 0.5 x 10(-4) cm/s at 25 degrees C in the presence of 500 micrograms/ml amphotericin; Ea was 6.5 kcal/mol. Thus Pd, but not Pf, is unstirred layer limited.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Water and urea permeability properties of Xenopus oocytes: expression of mRNA from toad urinary bladder. 198 78

High yields of human hepatocytes (up to 23 X 10(6) viable cells/g) were obtained from small surgical liver biopsies (1 to 3 g) by a two-step collagenase microperfusion method. Cell viability was about 95%, attachment efficiency of hepatocytes seeded on fibronectin-coated plates was 80% within 1 h after plating, and cells survived for about 2 wk in serum-free Ham's F12 containing 0.2% bovine serum albumin, 10(-8) M insulin, and 10(-8) M dexamethasone. To evaluate the metabolism of human hepatocytes in serum-free conditions, we measured their most characteristic biochemical functions and compared them to those reported for human liver. After 24 h in culture, glycogen content was 1250 +/- 177 nmol glucose/mg cell protein and remained stable for several days. Gluconeogenesis from lactate in hormone-free media was (3.50 +/- 0.17 nmol glucose.mg-1.min-1) similar to that reported for human liver. Insulin at 10(-8) M activated glycolysis (X1.40) and glycogenesis (X1.34), and glucagon at 10(-9) M stimulated gluconeogenesis (X1.35) and glycogenolysis (X2.18). Human hepatocytes synthesized albumin, transferrin, fibrinogen, alpha 1-antitrypsin, alpha 1-antichymotrypsin, alpha 1-acid glycoprotein, haptoglobin, alpha 2-macroglobulin, and plasma fibronectin and excreted them to the culture medium. Maximum protein synthesis was stimulated by 10(-9) M dexamethasone. Basal urea synthesis oscillated between 2.5 and 3.5 nmol.mg-1 cell protein.min-1, about 5 times the value estimated for human liver. Cytochrome P-450 decreased in culture but it was still 20% of freshly isolated hepatocytes by Day 5 in culture. In addition, ethoxycumarin-O-deethylase and aryl hydrocarbon hydroxylase could be induced in vitro by treatment with methyl cholanthrene. Glutathione levels were similar to those reported for human liver (35 nmol.mg-1). The results of our work show that adult human hepatocytes obtained from small surgical biopsies and cultured in chemically defined conditions express their most important metabolic functions to an extent that is similar to that reported for adult human liver.
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PMID:Culture of human hepatocytes from small surgical liver biopsies. Biochemical characterization and comparison with in vivo. 215 94

Regulation of urea transport by vasopressin in inner medullary collecting duct (IMCD) cells is thought to be important for the urinary concentrating mechanism. Isolated tubule perfusion studies suggest the existence of a saturable urea carrier. We have measured 14C-urea efflux in IMCD cells which were freshly isolated and grown in primary culture. Cells were isolated from rat papilla by collagenase digestion and hypotonic shock. In suspended cells, 14C-urea efflux (Jurea) from loaded cells was exponential with time constant 59 +/- 3 sec (SEM, n = 6, 23 degrees C). Jurea had an activation energy of 4.1 kcal/mole and was inhibited 42 +/- 7% by 0.25 mM phloretin and 30-40% by the high affinity urea analogues dimethylurea and phenylurea. Jurea was increased 40-60% by addition of vasopressin (10(-8) M) or 8-bromo-cAMP (1 mM); stimulated Jurea was inhibited 55 +/- 8% by the kinase A inhibitor H-8. Phorbol esters and epidermal growth factor did not alter Jurea. IMCD cells grown in primary culture were homogeneous in appearance with greater than fivefold stimulation of cAMP by vasopressin. The exponential time constant for urea efflux was 610 +/- 20 sec (n = 3). Jurea was not altered by vasopressin, cAMP or phloretin. Another function of in vivo IMCD cells, vasopressin-dependent formation of endosomes containing water channels, was absent in the cultured cells. These results demonstrate presence of a urea transporter on suspended IMCD cells which is activated by cAMP and inhibited by phloretin and urea analogues. The urea transporter and its regulation by cAMP, and cAMP-dependent apical membrane endocytosis, are lost after growth in primary culture.
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PMID:Urea transport in freshly isolated and cultured cells from rat inner medullary collecting duct. 217 46


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