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)

The metabolism and DNA binding of acetylaminofluorene (AAF) was investigated in human hepatocytes that were isolated from donor liver tissue by collagenase perfusion. Hepatocytes were treated with 0.01 microM pentachlorophenol (PCP), as a sulfotransferase inhibitor, to investigate the role of sulfotransferase in human bioactivation of aromatic amines. Concentrations of PCP greater than 0.1 microM resulted in cytotoxicity as noted by detachment of cells and atypical morphology. The metabolites of AAF were identified by HPLC as aminofluorene, 7-OH-AAF, 9-OH-AAF, 5-OH-AAF, N-OH-AAF, 1-OH-AAF and 3-OH-AAF. No consistent alteration in the metabolites produced occurred with PCP treatment compared to controls. PCP treatment increased total DNA binding of AAF metabolites compared with controls, suggesting that sulfotransferase does not activate AAF in human hepatocytes. Inhibition of sulfotransferase in human hepatocytes does not decrease DNA binding of AAF metabolites as noted previously with rat hepatocytes. Therefore, PCP may inhibit a detoxication pathway. This study supports N,O-acyltransferase as the critical enzyme for the formation of the major reactive metabolite in human liver.
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PMID:Inhibition of sulfotransferase in primary cultures of human hepatocytes affecting metabolism and binding of 2-acetylaminofluorene. 161 93

The sulfate and glucuronide conjugation of acetaminophen (APAP) by hepatocytes cultured on Matrigel or type 1 collagen was compared to APAP metabolism in vivo. The metabolic fate of low (15 mg/kg), medium (125 mg/kg), and high (300 mg/kg) doses of APAP injected intraperitoneally were determined in male and female rats. Males excreted more APAP as the sulfate conjugate than females, which correlated with the twofold greater APAP sulfotransferase activity in the male vs. females (301 +/- 24 vs. 156 +/- 18 pmol.mg-1 protein.min-1). Also, as sulfate conjugation became saturated, there was a dose-related shift in APAP metabolism from sulfate to glucuronide conjugation in both sexes. After death, the livers of the same animals were perfused with collagenase and the hepatocytes cultured in modified Waymouth's medium on either Matrigel or rat-tail collagen, with various doses of APAP (0, 0.125, 0.25, 0.5, and 1.0 mM). Sex differences in APAP sulfation and glucuronidation persisted in culture for up to 4 days, with sulfation predominating in the male similar to in vivo. With increasing APAP concentration (dose), there was a saturation of sulfate conjugation and a shift to glucuronidation as observed in vivo. Sex differences in APAP sulfation and glucuronidation were no longer significant by Day 4 in culture. Sulfation, and to a lesser extent, glucuronidation, were more stable on Matrigel than collagen. We concluded that APAP metabolism of freshly isolated hepatocytes could replicate in vivo sex differences in conjugation, and that Matrigel was superior to collagen as substrate.
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PMID:Sulfation and glucuronidation of acetaminophen by cultured hepatocytes reproducing in vivo sex-differences in conjugation on Matrigel and type 1 collagen. 175

Freshly isolated and preserved rat liver parenchymal cells were used as metabolizing system in the Salmonella mutagenicity assay. The liver cells were isolated with EDTA perfusion without the addition of collagenase and had a viability of 96% as judged by trypan blue exclusion. When freshly isolated liver parenchymal were cryopreserved with a computer controlled freezing protocol and stored at -196 degrees C they had a mean viability of 89% after thawing. Furthermore, freshly isolated cells were stored at 0 degree C in University of Wisconsin organ transplantation solution. After 1 day of hypothermic storage they had a viability of 95%. Four different indirect mutagens, 2-aminoanthracene, benzo[a]pyrene, 7,12-dimetylbenz[a]anthracene and cyclophosphamide, were used with the liver cells as metabolizing system in the preincubation assay with Salmonella typhimurium TA100. After cryopreservation, liver parenchymal cells were able to activate all tested indirect mutagens to ultimate mutagens. However, the induction of revertants was lower with three of the four tested compounds. Only 2-aminoanthracene was activated to the same extent by freshly isolated and cryopreserved liver cells. 7-Hydroxymethyl-12-methylbenz[a]anthracene, which is activated to its ultimate mutagen by sulfotransferase, also induced a reduced mutagenic effect with cryopreserved liver cells in comparison to freshly isolated liver parenchymal cells. This indicates that phase I and phase II enzyme activities are effected by cryopreservation. However, identical mutation frequencies were obtained when freshly isolated liver parenchymal cells or 1 day hypothermically preserved liver parenchymal cells were used in the cell-mediated Salmonella mutagenicity test. The use of hypothermic short-time storage of liver parenchymal cells could help to make the liver cell-mediated genotoxicity test simpler and thereby more attractive.
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PMID:Cryopreserved and hypothermically stored rat liver parenchymal cells as metabolizing system in the Salmonella mutagenicity assay. 852 46

To determine whether the dexamethasone (DEX)-inducible hepatic sulfotransferase gene expression that has been described in the rat is conserved in humans, the effects of DEX treatment on hydroxysteroid sulfotransferase (SULT2A1) and aryl sulfotransferase (SULT1A1) gene expression were investigated in primary cultured human hepatocytes. Hepatocytes were prepared from nontransplantable human livers by collagenase perfusion of the left hepatic lobe, and cultured in Williams' medium E that was supplemented with 0.25 U/ml insulin. As reported in the rat, DEX treatment produced concentration-dependent increases in SULT2A1 mRNA and protein expression, with maximum increases observed at concentrations of DEX that would be expected to activate the pregnane X receptor (PXR) transcription factor. In contrast to the rat, in which DEX-inducible SULT1A1 expression has been demonstrated, SULT1A1 expression in primary cultured human hepatocytes was not measurably increased by DEX. In transient transfections conducted in primary cultured rat hepatocytes, the PXR ligands DEX and pregnenolone-16 alpha-carbonitrile significantly induced transcription of human and rat SULT2A reporter gene constructs. Cotransfection of either the human or rat SULT2A reporter gene with a PXR dominant negative construct significantly reduced DEX-inducible transcription. These results underscore that while certain features of rat hepatic sulfotransferase gene regulation are conserved in humans, important differences exist across species. The findings also implicate a role for the PXR transcription factor in DEX-inducible rat and human SULT2A gene expression.
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PMID:Effects of dexamethasone on aryl (SULT1A1)- and hydroxysteroid (SULT2A1)-sulfotransferase gene expression in primary cultured human hepatocytes. 1216 65

Proximal-rich tubules were prepared from rat kidneys by using collagenase treatment. The isolated rat renal tubules were compared with the intact kidney on the following characteristics. (1) Composition of the sulfoglycolipid. (2) Sulfoglycolipid metabolism based on incorporation of [35S]sulfate or some properties of sulfoglycolipid metabolism, including the activities of anabolic and catabolic enzymes. The results indicated following characteristics of the isolated renal tubules in comparison to the kidney in vivo. (1) The sulfoglycolipid compositions are qualitatively similar, except that the content of glucosyl sulfatide, Gg3Cer II3-sulfate, and GM4 was slightly higher in the isolated tubules. (2) The apparent half-lives (15-55 min) of sulfoglycolipids in the isolated tubules could indicate the existence of a rapid turnover pool of these lipids. (3) The sulfotransferase and sulfatase activities related to sulfoamphiphiles in the renal tubule were similar to those reported for the whole kidney. Based on the above criteria, we conclude that the isolated rat renal tubule should be a useful metabolic system for clarification of the short-term physiological events, up to 90 min, of proximal tubular sulfoglycolipids. By using the present system, we showed that biosynthesis of the renal total sulfoglycolipid was significantly elevated in rats deprived of water for 24 h.
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PMID:Metabolism of sulfolipids in isolated renal tubules from rat. 1569 97