Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.99.3 (diaphorase)
 5,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human cytochrome CYP3A4 is the most abundant of all the P450s in human liver and is involved in the metabolism of many environmental toxicants and drugs. Kinetic studies with CYP3A4 have been hampered due to low activity of this enzyme obtained from recombinant gene expression systems or difficulty in reconstituting activity with the native enzyme purified from human liver. To overcome these obstacles, we have expressed high levels of catalytically active CYP3A4 and human NADPH-cytochrome P450 reductase (CYPOR) together in two insect cell lines, Spodoptera frugiperda (Sf9) and Trichoplusia ni (T.ni), via a single recombinant baculovirus carrying both cDNAs (CYP3A4-OR). Microsomes containing recombinant CYP3A4-OR from these cell lines were up to 50-times more active in testosterone 6 beta-hydroxylase activity than recombinant CYP3A4 expressed alone and supplemented with purified rabbit CYPOR. The spectral P450 content of CYP3A4-OR T.ni microsomes was 107 pmol/mg microsomal protein and the cytochrome c reductase activity was 3904 units/mg. Recombinant CYP3A4-OR was catalytically similar to human liver CYP3A4 based on similarities in the testosterone metabolite profile, time course of metabolite formation, Vmax and Km values (for CYP3A4-OR, Vmax was 8.8 nmol/min/mg microsomal protein [70 nmol/min/nmol CYP3A4] and Km was 33 microM), the extent of inhibition by 100 microM troleandomycin (> 75%) in the presence of 25 microM testosterone, and the degree of P450 activation in the presence of 20 microM 7,8-benzoflavone. The coexpression of recombinant cytochrome b5 with CYP3A4-OR did not result in an additional increase in CYP3A4-OR activity.
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PMID:CYP3A4 expressed by insect cells infected with a recombinant baculovirus containing both CYP3A4 and human NADPH-cytochrome P450 reductase is catalytically similar to human liver microsomal CYP3A4. 777 80

The influence of cell density and epidermal growth factor (EGF) on the expression and inducibility of cytochrome P450 (CYP) genes of the CYP3A and CYP1A families in adult human hepatocytes in primary culture has been evaluated. Only when cultured at subconfluence and in the presence of EGF did hepatocytes exhibit a proliferative response, assessed by measuring DNA synthesis and cyclin A accumulation. In the absence of EGF, the accumulation of CYP3A4 and CYP1A2 messenger RNAs (mRNAs) in response to their respective inducers (rifampicin and dioxin) was dramatically decreased in subconfluent culture with respect to confluent cultures. The presence of EGF only slightly decreased the accumulation of these mRNAs in both confluent and subconfluent cultures. The accumulation of CYP2D6 and CYP2E1 proteins, which are constitutively expressed in confluent cultures, and the production of fibrinogen and apolipoprotein (Apo) B100 exhibited similar behavior, while nicotinamide adenine dinucleotide phosphate cytochrome c reductase activity was affected neither by cell density nor by EGF. In contrast, the accumulation of CYP1A1 mRNA in response to dioxin was similar in confluent and subconfluent cultures, irrespective of the presence of EGF. Interestingly, CYP3A7, a gene that is preferentially expressed in the fetal liver, was expressed constitutively neither in confluent nor in subconfluent cultures, irrespective of the presence of EGF. It is concluded that the loss of cell-cell contacts rather than the proliferative status of cells per se is responsible for the dramatic decrease in the expression of CYP genes, normally expressed in the adult human liver.
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PMID:Effect of cell density and epidermal growth factor on the inducible expression of CYP3A and CYP1A genes in human hepatocytes in primary culture. 914 35

Ketamine is an intravenous anesthetic agent often used for inducing and maintaining anesthesia. Cytoskeletons contribute to the regulation of hepatocyte activity of drug biotransformation. In this study, we attempted to evaluate the effects of ketamine on F-actin and microtubular cytoskeletons in human hepatoma HepG2 cells and its possible molecular mechanisms. Exposure of HepG2 cells to ketamine at <or=100 microM, which corresponds to clinically relevant concentrations for 1, 6, and 24 h, did not affect cell viability. Meanwhile, administration of therapeutic concentrations of ketamine obviously interrupted F-actin and microtubular cytoskeletons. In parallel, levels of intracellular calcium concentration- and time-dependently decreased after ketamine administration. Analysis by confocal microscopy further revealed that ketamine suppressed calcium mobilization from an extracellular buffer into HepG2 cells. Exposure to ketamine decreased cellular ATP levels. The mitochondrial membrane potential and complex I NADH dehydrogenase activity were both reduced after ketamine administration. Ketamine did not change the production of actin or microtubulin mRNA in HepG2 cells. Consequently, ketamine-caused cytoskeletal interruption led to suppression of CYP3A4 expression and its metabolizing activity. Therefore, this study shows that therapeutic concentrations of ketamine can disrupt F-actin and microtubular cytoskeletons possibly through suppression of intracellular calcium mobilization and cellular ATP synthesis due to down-regulation of the mitochondrial membrane potential and complex I enzyme activity. Such disruption of the cytoskeleton may lead to reductions in CYP3A4 activity in HepG2 cells.
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PMID:Cytoskeleton interruption in human hepatoma HepG2 cells induced by ketamine occurs possibly through suppression of calcium mobilization and mitochondrial function. 1884 61