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)

NADPH cytochrome c (P-450) reductase was purified from human placental microsomes using a combination of affinity and gel filtration chromatography. Affinity chromatography using agarose-hexane-adenosine 2'5 diphosphate resulted in two protein bands being detected by SDS-PAGE of approximate MwS 68 and 75 kDa. Fractions containing the two proteins were pooled, and then resolved using Sephacryl S-200. Both of the purified proteins displayed enzyme activity, measured by their ability to reduce cytochrome c. The 75 kDa protein obtained was used to immunize three female New Zealand white rabbits. The IgG fraction was partly purified from rabbit sera which suppressed placental microsomal NADPH cytochrome c reductase activity by > 80% using 33% ammonium sulphate. The procured antibody suppressed androstenedione aromatase activity in microsomal preparations of human placental and breast adipose tissue, and NADPH cytochrome c reductase activity in prostate (benign and malignant), MDA-MB-231 breast cancer cells, breast adipose, Hep G2 hepatoma cells and placental microsomal preparations. The extent of NADPH cytochrome c reductase inhibition varied in the order of malignant prostate < benign prostate < MDA < breast adipose < Hep G2 < placenta. The results suggest that human placental NADPH cytochrome c (P-450) reductase shares common antigenic epitopes pertinent to its capability of reducing cytochrome c in all of the above-mentioned tissues. In attempting to associate possible changes in NADPH cytochrome c reductase activity imposed by neoplasia to the obtained immunochemical cross reactivity and enzyme activity results, it was noted that microsomes obtained from MDA cells exhibited enzyme activity significantly less than that of breast adipose microsomes (1.6 and 8.1 nmol/min/mg protein, respectively) and by comparison showed 6% less homology towards the placental antibody. The results obtained for benign and malignant prostate showed no significant difference between the neoplastic states as adjudged by enzyme activity and immunochemical assays.
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PMID:Immunochemical specificity of placental NADPH cytochrome c (P-450) reductase in neoplastic and non-neoplastic human tissue. 141 86

Two distinct aromatase-active protein complexes are solubilized by use of deoxycholate and separated by diethylamino-ethyl-cellulose chromatography from lyophilized powder of 900 X g precipitate fraction of human term placenta. Aromatase activity to produce estriol, the major estrogen of human pregnancy, was designated to be aromatase I activity and measured by estriol formation from 16 alpha-hydroxytestosterone. Aromatases II activity was the designation for that which produces estrone plus estradiol and was measured by androstenedione aromatization. Aromatases II and I are eluted with 0.25 M and 0.5 M Tris buffer, respectively, from diethylaminoethyl-cellulose column in an Mr 2 million soluble complex. Each has a minimum active Mr 135,000 subunit, which is isolated by Bio-Gel filtration in the presence of detergents, and consists of a reduced nicotinamide adenine dinucleotide phosphate:cytochrome P-450 reductase (Mr 83,000) and a cytochrome P-450 (Mr 52,000). Aromatase II was found to be the major aromatase, containing approximately five times more aromatase activity, reduced nicotinamide adenine dinucleotide phosphate:cytochrome c reductase activity, cytochrome P-450, and protein than did aromatase I. Antibodies raised in rabbits against aromatase II and its reductase suppressed aromatase II activity of breast cancer tissues, as well as of adult male lung tissue, placental microsomes, and solubilized aromatase. The breast carcinoma specimens responded to the antibodies in different degrees, but there was no response to antibodies against rat liver cytochrome P-450. The results indicate similar antigenic structures for breast cancer and placental aromatase but not for rat liver cytochrome P-450.
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PMID:Multiple forms of aromatase and response of breast cancer aromatase to antiplacental aromatase II antibodies. 617 1

To analyze the mechanism by which nitric oxide (NO) exerts its antisteroidogenic action, human luteal cells were cultured during 24 and 48 h with L-arginine (L-Arg, 1 mmol/L); 1,2(2-trifluoromethylphenyl)imidazole (TRIM) (50 micromol/L and 1 mmol/L) and cyclic guanosine monophosphate (cGMP) analog (8-Br-cGMP, 1 mmol/L). Estradiol, nitrite, and P450 AROM activity were determined in culture media. Total cGMP concentration was evaluated in the cells and culture media by radioimmunoassay, and NADPH diaphorase was used as a histochemical marker for NO synthase (NOS) activity. During the corpus luteum (CL) life-span, NO affected estradiol secretion in an age-dependent manner, with an inhibition in mid-CL (37%; p < 0.05) in agreement with our previous results, and no significant modification in early and late CL. Basal nitrite concentration in 24 and 48 h of midluteal cell cultures (42 and 93 pmol/10(6) cells, respectively) was increased by L-Arg (53% and 88%) and inhibited by the two TRIM concentrations; also, an intense diaphorase reactivity was observed in endothelial cells and luteal parenchyma. Total cGMP was not detected in cell cultures and 8-Br-cGMP did not modify estradiol secretion, whereas aromatase activity was strongly inhibited by L-Arg (70%, p < .05). These results suggest that both NOS isoforms are active in midluteal cells, and the mechanism of action for NO on in vitro estradiol secretion may be an inhibition of P450 AROM activity.
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PMID:Antisteroidogenic action of nitric oxide on human corpus luteum in vitro: mechanism of action. 1066 38

Normal and imposex-affected female Buccinum undatum were sampled from the open North Sea at three locations, one with low, and two with high shipping densities. Cytochrome P450 components and P450 aromatase activity were determined in the microsomal fractions isolated from pooled digestive gland/gonads. Cytochrome P450 aromatase activity was significantly higher (P < 0.05) in normal females collected in the low shipping density area (1,325 +/- 295 fmol/h/mg protein) than levels from imposex animals from a high shipping density area (620 +/- 287 fmol/h/mg protein). A negative correlation was found between aromatase activity and organotin body burden (r = -0.99). Levels of CYP450, cytochrome b5 and NADPH cytochrome c reductase activity did not show differences among groups. This is the first field evidence of depressed aromatase activity in imposex affected females, although additional research under laboratory controlled conditions is required to fully understand the mechanisms underlying the development of imposex in this species.
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PMID:Cytochrome P450 differences in normal and imposex-affected female whelk Buccinum undatum from the open North Sea. 1240 32

In Japanese quail (Coturnix japonica), previous studies indicated that the distribution of reduced nicotinamide dinucleotide phosphate (NADPH) diaphorase overlaps with steroid-sensitive areas that contain dense populations of aromatase-immunoreactive (ARO-ir) cells. We investigated here the anatomical relationships between aromatase (ARO) and nitric oxide synthase (NOS)-containing cells that were visualized both by NOS-immunohistochemistry and NADPH-histochemistry. The distribution of ARO-ir and of NADPH-positive cells in the forebrain observed here matched exactly the distribution previously reported. The distribution of NOS-immunoreactive material in the vicinity of ARO-ir cell groups appeared similar to the distribution of NADPH-positive structures previously identified by histochemistry. The number of NOS-immunoreactive cells was similar to the number of NADPH-positive cells and they were found in the same brain regions. In contrast, few NOS-immunoreactive fibers were observed whereas numerous NADPH-positive fibers and punctuate structures were present in many areas. Major groups of NOS-immunoreactive/NADPH-positive neurons were adjacent to the main ARO-ir cell groups, such as the medial preoptic nucleus, the bed nucleus of the stria terminalis and the nucleus ventromedialis hypothalamic. However, examination of adjacent sections indicated that there is very little overlap between the NOS-immunoreactive and ARO-ir cell populations. This notion got further support by double-labeled sections where no double-labeled cells could be identified. In sections stained simultaneously by histochemistry for NADPH and immunohistochemistry for ARO, many NADPH-positive fibers and punctate structures were closely associated with ARO-ir perikarya. Taken together, the present data indicate that NOS is not or very rarely colocalized with ARO but that NOS inputs are closely associated with ARO-ir cells. Based on previous work in a variety of model systems, it can be hypothesized that these inputs modulate the expression or activity of ARO in the quail brain.
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PMID:Anatomical relationships between aromatase-immunoreactive neurons and nitric oxide synthase as evidenced by NOS immunohistochemistry or NADPH diaphorase histochemistry in the quail forebrain. 1257 58