Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.5.2 (NQO1)
 6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

NO synthase (NOS; EC 1.14.23) catalyzes the conversion of L-arginine into L-citrulline and a guanylyl cyclase-activating factor (GAF) that is chemically identical with nitric oxide or a nitric oxide-releasing compound (NO). Similar to the other isozymes of NOS that have been characterized to date, the soluble and Ca2+/calmodulin-regulated type I from rat cerebellum (homodimer of 160-kDa subunits) is dependent on NADPH for catalytic activity. The enzyme also possesses NADPH diaphorase activity in the presence of the electron acceptor nitroblue tetrazolium (NBT). We investigated the requirements of NOS and its content of the proposed additional cofactors tetrahydrobiopterin (H4biopterin) and flavins, further characterized the NADPH diaphorase activity, and quantified the NADPH binding site(s). Purified NOS type I Ca2+/calmodulin-independently bound the [32P]2',3'-dialdehyde analogue of NADPH (dNADPH), which, at near Km concentrations during 3-min incubations was utilized as a substrate and at higher concentrations or after prolonged incubations and cross-linking inhibited NOS activity. The NADPH diaphorase activity was Ca2+/calmodulin-independent, required higher NADPH concentrations than NOS activity, and was affected by dNADPH to a lesser degree. Divalent cations interfered with the diaphorase assay. Per dimer, native NOS contained about 1 mol each of H4biopterin, FAD, and FMN, classifying it as a biopteroflavoprotein, and incorporated 1 mol of dNADPH. No dihydrobiopterin (H2biopterin), biopterin, or riboflavin was detected. These findings suggest that NOS may share cofactors between two identical subunits via high-affinity binding sites.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Ca2+/calmodulin-dependent NO synthase type I: a biopteroflavoprotein with Ca2+/calmodulin-independent diaphorase and reductase activities. 137 27

The application of enzymatic staining techniques, using tetrazolium dyes, to aldehyde-treated brain sections has revealed the presence of NADPH-diaphorase activity attributed to nitric oxide synthase. When evaluating the specificity of the putative guanylyl cyclase inhibitor LY 83583, a robust and novel staining pattern was noted in epithelial, endothelial, and astrocytic cells when LY 83583 was included in the NADPH-diaphorase histochemical reaction. This LY 83583-dependent staining could be blocked by the NAD(P)H:quinone oxidoreductase inhibitor dicumarol. Based on its quinone structure, we hypothesized that LY 83583 was a substrate for the enzyme NAD(P)H:quinone oxidoreductase. Transfection of human embryonic kidney 293 cells with the rat liver isoform of NAD(P)H:quinone oxidoreductase resulted in robust NADPH- and LY 83583-dependent staining that was completely blocked by dicumarol and was not observed in untransfected cells. Analysis of transfected cell extracts and brain homogenates indicated that LY 83583 was a substrate for NAD(P) H:quinone oxidoreductase, with a Km similar to the well-characterized substrate menadione. Sensitivity of the nitroblue tetrazolium reduction to superoxide dismutase indicated that the reduction of LY 83583 by NAD(P)H:quinone oxidoreductase leads to superoxide generation. The localization of NAD(P)H:quinone oxidoreductase activity to astrocytic cells suggests a role for glia in combating oxidative insults to brain and in activating quinone-like drugs such as LY 83583.
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PMID:Histochemical detection of quinone reductase activity in situ using LY 83583 reduction and oxidation. 957 3

Non-restrictive, porous, external stents inhibit neointima formation in porcine vein grafts. Since the mechanisms underlying these effects are unknown we investigated the impact of this external stent on factors known to inhibit vascular smooth muscle cell proliferation: prostacyclin (PGI2), nitric oxide (NO), cAMP and cGMP formation in different regions of stented and unstented porcine vein grafts. Paired stented and unstented saphenous vein-carotid artery interposition grafting was carried out in Landrace pigs. One month after surgery, the vessels were excised and the formation of PGI2, cAMP and cGMP determined using radioimmunoassay and nitric oxide synthase (NOS) distribution studied using autoradiography and histochemistry. There were no significant differences between PGI2, cAMP and cGMP (nitroprusside-stimulated) formation in the medial/intimal regions of grafts of stented vein graft and ungrafted saphenous vein whereas all were significantly reduced in unstented vein graft. A23187-stimulated cGMP formation (mediated by NO release) and NOS content was significantly greater in the medial/intimal region of stented and unstented vein graft compared to ungrafted saphenous vein, indicating induction of endothelial NOS (eNOS) in both types of graft. This normalisation of the PGI2-cAMP axis and guanylyl cyclase activity in the medial/intimal region may contribute to the beneficial impact of the external stent on vein graft thickening. The increase in eNOS in both stented and unstented vein grafts mitigates against this isoform as playing a role in mediating the inhibitory effect of the stent on neointima formation. In the adventitia of both stented and unstented grafts there was an increase in PGI2, cAMP and cGMP formation compared to ungrafted saphenous vein, the production being greater in the stented compared to the unstented graft. In the adventitia of stented veini grafts, NOS, detected with NAPDH diaphorase staining, was associated with microvessels as well as with inflammatory cells. Taken together, these data are suggestive of a role for PGI2 and NO in promoting microangiogenesis in the adventitia of stented vein grafts which may in turn minimize graft hypoxia, an established contributory factor to neointima formation.
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PMID:Nitric oxide, prostacyclin and cyclic nucleotide formation in externally stented porcine vein grafts. 986 78

It has recently been suggested that, in addition to nitric oxide (NO), carbon monoxide (CO) is an important gaseous messenger which might be involved in vertebrate olfactory transduction because its effects include activation of guanylyl cyclase and the formation of cGMP. As there is no information regarding the presence of heme oxygenase-2 -- the constitutive isoform of the heme oxygenase system -- in olfactory neurons of non-rodent species, we have investigated the distribution pattern of heme oxygenase-2 in the olfactory epithelium of the bovine, a representative of macrosmatics. Localization of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) activity of the olfactory epithelium was compared with heme oxygenase-2 and NO synthase (NOS) immunoreactivities in order to obtain possible hints at functional significance. NADPH-d activity was particularly intense in apical dendrites of receptor neurons. It was also found in Bowman glands and intraepithelial duct cells. Less intense, discrete NADPH-d activity was present also at intermediate and basal levels of the olfactory epithelium, corresponding to the layer of receptor neuron somata and basal cells. While heme oxygenase-2 activity mainly occurred in neuronal perikarya, a very intense NOS immunoreactivity, exclusively for the inducible isoform, was detected in the apical dendrites. Ultrastructurally, NADPH-d histochemistry showed distinct labelling of membranes, in particular of endoplasmic reticulum, mitochondria and nucleus. The coincident localization of the moderate NADPH-d activity and heme oxygenase-2 immunoreactivity in receptor cell perikarya suggest a functional association between NADPH-cytochrome P450 reductase and heme oxygenase-2. In contrast, dendritic localization of NADPH-d activity is topically and possibly functionally related to the presence of the inducible isoform of NOS. The results suggest that both CO and NO may be generated in bovine receptor neurons and thus involved in odorant stimulation. Based on immunocytochemical localization of synthesizing enzymes, NO might be regarded as a direct regulator of transduction related processes while CO might act as a modulator of the initial signal.
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PMID:Heme oxygenase-2 and nitric oxide synthase immunoreactivity of bovine olfactory receptor neurons and a comparison with the distribution of NADPH-diaphorase staining. 1094 53

In the current report, we summarize our findings related to the involvement of nitric oxide (NO) in the pathology of spinal cord trauma. We initially studied the distribution of nitric oxide synthase (NOS)-immunolabeled and/or nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd; which is highly colocalized with NOS)-stained somata and fibers in the spinal cord of the rabbit. Segmental and laminar distribution of NADPHd-stained neurons in the rabbit revealed a large number of NADPHd-stained neurons in the spinal cord falling into six categories, N1-N6, while others could not be classified. Large numbers of NADPHd-stained neurons were identified in the superficial dorsal horn and around the central canal. Four morphologically distinct kinds of NADPHd-stained axons 2.5-3.5 microm in diameter were identified throughout the white matter in the spinal cord. Moreover, a massive occurrence of axonal NADPHd-staining was detected in the juxtagriseal layer of the ventral funiculus along the rostrocaudal axis. The prominent NADPHd-stained fiber bundles were identified in the mediobasal and central portion of the ventral funiculus. The sulcomarginal fasciculus was found in the basal and medial portion of the ventral funiculus in all cervical and thoracic segments. Since the discovery that NO may act as a neuronal transmitter, an increasing interest has focused on its ability to modulate synaptic function. NO passes through cell membranes without specific release or uptake mechanisms inducing changes in signal-related functions by several means. In particular, the activation of the soluble guanylyl cyclases (sGC), the formation of cyclic guanosine 3',5'-monophosphate (cGMP) and the action of cGMP-dependent protein kinases has been identified as the main signal transduction pathways of NO in the nervous system including spinal cord. It is known that the intracellular level of cGMP is strictly controlled by its rate of synthesis via guanylyl cyclases (GC) and/or by the rate of its degradation via 3',5'-cyclic nucleotide phosphodiesterases (PDE). GC can be divided into two main groups, i.e., the membrane-bound or particular guanylyl cyclase (pGC) and the cytosolic or sGC. In the spinal cord, the activation of pGC has only been demonstrated for natriuretic peptides, which stimulate cGMP accumulation in GABA-ergic structures in laminae I-III of the rat cervical spinal cord. These neurons are involved in controlling the action of the locomotor circuit. In view of the abundance of NO-responsive structures in the brain, it is proposed that NO-cGMP signaling will be part of neuronal information processing at many levels. In relation to this, we found that surgically induced Th7 constriction of 24 h duration stimulated both the constitutive NOS activity and cGMP level by 120 and 131%, respectively, in non-compartmentalized white matter of Th8-Th9 segments, located just caudally to the site of injury. NO-mediated cGMP formation was only slightly increased in the dorsal funiculus of Th5-Th9 segments. There are some other sources that may influence the NO-mediated cGMP formation in spinal cord. A high level of glutamate produced at the site of the lesion and an excessive accumulation of intracellular Ca2+ may stimulate NOS activity and create suitable conditions for NO synthesis and its adverse effect on white matter. An increased interest has focused on the role of NO at the site of injury and in areas located close to the epicenter of the impact site and, in these connections an upregulation of NOS was noted in neurons and interneurons. However, the upregulation of NOS expression was also seen in interneurons located just rostrally and caudally to the lesion. A quantitative analysis of laminar distribution of multiple cauda equina constriction (MCEC) induced NADPHd-stained neurons revealed a considerable increase in these neurons in laminae VIII-IX 8h postconstriction, and a highly statistically significant increase of such neurons in laminae VII-X 5 days postconstriction in the lumbosacral segments. Concurrently, the number of NADPHd-stained neurons on laminae I-II in LS segments was greatly reduced. It is concluded that a greater understanding of NO changes after spinal cord trauma is essential for the possibility of targeting this pathway therapeutically.
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PMID:Traumatic injury of the spinal cord and nitric oxide. 1761 76