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:1.6.99.1 (NADPH-diaphorase)
3,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To define whether astrocytes express a constitutive nitric oxide synthase (NOS), nitric oxide (NO) producing activity in astrocytes derived fetal rat cerebra was examined. We found that the addition of an NOS inhibitor to cultures caused decrease in the basal cGMP levels in unstimulated astrocytes and the decrease was dose-dependent. Further, the expression of cNOS activity in unstimulated astrocytes was confirmed by histochemical staining for NADPH diaphorase and by measuring conversion of L-arginine to L-citrulline. We conclude that cultured astrocytes express constitutive NOS, in addition to inducible one.
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PMID:Presence of constitutive type nitric oxide synthase in cultured astrocytes isolated from rat cerebra. 752 82

Nitric oxide can act as a neurotransmitter and a retrograde modulator of synaptic transmission, but uncontrolled nitric oxide synthase activity has been associated with neural degeneration. Although earlier studies using immunohistochemistry, in situ hybridization, and NADPH-diaphorase staining had suggested that nitric oxide synthase is not expressed in the CA1 neurons of the hippocampus, we have recently demonstrated that NADPH-diaphorase activity can be detected in CA1 neurons of the hippocampus. To confirm that this diaphorase activity reflects nitric oxide synthase, we have developed a more sensitive in situ hybridization procedure, and an RNase protection assay to detect message for constitutive nitric oxide synthase, the form constitutively expressed in many neurons. Message for constitutive nitric oxide synthase is expressed in the hippocampus, and it is localized to neural cell layers CA1, CA3, the dentate gyrus and some displaced neurons, but not to CA2. Expression of constitutive nitric oxide synthase message in the CA1 region was lost when pyramidal neurons died due to transient forebrain ischemia, supporting the conclusion that CA1 pyramidal cells express constitutive nitric oxide synthase. Although constitutive nitric oxide synthase message is strongly expressed in CA3 and the dentate gyrus, there is little diaphorase activity in these cells, suggesting that there may be post-transcriptional controls that limit constitutive nitric oxide synthase expression in some cells. Message for constitutive nitric oxide synthase is also present in a number of other regions, including the amygdala, several hypothalamic nuclei, the cerebellum, the olfactory bulb, two distinct regions of the perirhinal cortex, the subthalamic nuclei, a neuronal layer in the retrosplenial granular cortex, the lateral geniculate nucleus, the presubiculum, the inferior colliculus, the superior colliculus, the pedunculopontine tegmental nucleus, and scattered individual neurons in the cortex, hippocampus and brainstem. These studies support a role for nitric oxide in multiple regions of the central nervous system. In particular, nitric oxide synthase, the enzyme responsible for the synthesis of nitric oxide, is expressed in the CA1 region of the hippocampus, where there is evidence that nitric oxide may play a major role in long-term potentiation. CA1 hippocampal neurons are an example of a population of neurons that express constitutive nitric oxide synthase but are very sensitive to excitotoxicity and ischemic insults.
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PMID:Expression of the neural form of nitric oxide synthase by CA1 hippocampal neurons and other central nervous system neurons. 753 83

Nitric oxide (NO) is a recently recognized messenger molecule that has been shown to possess pleiotropic properties, including vasodilation, neurotransmission, cytotoxicity and antimicrobial activity. Constitutive and inducible forms of NO synthase (NOS) have been identified. Activation of cNOS releases relatively low levels of NO for short periods of time whereas induction of iNOS releases high levels of NO for extended periods of time. In rodents, iNOS is predominantly found in cells of the monocyte/macrophage series, including microglia, where it is induced by a combination of bacterial products and cytokines. cNOS and iNOS have also been reported in rodent astrocytes. Activation of iNOS in the CNS could be toxic to many different cell types, including neurons and oligodendrocytes. iNOS, however, has been difficult to demonstrate in human peripheral blood cells, suggesting that the regulation of expression of this enzyme in humans is different from that found in rodents. In this overview, we show that in human glial cells cultured in vitro, astrocytes, but not microglia, can be induced by cytokines to express NO-like activity. Bacterial products are without effect, but a combination of IL-1 and TNF alpha or IFN gamma is a potent stimulus. NO production by astrocytes inhibits Cryptococcus neoformans growth in vitro. In vivo, we show in acute multiple sclerosis lesions, intense NADPH-diaphorase activity is present in hypertrophic astrocytes in the lesion center and at the lesion edge, whereas microglia are nonreactive. Increased NADPH-diaphorase activity colocalizes with immunoreactivity for IL-1 and TNF. These results suggests that the induction of reactive nitrogen intermediates in humans differs from that found in rodents, and supports the conclusion that hypertrophic astrocytes are the major source of NO-like activity in the inflamed CNS.
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PMID:Reactive nitrogen intermediates in human neuropathology: an overview. 753 80

Nitric-oxide-releasing nerves regulate esophageal smooth muscle function. The density of such nerve fibers may differ in the different functional parts of the esophagus. We used both inspection and gray-scale analysis of digitized images to seek differences in density of such nerve fibers, stained for reduced nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase), between esophageal body and esophago-gastric sphincter and between smooth muscle layers in the opossum esophagus. Sections of Swiss roll preparations of the entire organ were stained for NADPH-diaphorase and for immunoreactivity to vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP), galanin (GAL), substance P (SP) and constitutive nitric oxide synthase (cNOS). In the circular muscle layer, NADPH-diaphorase-positive fibers were most abundant at the cephalic end of the esophageal body with a significant decline toward and through the esophago-gastric sphincter. In the longitudinal muscle layer and the longitudinally-oriented muscularis mucosae, NADPH-diaphorase-positive nerve fibers were most abundant at the esophago-gastric sphincter with a significant decline toward and through the striated-smooth muscle junction. cNOS immunoreactivity co-localized with NADPH-diaphorase activity. Fibers stained for CGRP immunoreactivity were distributed like the NADPH-diaphorase-positive fibers. Fibers stained for immunoreactivity to the other peptides (VIP, GAL, SP) showed no clear differences in distribution along the esophagus in any of the muscle layers.
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PMID:NADPH-diaphorase-positive nerve fibers in smooth muscle layers of opossum esophagus: gradients in density. 754 93

Nitric oxide has recently been implicated as a cellular molecule that mediates interleukin-1 beta (IL-1 beta)-induced inhibition of glucose-stimulated insulin secretion by islets of Langerhans. In this study evidence is presented which demonstrates that islets contain both the cytokine inducible and the constitutive isoforms of nitric oxide synthase as determined by NADPH diaphorase staining and immunohistochemical localization. Untreated islets contain NADPH diaphorase activity, and the intensity of NADPH diaphorase staining is dramatically increased after culture for 18 hrs with IL-1 beta. Both control and IL-1 beta-induced NADPH diaphorase staining of islets is inhibited by the nitric oxide synthase inhibitor NG-monomethyl-L-arginine (NMMA). Importantly, approximately 60-70% of islet cells stained positive for NADPH diaphorase (under both IL-1 beta treated and control conditions), suggesting that a subset of islet cells contain nitric oxide synthase. The beta-cell appears to be the endocrine cell type which contains constitutive nitric oxide synthase as demonstrated by immunohistochemical co-localization of constitutive nitric oxide synthase and insulin. IL-1 beta is believed to stimulate the expression of cytokine inducible nitric oxide synthase because the synthetic glucocorticoid, dexamethasone, prevents IL-1 beta induced inhibition of glucose stimulated insulin secretion and cGMP accumulation by islets. Both dexamethasone, and the nitric oxide synthase inhibitors NMMA and aminoguanidine also prevent IL-1 beta induced islet degeneration. These results indicate that nitric oxide produced by the inducible isoform of nitric oxide synthase mediates cytokine induced islet dysfunction and destruction, and that the beta-cell is the islet endocrine cellular source of constitutive nitric oxide synthase.
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PMID:Nitric oxide mediates IL-1 beta-induced islet dysfunction and destruction: prevention by dexamethasone. 769 96

The cellular localization of inducible (iNOS) and constitutive (cNOS) nitric oxide synthase was studied in rats by immunocytochemical techniques involving specific iNOS and cNOS directed antibodies and by NADPH-diaphorase histochemistry. Paraformaldehyde-fixed vibratome sections of brains and cryostat sections of peripheral lymph nodes were studied of rats treated with endotoxin (2.5 micrograms/kg or 2.5 mg/kg i.v.), rats infected with rabies virus, and rats exposed to experimental allergic encephalomyelitis (EAE). Endotoxin-treated animals showed no appearance of immunoreactive iNOS (ir-iNOS) cells in the brain with the exception of a few microglial cells near the median eminence and some meningeal macrophages. In the same animals however, iNOS-immunoreactive cells were found in peripheral lymph nodes. Neurons that stain positive for cNOS and for NADPH-diaphorase could be observed in brains of control as well as of endotoxin-treated animals with a similar distribution and staining intensity. In contrast, animals that had been infected with rabies virus or subjected to EAE, showed the appearance of ir-iNOS-positive cells in several brain areas. These cells are located near blood vessels and lesion sites. The majority of these cells are GSA-I-B4 isolectin-positive and therefore are likely to represent macrophages. Our data suggest that increased production of nitric oxide may play a role in the altered brain functions in rabies-infected and EAE rats. On the contrary, increased nitric oxide production is probably not involved in the non-specific symptoms of sickness induced by endotoxin.
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PMID:Appearance of inducible nitric oxide synthase in the rat central nervous system after rabies virus infection and during experimental allergic encephalomyelitis but not after peripheral administration of endotoxin. 774 18

Sodium and chloride transport by the macula densa and thick ascending limb of Henle's loop participates importantly in extracellular fluid volume homeostasis, urinary concentration and dilution, control of glomerular filtration, and control of renal hemodynamics. Transepithelial Na and Cl transport across the apical membrane of thick ascending limb (TALH) cells is mediated predominantly by a loop diuretic sensitive Na-K-2Cl cotransport pathway. The corresponding transport protein has recently been cloned. Functional studies suggest that the cotransporter is expressed by macula densa cells as well as by TALH cells. The current studies were designed to identify sites of Na-K-2Cl cotransporter expression along distal nephron in rabbit and rat. Non-isotopic high-resolution in situ hybridization, using an antisense probe for the apical form of the Na-K-2Cl cotransporter identified expression throughout the TALH, from the junction between inner and outer medulla to the transition to distal convoluted tubule. Expression by macula densa cells was confirmed by colocalization using markers specific for macula densa cells. First, Na-K-2Cl cotransporter mRNA was detected in macula densa cells that did not stain with anti-Tamm-Horsfall protein antibodies. Second, Na-K-2Cl cotransporter mRNA was detected in macula densa cells that show positive NADPH-diaphorase reaction, indicating high levels of constitutive nitric oxide synthase activity. In rat, levels of Na-K-2Cl cotransporter mRNA expression were similar in TALH and macula densa cells. In rabbit, expression levels were higher in macula densa cells than in surrounding TALH cells. The present data provide morphological support for a previously established functional concept that Na-K-2Cl cotransport at the TALH is accomplished by the expression of a well-defined cotransporter. At the macula densa, this transporter may establish a crucial link between tubular salt load and glomerular vascular regulation.
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PMID:Expression of the Na-K-2Cl cotransporter by macula densa and thick ascending limb cells of rat and rabbit nephron. 869 54

To better understand the role of nitric oxide (NO) in fetal lung development, specifically in the transition of the fetal circulation at birth, we studied the timing of cell-specific expression of NO synthase (NOS) isoforms from formation of lung buds (13th day of gestation) to 7 days postnatal. Expression of NOS was studied using immunohistochemical labeling with antibodies against the three known NOS isoforms and the NADPH diaphorase technique (NADPH-d). Endothelial NOS (eNOS) immunoreactivity was found in the cells of the 14-day fetal lung. As gestation proceeded, the quantity of these immunopositive cells increased, and they coalesced to form an inner (endothelial) layer of pulmonary vessels. This process of angiogenesis marked by eNOS-positive cells was seen from 15 days of gestation to at least 7 days postnatal. A majority of the eNOS immunoreactivity appeared densely in one focal spot in the cytoplasm, indicating that during development the eNOS may be primarily located in a cytoplasmic organelle. Epithelial cells of the rat airway from the same developmental period were positively stained with both brain NOS antibody (bNOS) and NADPH-d at the beginning of 13 days of gestation. Then the intensity of stainings began to decrease and reached the lowest level in the 16-day fetal lung. However, the NOS stainings of the epithelium, especially in small canalicular structures of the airways, began to increase at 18 days of gestation and was dramatically elevated at 20 days of gestation (term is 22 days). Postnatally, NOS in epithelium was decreased in distal airways in conjunction with the formation of alveolar structure. Inducible NOS (iNOS) immunoreactivity was also found in the epithelium of rat lung airways after 16 days of gestation. Unlike the bNOS staining, iNOS immunoreactivity exhibited a pattern of a small dot-like staining within epithelial cytoplasm during gestation and the first day postnatal, then changed to a pattern of diffuse cytoplasmic staining by the 7th postnatal day. This study concludes that 1) expression of three isoforms of NOS is present and regulated during lung development; 2) markedly increased NOS in epithelium near term supports a role for NO in mediating the pulmonary transition from fetal to neonatal life; and 3) eNOS immunohistochemistry serves as an effective marker to follow the process of pulmonary angiogenesis and suggests the concept of in situ formation of endothelial vesicles in developing mesenchyme.
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PMID:Developmental expression of NOS isoforms in fetal rat lung: implications for transitional circulation and pulmonary angiogenesis. 877 31

Renal haemodynamic changes are suggested to be an early sign of diabetic glomerulopathy. The juxtaglomerular apparatus relevant to the renin angiotensin system, known to be the site of nitric oxide (NO) production, is considered to play a role in the regulation of glomerular blood flow. This study was therefore designed to clarify whether in situ expression of nitric oxide synthase (NOS) is altered in the kidney of diabetic rats. Streptozotocin-induced diabetic rats with 6, 8, 12 and 32 weeks diabetes duration and age-matched normal control rats were used. The expression of a constitutive form of NOS (cNOS, neural type) and NADPH diaphorase activity in the renal cortex were studied immunohistochemically and histochemically. Diabetic rats had lower body weight and heavier kidney mass compared to control rats at each time point examined. Mean glomerular surface area was greater in 6, 8 and 12-week diabetic rats compared to age-matched control rats. cNOS reaction was localized in the macula densa and appeared less intense in diabetic rats compared to age-matched control rats. The mean number of macula densa cells positive for cNOS in each glomerulus and in each glomerular area was significantly lower in diabetic rats compared to control rats at any time examined. In contrast, NADPH diaphorase activity was detected in both juxtaglomerular arterioles and macula densa cells. The staining reaction of NADPH diaphorase in the arterioles remained positive but appeared less intense in macula densa cells in diabetic rats. These results suggest that NO production in macula densa cells may be reduced in diabetic rats, modulating the vasodilatory function of afferent arterioles. Further investigation on the changes in inducible NOS as well as endothelial cNOS are necessary to clarify mechanisms of haemodynamic changes in the diabetic kidney.
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PMID:Expression of nitric oxide synthase in macula densa in streptozotocin diabetic rats. 881 3

The expression of nitric oxide synthase (NOS) in the mucosa of the canine colon was investigated with in situ hybridzation, immunohistochemistry (using isoform specific antibodies), western analysis, and NADPH diaphorase (NADPH-d) histochemistry. In situ hybridization using a common probe for known isoforms of NOS showed that NOS mRNA was strongly expressed in mucosal cells. A gradient in the degree of hybridization was noted from the base of the crypts to the luminal surface. This gradient was also apparent using an endothelial NOS (eNOS)-specific probe. Neural NOS-like immunoreactivity (nNOS-LI) was observed in columnar epithelial cells, and the same population of cells was stained with NADPH-d. Endothelial NOS-like immunoreactivity (eNOS-LI) was also found in mucosal cells; however, this eNOS-LI was confined to mucous cells. These cells were not stained with NADPH-d. The existence of eNOS in mucosal cells was confirmed by in situ hybridization using the probe which specifically hybridized with mRNA of eNOS and by western blots which demonstrated the expression of a 135-kDa protein in mucosal homogenates. The differential expression of NOS isoforms and the gradient in expression along the length of the crypts suggest complex roles for NO in the development of colonic epithelial cells and in secretion and transport functions of the colonic mucosa.
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PMID:Expression of nitric oxide synthase in mucosal cells of the canine colon. 882 4


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