Amount Of Nitric Oxide Synthase Research Articles

Characterization of L-arginine transporters in rat renal inner medullary collecting duct.

Previous work from our laboratory has demonstrated that the inner medullary collecting duct (IMCD) expresses a large amount of nitric oxide synthase (NOS) activity. The present study was designed to characterize the transport of NOS substrate, L-arginine, in a suspension of bulk-isolated IMCD cells from the Sprague-Dawley rat kidney. Biochemical transport studies demonstrated an L-arginine transport system in IMCD cells that was saturable and Na(+) independent (n = 6). L-Arginine uptake by IMCD cells was inhibited by the cationic amino acids L-lysine, L-homoarginine, and L-ornithine (10 mmol/l each) and unaffected by the neutral amino acids L-leucine, L-serine, and L-glutamine. Both L-ornithine (n = 6) and L-lysine (n = 6) inhibited NOS enzymatic activity in a dose-dependent manner in IMCD cells, supporting the important role of L-arginine transport for NO production by this tubular segment. Furthermore, RT-PCR of microdissected IMCD confirmed the presence of cationic amino acid transporter CAT1 mRNA, whereas CAT2A, CAT2B, and CAT3 were not detected. These results indicate that L-arginine uptake by IMCD cells occurs via system y(+), is encoded by CAT1, and may participate in the regulation of NO production in this renal segment.

Expression of nitric oxide synthase in inflammatory bowel disease is not affected by corticosteroid treatment.

AIM: To examine the effect of corticosteroid treatment on the expression of inducible nitric oxide synthase (iNOS) in the colon of patients with inflammatory bowel disease. METHODS: Four groups of...

Endothelial nitric oxide synthase in hypoxic newborn porcine pulmonary vessels

AIMSTo determine if the failure of neonatal pulmonary arteries to dilate is due to a lack of nitric oxide synthase (NOS).METHODSA monoclonal antibody to endothelial NOS was used to demonstrate...

Synergistic cooperation between thapsigargin and phorbol ester for induction of nitric oxide synthesis in murine peritoneal macrophages

The biochemical transductional events involved in NO synthesis are not fully understood. These studies, therefore, were undertaken to elucidate the role of intracellular calcium and protein kinase C (PKC) in the induction of nitric oxide (NO) synthesis in murine peritoneal macrophages. Thapsigargin (TG), Ca 2+ -ATPase inhibitor of endoplasmic reticulum, had modest activity on NO synthesis by itself, whereas phorbol ester, PKC activator, alone had no effect. When TG was used in combination with phorbol ester, there was a marked cooperative induction of NO synthesis in a dose-dependent manner. The optimal effect of phorbol ester was shown in the first 6 h after TG treatment. In addition, the ability of TG with phorbol ester on NO synthesis could be mimicked by another chemically unrelated inhibitor of Ca 2+ -ATPase, 2,5-di-( t-butyl)-1,4-benzohydroquinone, and Ca 2+ ionophore, A23187. This increase of NO synthesis was reflected as increased amount of NO synthase (NOS) mRNA, as determined by Northern blotting. Intracellular Ca 2+ transient by TG was not affected in the presence or absence of extracellular Ca 2+, indicating that TG must be effective on cytosolic Ca 2+ pool. In addition, chelation of intracellular Ca 2+ by acetoxymethyl ester of 1,2- bis-(2-aminophenoxy)-ethane- N,N,N′,N′-tetraacetic acid (BAPTA/AM), an intracellular Ca 2+ chelating agent, blocked TG- or TG + PMA-induced NO production. PKC inhibitors such as staurosporine or polymyxin B reduced only the synergistic cooperative effect of TG with phorbol ester without affecting TG-induced NO production. In addition, when the cells were pretreated with phorbol ester before TG treatment, there was no synergy between TG and phorbol ester, indicating that PKC is not directly involved in the expression of NOS but involved in “triggering” signal. Secretion of NO corresponded with tumor cell killing, but TG plus phorbol ester-activated macrophages failed to kill tumor cell targets in the presence of N G-monomethyl- l-arginine. Collectively, these data illustrate that mobilization of intracellular Ca 2+ provides a “priming” signal for induction of NOS gene expression by itself and it also requires PKC as a “triggering” signal for macrophage tumoricidal activity.

Plasma nitrate clearance in mice: modeling of the systemic production of nitrate following the induction of nitric oxide synthesis.

Nitric oxide (NO) is produced in mammals by the enzyme NO synthase (NOS) in response to a number of agents, including the experimental antitumour agent flavone acetic acid (FAA) and the cytokine tumour necrosis factor-alpha (TNF). NO is converted rapidly in the presence of oxygen, water and haemoglobin to oxidation products, largely nitrate. To quantitate the production of nitric oxide it is necessary to know the clearance of nitrate. The concentration of nitrite and nitrate ion in the plasma of C3H and BDF1 (C57BL6 x DBA2) mice was assessed before and after injection of sodium nitrate and sodium nitrite. Nitrite was covered rapidly to nitrate and the kinetics of elimination of nitrate were determined. There was no significant difference between results obtained with different mouse strains, between levels of nitrite and nitrate, or between i.p. and i.v. administration, and the observations were therefore combined. The volume of distribution of nitrate was 0.71 +/- 0.04 l/kg and the clearance was 0.32 +/- 0.02 l/h-1/kg-1 (plasma half-life, 1.54 h). Using previously published data, we developed a pharmacokinetic-pharmacodynamic model that relates the production of TNF in response to administration of FAA, the enhancement of NOS activity in response to TNF, and the elevation of plasma nitrate in response to NO production. This information permits the prediction from observed plasma nitrate values of the amount of NOS induced in vivo.