| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1998: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Research Abstract |
1) Methods in Measurement of Nitric Oxide (NO)
There are several methods to measure nitric oxide. In this study we employed three methods, a measurement of oxidative products of NO (NOィイD2XィエD2=NOィイD22ィエD2 and NOィイD23ィエD2) by Griess reaction, electrochemical method and direct detection by measuring chemiluminescence. In this investigation, we clarified an unreliability of NO measurement by Griess reaction or electrochemical method. Firstly, the measurement of NOィイD2XィエD2 by Griess reaction does not always predict NO release because NOィイD2XィエD2 rather than NO could be released from L-canavanine and L-NAME by the action of hydrogen peroxide. Secondly, electrochemical method is interfered by hydrogen peroxide or superoxide.
2) Glomerular Nitric Oxide Production in Experimental Nephritis induced by Puromycin Aminonucleoside
In the early phase (2 days after administration) of the experimental nephritis in rats induced by puromycin aminonucleoside, we clarified increased urinary NOィイD2XィエD2 exc
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retion though there are no increase of immunohistochemical NOS staining in the kidney. Also, NO production increased in the incubation of glomeruli prepared at the same phase. This phenomenon suggests the origin of increased urinary NOィイD2XィエD2 is derived from the kidney. In the very early phase of this nephritis, enhanced production of reactive oxygen species without proteinuria is reported. We speculate this oxidative state bring about non-enzymatic NO production in the kidney.
3) Involvement of a Non-enzymatic Nitric Oxide Generation Pathway in Hemodialysis Patients
We have already reported that NO production in hemodialysis patients was significantly higher than healthy control. And uremic condition is known to be peroxidative and a state of increased production of reactive oxygen species. In this study, we investigated the serum inhibitory effect on non-enzymatic NO synthesis by the reaction of L-arginine and hydrogen peroxide. As a result, the inhibitory effect on the NO synthesis of pre-hemodialysis sera is significantly weaker than that of healthy control, and this less inhibitory effect recovered to healthy level after hemodialysis treatment. This study proposes that an increased NO production in hemodialysis patients is a result of peroxidative state in uremia.
4) Evaluation of Redox State in the Kidney by in vivo Electron Spin Resonance (ESR)
This study employed a new technique that enables in vivo detection of free radicals. In an experimental nephrosis caused by puromycin aminonucleoside (PAN), an oxidative state is thought to play a key role. By the combination of this new ESR technique and newly developed spin probing reagent, 4-hydroxy 2,2,6,6-tetramethyl - piperidine-1-oxyl (TEMPOL), the ESR signal derived from this decreases with oxidation, we investigated the redox state of the kidney in vivo in PAN nephrosis. As a result, the reducing activity of the kidney in nephritic group was significantly weaker than that of control. This investigation directly proved the decreased reducing activity of nephritic kidney in vivo.
5) Influence of Endothelial Nitric Oxide Synthase Gene Polymorphism in Intron 4 on the Progression of Renal Diseases
There are several polymorphisms in endothelial nitric oxide synthase (eNOS) gene, and the homozygosity for the eNOS intron 4 a allele is reported to be a risk factor for coronary artery disease among smokers. In this study, we clarified that the frequency of the eNOS intron 4 a allele was significantly higher in the patients with non-diabetic end stage renal failure compared with healty control. This result indicates that the eNOS 4 a allele is a risk factor for end stage renal failure in non diabetic renal diseases. Less
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