Abrantes Daniele C, Brunini Tatiana M C, Matsuura Cristiane, Mury Wanda Vianna, Corrêa Carolina R, Santos Sérgio F, Ormonde do Carmo Monique B O, Mendes-Ribeiro Antônio Cláudio
Department of Pharmacology and Psychobiology, University of the State of Rio de Janeiro, Rio de Janeiro, 20551-030, Brazil.
Mol Cell Biochem. 2015 Mar;401(1-2):147-53. doi: 10.1007/s11010-014-2302-1. Epub 2014 Dec 19.
Chronic renal failure (CRF) is a complex clinical condition associated with accelerated atherosclerosis and thrombosis leading to cardiovascular events. The aim of this study was to investigate in detail the NO pathway in neutrophils obtained from hemodialysis patients and its association with platelet function and oxidative status. Fifteen CRF patients on hemodialysis and fifteen controls were included in this study. Laboratory and experimental evaluations were performed after hemodialysis in CRF patients. We evaluated L-[³H] arginine transport, NO synthase (NOS) activity, amino acid concentration in neutrophils, and expressions of NOS isoforms and p47(phox) by western blotting. Platelet aggregation was analyzed in the presence or absence of neutrophils. Oxidative status was measured through glutathione peroxidase, catalase activities, protein oxidation, lipid peroxidation, and DNA/RNA oxidation in serum. Basal NOS activity (pmol/10⁶ cells/min) was impaired in CRF patients on hemodialysis (0.33 ± 0.17) compared to controls (0.65 ± 0.12), whereas the expression of NOS isoforms remained unaltered. L-Arginine transport into neutrophils was similar in CRF patients on hemodialysis and controls. In addition, intracellular concentration of L-arginine was increased fourfold in the patient group. Systemic oxidative stress markers were not affected by CRF. On the other hand, NADPH oxidase subunit p47(phox) in neutrophils was overexpressed in CRF. In the presence of neutrophils, there was a reduction time-dependent in platelet aggregation in both groups with no difference between them. This data suggest that reduced basal generation of NO by neutrophils in CRF patients on hemodialysis occurs independently of L-arginine bioavailability and is able to suppress platelet activation.
慢性肾衰竭（CRF）是一种复杂的临床病症，与加速的动脉粥样硬化和血栓形成相关，可导致心血管事件。本研究的目的是详细调查血液透析患者中性粒细胞中的一氧化氮（NO）途径及其与血小板功能和氧化状态的关联。本研究纳入了15例接受血液透析的CRF患者和15例对照者。对CRF患者在血液透析后进行了实验室和实验评估。我们通过蛋白质免疫印迹法评估了L-[³H]精氨酸转运、一氧化氮合酶（NOS）活性、中性粒细胞中的氨基酸浓度以及NOS同工型和p47（phox）的表达。在有或无中性粒细胞存在的情况下分析血小板聚集情况。通过血清中的谷胱甘肽过氧化物酶、过氧化氢酶活性、蛋白质氧化、脂质过氧化和DNA/RNA氧化来测定氧化状态。与对照者（0.65±0.12）相比，接受血液透析的CRF患者的基础NOS活性（pmol/10⁶细胞/分钟）受损（0.33±0.17），而NOS同工型的表达保持不变。血液透析的CRF患者和对照者中L-精氨酸向中性粒细胞的转运相似。此外，患者组中L-精氨酸的细胞内浓度增加了四倍。CRF未影响全身氧化应激标志物。另一方面，CRF患者中性粒细胞中的NADPH氧化酶亚基p47（phox）过度表达。在有中性粒细胞存在的情况下，两组的血小板聚集均随时间减少，且两组之间无差异。这些数据表明，接受血液透析的CRF患者中性粒细胞基础NO生成减少独立于L-精氨酸的生物利用度，并且能够抑制血小板活化。
