Yokota Takashi, Kamimura Naomi, Igarashi Tsutomu, Takahashi Hiroshi, Ohta Shigeo, Oharazawa Hideaki
Department of Biochemistry and Cell Biology, Institute of Development and Aging Sciences, Graduate School of Medicine, Nippon Medical School, Kawasaki, Kanagawa, Japan.
Department of Ophthalmology, Nippon Medical School, Tokyo, Japan.
Clin Exp Ophthalmol. 2015 Aug;43(6):568-77. doi: 10.1111/ceo.12525. Epub 2015 May 13.
Oxidative and nitrative processes have an important role in the pathogenesis of glaucomatous neurodegeneration. Oxidative stress occurs when cellular production of reactive oxygen species outweighs the protective capacity of antioxidant defences. Reactive oxygen species are generated as by-products of cellular metabolism, primarily in the mitochondria. Herein, we present a novel investigation of the effects of molecular hydrogen (H2 ) on retinal cells exposed to oxidative stress.
We cultured adult rat retinal tissues in an organotypic culture system with a nitric oxide donor, S-nitroso-N-acetylpenicillamine, in the presence or absence of H2 . Loss of mitochondrial membrane potential and apoptosis of retinal cells were analysed using a MitoTMRE detection kit and TdT-mediated dUTP nick end labeling (TUNEL) assay, respectively. Tyrosine nitration levels and oxidative stress damage in the retina were evaluated using immunohistochemical staining. Retinal damage was quantified by measuring the numbers of cells in the ganglion cell and inner nuclear layers and the thickness of the retina.
H2 suppressed loss of mitochondrial membrane potential and apoptosis in retinal cells. Moreover, H2 decreased the tyrosine nitration level and suppressed oxidative stress damage in retinal cells. S-nitroso-N-acetylpenicillamine treatment decreased the cell numbers in the ganglion cell layer and inner nuclear layer, but the presence of H2 inhibited this reduction. These findings suggest that H2 has a neuroprotective effect against retinal cell oxidative damage, presumably by scavenging peroxynitrite.
H2 reduces cellular peroxynitrite, a highly toxic reactive nitrogen species. Thus, H2 may be an effective and novel clinical tool for treating glaucoma and other oxidative stress-related diseases.
氧化和硝化过程在青光眼性神经退行性变的发病机制中起重要作用。当细胞内活性氧的产生超过抗氧化防御的保护能力时，就会发生氧化应激。活性氧作为细胞代谢的副产物产生，主要在线粒体中。在此，我们对分子氢（H₂）对暴露于氧化应激的视网膜细胞的影响进行了一项新的研究。
我们在器官型培养系统中，在有或没有H₂的情况下，用一氧化氮供体S-亚硝基-N-乙酰青霉胺培养成年大鼠视网膜组织。分别使用MitoTMRE检测试剂盒和TdT介导的dUTP缺口末端标记（TUNEL）分析来检测视网膜细胞的线粒体膜电位丧失和凋亡。使用免疫组织化学染色评估视网膜中的酪氨酸硝化水平和氧化应激损伤。通过测量神经节细胞层和内核层中的细胞数量以及视网膜厚度来量化视网膜损伤。
H₂抑制了视网膜细胞的线粒体膜电位丧失和凋亡。此外，H₂降低了视网膜细胞中的酪氨酸硝化水平，并抑制了氧化应激损伤。S-亚硝基-N-乙酰青霉胺处理减少了神经节细胞层和内核层中的细胞数量，但H₂的存在抑制了这种减少。这些发现表明，H₂对视网膜细胞氧化损伤具有神经保护作用，可能是通过清除过氧亚硝酸盐来实现的。
H₂可减少细胞内的过氧亚硝酸盐，这是一种剧毒的活性氮物质。因此，H₂可能是治疗青光眼和其他氧化应激相关疾病的一种有效且新颖的临床工具。
