基于线粒体损伤信号通路研究叶酸对高同型半胱氨酸血症小鼠急性肾损伤的作用和分子机制

Acute kidney injury (AKI) is a common and multiphasic clinical syndrome. The prevalence of AKI has been increasing rapidly around the world. AKI is also known for its association with unacceptably high in-hospital mortality. Whether or not patients have preexisting chronic kidney disease (CKD), those who have had AKI have a high risk of developing progressive CKD and end-stage renal disease(ESRD) over time. Renal tubular cell apoptosis are recognized as the precipitating factors in AKI. Mitochondrial damage is one of the three apoptotic pathways involved in apoptosis of renal tubular epithelial cells. Homocysteine (Hcy) is a sulfur-containing amino acid. An elevated plasma or serum Hcy level is denoted hyperhomocysteinemia (HHcy). Our previous study has shown that HHcy can aggravate cisplatin induced acute renal injury via endoplasmic reticulum stress and inhibition of Akt phosphorylation. However, there is no research on how to interfere with the role of hyperhomocysteinemia in AKI. The aim of this study was to investigate the effects of folic acid on the occurrence and development of acute renal injury in mice with hyperhomocysteinemia by mitochondrial damage pathway.

急性肾损伤（AKI）是一组多因素参与的复杂而常见的临床综合征。全世界范围内，AKI的发病率正迅速增加，且与增高的住院死亡率有关。部分AKI可能发展为进展性CKD，甚至进入终末期肾病（ESRD）。在急性肾损伤中，肾小管细胞凋亡起至关重要的致病作用。线粒体损伤是肾小管上皮细胞凋亡主要涉及3条凋亡通路之一。同型半胱氨酸（Hcy）为一种含硫氨基酸。血清或血浆Hcy浓度升高，称为高同型半胱氨酸血症(HHcy)。我们之前的研究显示HHcy可通过内质网应激及抑制Akt的磷酸化加重顺铂诱导的急性肾损伤。但对于如何干预高同型半胱氨酸血症在AKI上的作用目前尚无相关研究。本研究旨在急性损伤小鼠模型中，观察叶酸是否通过线粒体损伤途径影响高同型半胱氨酸血症小鼠急性肾损伤的发生、发展。

急性肾损伤（AKI）严重危害人类健康，已是全球公共卫生的重要挑战。我们既往的研究发现在AKI小鼠中，高同型半胱氨酸血症(HHcy)可能通过增加内质网应激及减少Akt磷酸化，促进肾小管上皮细胞的凋亡及减少细胞的增殖。但HHcy是否通过其它途径加重AKI以及叶酸的干预是否可改善HHcy小鼠的急性肾损伤尚不明确。为了探讨上述问题，本项目建立了小鼠的HHcy模型及顺铂诱导的AKI模型，检测血清肌酐、肾小管凋亡细胞数，肾小管上皮细胞线粒体损伤评分，肾脏线粒体DNA及cleaved-caspase3、caspase12、γH2AX、细胞色素C和ATP合酶β的表达；予同型半胱氨酸（Hcy）或/和线粒体损伤保护剂（mdivi-1）刺激肾小管上皮细胞，检测细胞凋亡、线粒体DNA、γH2AX、细胞色素C的表达；并进一步在HHcy的AKI小鼠中给予叶酸、mdivi-1干预，观察Hcy、血清肌酐、肾小管损伤评分、肾小管上皮细胞凋亡及线粒体损伤情况。研究结果显示：1）在AKI小鼠中，与Hcy正常的小鼠相比，HHcy小鼠血清肌酐、肾小管损伤评分、肾小管上皮细胞的凋亡以及肾小管上皮细胞线粒体损伤评分明显增加，肾脏线粒体DNA水平明显降低，ATPβ的表达略低，γH2AX的表达明显升高，且Cytc向胞浆的释放增加。2）Hcy刺激NRK细胞16h后，细胞中凋亡相关分子（caspase3、caspase12）以及DNA损伤标志物（γH2AX）的表达明显升高，ATPβ的表达明显降低，Cytc向胞浆的释放增加；而Mdivi-1干预能明显降低Hcy诱导的细胞凋亡、线粒体损伤以及DNA损伤。3）予叶酸、线粒体保护剂（mdivi-1）干预后，与HHcy+AKI小鼠相比，HHcy+AKI+叶酸小鼠及HHcy+AKI+Mdivi-1小鼠血清肌酐、肾小管损伤评分、肾小管上皮细胞的凋亡及线粒体结构损伤减少，肾脏线粒体DNA水平升高，ATPβ的表达增加，γH2AX的表达减少，Cytc向胞浆的释放减少，且HHcy+AKI+叶酸小鼠血清Hcy减低。本项目的结果证实HHcy可通过促进线粒体损伤，增加肾小管上皮细胞的凋亡，加重急性肾损伤；叶酸的干预可降低血清Hcy水平，改善线粒体损伤，减少肾小管上皮细胞的凋亡，减轻HHcy小鼠急性肾损伤。该研究结果可能为急性肾损伤的防治提供新的靶点。