CD36膜转位抑制线粒体脂肪酸氧化在糖尿病肾病肾小管间质炎症中的作用及机制研究

Renal tubular interstitial inflammatory is an important pathogenesis of diabetic nephropathy. Reduction of fatty acid oxidation promotes the progression of tubulointerstitial inflammation, but its mechanism is unclear. It has been reported that the translocation of CD36 to plasma membrane can regulate mitochondrial fatty acid oxidation and maintain cell metabolic balance. In our previous study, we have found that the expression of CD36 in high-glucose cultured renal tubular cells, especially in plasma membrane, was increased, and promoted inflammatory response. However, it is not clear that whether CD36 mediates this effect by regulating fatty acid oxidation. In this study, the relationship between the localization of CD36 in tubular cells and the expression of fatty acid oxidation, inflammation/fibrosis markers in renal tissue of patients with diabetic nephropathy were analyzed using the tissue microenvironment multi-scale total landscape analysis system. Experimental techniques such as immunoco-localization were used to demonstrate the existence of CD36 on the plasma membrane of renal tubular cells and its regulation by palmitoylation. Overexpression and inhibition of CD36 translocation in renal tubular epithelial cells, genetically modified mice with specific high expression of CD36 and specific knockout of the palmitoylation modification site of CD36 in renal tubular cells were constructed, respectively. Using in vivo and in vitro experiments confirmed that CD36 palmitoylation induced translocation, activate AMPK signaling pathway, regulate mitochondrial fatty acid oxidation of renal tubular cells, and promote the inflammatory response of diabetic tubulointerstitium. This study would explore new pathways and molecular mechanisms of renal interstitial inflammation in diabetes mellitus, and provide new ideas and targets for preventing renal damage in diabetes mellitus from the perspective of anti-inflammatory.

肾小管间质炎症反应是糖尿病肾病的重要发病机制。脂肪酸氧化减少促进小管间质炎症进展，但其发生机制未明。文献报道CD36膜转位能够调节线粒体脂肪酸氧化，参与细胞代谢性炎症。我们前期研究发现CD36在高糖培养的肾小管细胞膜表达增加，且参与肾小管炎症反应，但CD36是否通过调控脂肪酸氧化介导该作用尚不明确。本项研究将应用组织微环境多标全景观分析系统分析糖尿病肾病患者肾组织CD36在小管细胞的定位与脂肪酸氧化、炎症/纤维化标志的表达关系，利用免疫共定位等实验技术证实CD36存在膜转位且受棕榈酰化调节。过表达和抑制肾小管上皮细胞CD36膜转位，分别构建肾小管细胞特异性高表达CD36和特异性敲除CD36棕榈酰化修饰位点的基因修饰小鼠，体内外实验证实CD36棕榈酰化修饰引起膜转位调控AMPK信号通路，调节肾小管细胞线粒体脂肪酸氧化，促进糖尿病肾小管间质炎症反应。解析糖尿病肾间质炎症反应的新途径及分子机制。

肾小管间质炎症反应是糖尿病肾病（DKD）的重要发病机制。肾小管上皮细胞（TECs）脂肪酸氧化（FAO）障碍促进小管间质炎症进展，但在DKD中的作用和机制未明。文献报道CD36膜转位能够调节线粒体FAO，参与细胞代谢性炎症。所以本项目旨在研究CD36通过调节TECs代谢，促进糖尿病小管间质炎症反应的作用及机制探讨。主要研究结果如下：1）高糖环境诱导TECs能量代谢重编，即线粒体FAO减少，糖酵解增强；TECs能量代谢重编促进糖尿病肾小管间质炎症；药物改善TECs线粒体FAO及药物保护线粒体均能够逆转糖尿病肾小管间质损伤。2）抑制CD36表达，敲除CD36基因，以及抑制CD36细胞膜定位（CD36棕榈酰化位点突变）均可改善DKD小管间质炎症/纤维化损伤，延缓肾病进展。3） 抑制CD36表达能够上调线粒体FAO，逆转糖尿病TECs能量代谢重编，通过减少线粒体ROS生成，抑制小管上皮细胞模式识别受体（NLRP3，TLR2/4）启动小管间质炎症反应。4）CD36棕榈酰化能够抑制小管上皮细胞模式识别受体（NLRP3，TLR2/4）启动小管间质炎症反应，但该作用不通过改善线粒体FAO。5）CD36能够介导线粒体核糖体缺陷，从而损害线粒体氧化磷酸化，促进糖尿病肾病小管间质纤维化进展。6）CD36是维持糖尿病肾小管上皮细胞损伤持续进展的因素之一，参与DKD代谢记忆。本项目通过阐明TECs能量代谢重编参与DKD小管间质炎症损伤的机制，证实CD36在DKD发生和发展中的关键作用，为临床防止DKD提供了理论依据。本项目已发表SCI论文2篇（Cell Death Dis. 1篇；Front Pharmacol. 1篇），在投稿中SCI论文2篇（Redox Biol.1篇；NDT 1篇），撰写中论文3篇。