Supplementary Material for: Pterostilbene, a resveratrol derivative, improves ectopic lipid deposition in the kidneys of mice induced by a high-fat diet

Background: Diabetic kidney disease is a major cause of global end-stage renal diseases. Ectopic lipid deposition in the renal tissues of diabetic kidney disease is one major factor leading to renal fibrosis and chronic kidney disease. Pterostilbene has been reported to display lipid-lowing activity and participate in many kidney diseases. However, the influence of pterostilbene on the ectopic lipid deposition is unclear. We intend to explore the influence of pterostilbene on the ectopic lipid deposition in the kidney of mice induced by high fat. Methods: A high-fat diet-induced diabetic mouse model was established to detect the alleviative effect of pterostilbene on the ectopic lipid deposition in the kidney of diabetic mice. Biochemical analysis was performed to examine the level of urine albumin, urine creatinine, serum creatinine and blood urea nitrogen in mice after pterostilbene treatment. Histological analysis was conducted to detect the degree of renal injury and fibrosis. Oil red O staining and immunohistochemical staining were carried out to evaluate lipid droplets and the expression of adipose differentiation-related protein (ADRP) in renal tissues of the mice treated by pterostilbene. The protein levels were assessed by western blotting. Results: Pterostilbene inhibits the expression of the TGF-β1 and p-smad3 and suppresses the protein levels of SREBP-1 and FAS, and ultimately reduces the ectopic lipid deposition and alleviates the renal tubular damage and renal fibrosis in the kidneys of diabetic mice induced by high fat, and improved kidney function. Conclusion: Pterostilbene alleviates renal fibrosis and ectopic lipid deposition in the kidneys of diabetic mice induced by high fat diet by inhibiting the TGF-β1/smad3 signaling.

背景：糖尿病肾病（Diabetic kidney disease）是全球终末期肾病的主要致病原因。糖尿病肾病患者肾组织内的异位脂质沉积是引发肾纤维化与慢性肾病的关键诱因之一。紫檀芪（Pterostilbene）已被证实具有降脂活性，并参与多种肾脏疾病的病理进程，但目前关于紫檀芪对异位脂质沉积的影响尚不明确。本研究旨在探讨紫檀芪对高脂诱导小鼠肾脏异位脂质沉积的影响。

方法：本研究构建高脂饮食诱导的糖尿病小鼠模型，以检测紫檀芪对糖尿病小鼠肾脏异位脂质沉积的改善效果。通过生化分析检测紫檀芪干预后小鼠的尿白蛋白、尿肌酐、血清肌酐及血尿素氮水平。采用组织学分析评估肾损伤与纤维化程度。通过油红O（Oil red O）染色与免疫组织化学染色，检测紫檀芪处理小鼠肾组织内的脂滴及脂肪分化相关蛋白（ADRP）的表达水平。采用蛋白质免疫印迹（western blotting）检测相关蛋白的表达量。

结果：紫檀芪可抑制转化生长因子-β1（TGF-β1）与磷酸化Smad3（p-smad3）的表达，下调固醇调节元件结合蛋白-1（SREBP-1）及脂肪酸合酶（FAS）的蛋白水平，最终减少高脂诱导糖尿病小鼠肾脏内的异位脂质沉积，缓解肾小管损伤与肾纤维化，并改善肾功能。

结论：紫檀芪可通过抑制TGF-β1/Smad3信号通路，改善高脂饮食诱导糖尿病小鼠的肾纤维化与肾脏异位脂质沉积。