糖尿病肾病（DN）是糖尿病常见的慢性微血管并发症，也是引起终末期肾功能衰竭的最重要原因之一。肾脏高表达细胞因子信号传导抑制蛋白-1(SOCS-1)能够减轻糖尿病早期肾脏病变及减轻肾脏的炎症，在DN病变中起到重要的保护作用。荔枝核是治疗DN的传统中药，本项目拟采用建立在体小鼠糖尿病肾病模型和体外诱导肾小球系膜细胞，通过免疫组化、ELISA、Western blot和实时定量PCR等实验方法，从核酸和蛋白两个水平研究荔枝核活性成分荔枝核皂苷影响SOCS-1下调JAK/STAT 信号通路，进而延缓DN进程的作用机制，为临床应用中药防治DN提供理论与实验依据，有助于荔枝核皂苷成为具有自主知识产权的以SOCS-1为靶点的DN治疗药物，具有重要的科学意义和广阔的应用前景。

Diabetic nephropathy is an important microvascular complication of diabetic mellitus as well as the common reason resulting in chronic renal failure. suppressors of cytokine signaling-1 could ameliorate renal pathological changes and Inflammation,and SOCS has protective functions against renal pathological changes. Semen litchi,one of traditional Chinese medicinal, was mainly prescribed in treatment of diabetic nephropathy. This project aims to establish Mouse models of diabetic nephropathy and In vitro activated glomerular mesangial cell model, to study on mechanisms of Influence of saponin of litchi seed on SOCS-1 and Suppress JAK/STAT Pathway to Postpone the process of Diabetic nephropathy through immunohistochemistry, ELISA, Western blot and real-time quantitative PCR and other experimental methods, to provide theoretical and experimental evidence for preventing and treating diabetic nephropathy,and can assist saponin of litchi seed becoming an attractive therapeutic agent with independent intellectual property right for the treatment of fibrotic diabetic nephropathy with SOCS-1 as target.The study would have important scientific significance and application value.

本项目按计划进行了体内与体外实验的系统研究，建立了肾细胞高糖及相关细胞因子诱导活化模型，观察了荔枝核皂苷（saponin of litchi seed，SA）干预作用。完成了AngII、TNF-α对HKC增殖影响的最佳浓度的选择。20mg/L SA对TNF-α刺激的HKC具有明显的干预作用（P＜0.05）。TNF-α对HKC有促增殖作用, 且能被SA所拮抗。观察了SA对AngII诱导的HMC增殖及IL-6、IL-1β影响，结果SA对HMC增殖抑制作用呈浓度和时间依赖性。进行了SA对高糖诱导的HMC增殖和凋亡的作用机制研究，通过流式Annexin V-FITC/PI双染法检测细胞凋亡，结果高糖组细胞凋亡受到抑制(P<0.01)；SA各组明显促进HMC的凋亡（P<0.01），随浓度增加凋亡率亦增加。在体实验选择8周龄CD-1小鼠实验，结果模型小鼠血糖及24h尿蛋白可达标，但死亡率较高。实验中发现，与模型组比较，SA高剂量组及阳性对照组表达均显著降低(P<0.01 )。HE染色发现阳性对照组及SA各组与模型组比较均有显著性变化，SA高剂量组改变最为明显。Masson染色亦支持此结果。电镜检查结果：阳性对照组及SA组与模型组比较病变有所减轻。免疫组化法检测了α-SMA、TGF-β1 的表达情况，模型组a-SMA表达明显增多，阳性对照组与SA高剂量组TGF-βl有较弱的表达；模型组TGF-βl胞质内表达呈强阳性。观察了SA对小鼠肾脏SOCS-1表达的影响，经高糖刺激HKC后SOCS-1蛋白表达水平有所降低，TGF-β1 及FN炎症因子表达明显升高；阳性干预组可显著提高HKC细胞内SOCS-1蛋白表达，降低TGF-β1及FN 表达。在体动物实验中，模型组SOCS-1的表达量明显升高，与模型组相比较，阳性对照组及SA各组蛋白表达量有所下降。本项目为临床应用中药防治DN提供了理论与实验依据，提示我们通过抑制细胞增殖，降低相关细胞因子的表达，减少ECM积聚对防治肾小球硬化具有重要意义。