化瘀通络中药通过TAK1/JNK通路抑制巨噬细胞浸润及活化改善糖尿病肾病大鼠肾脏炎症

Huayu Tongluo herbs alleviate renal inflammation by inhibiting macrophage infiltration and activation via TAK1/JNK pathway in diabetic nephropathy rats

观察化瘀通络中药对糖尿病肾病大鼠肾脏炎症的干预作用,并探讨其机制。40只SD大鼠随机选取10只为正常组(C组),其余30只采用一次性腹腔注射链脲佐菌素(STZ,40 mg/kg)联合高糖高脂饲料喂养建立糖尿病模型。造模成功大鼠随机分为模型组(M组)和化瘀通络中药组(Z组)。Z组予以中药干预。16周后,各组大鼠留尿检测24 h尿蛋白定量(24 h UTP);ELISA法检测血清中单核细胞趋化蛋白-1(MCP-1)、白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的表达;取肾组织,应用免疫荧光法检测巨噬细胞标志蛋白CD68、经典活化巨噬细胞标志蛋白诱导性一氧化氮合成酶(iNOS)的表达,应用Western blot法检测TAK1/JNK信号通路相关蛋白转化生长因子激活激酶-1(TAK1)、p-TAK1、c-Jun氨基末端激酶(JNK)和p-JNK的表达。实验结果显示,与C组比较,M组大鼠24 h UTP、MCP-1、IL-1β和TNF-α水平均明显升高(P<0.001);肾间质可见大量CD68和iNOS蛋白的沉积;p-TAK1和p-JNK蛋白的表达量明显增多(P<0.001)。与M组比较,Z组大鼠24 h UTP、MCP-1、IL-1β和TNF-α水平均显著降低(P<0.001,P<0.001,P<0.01,P<0.001);CD68和iNOS蛋白的沉积显著减少;p-TAK1和p-JNK蛋白的表达量均显著减少(P<0.05)。结果表明化瘀通络中药可降低糖尿病肾病大鼠蛋白尿,改善肾脏炎症反应,其机制可能与其抑制TAK1/JNK信号通路减少肾组织内巨噬细胞浸润、活化和炎症因子释放相关。

To observe the effect of Huayu Tongluo herbs on renal inflammation in diabetic nephropathy(DN)rats and explore its mechanism.Ten SD rats were randomly selected as normal control group(C group),and the other 30 rats were fed with one-time intraperitoneal injection of streptozotocin(STZ,40mg/kg)combined with high-glucose and high-fat diet to establish diabetic model.The successfully modeled rats were randomly divided into model group(M group)and Huayu Tongluo herbs group(Z group).Z Group was treated with traditional chinese medicine.After 16 weeks,24 h urine total protein(24 h UTP),monocyte chemotactic protein 1(MCP-1),interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)were detected by ELISA method,the expression of CD68 and inducible nitric oxide synthase(iNOS)was detected by Immunofluorescence,and the expression of TAK1/JNK pathway related proteins transforming growth factorβ-activated kinase 1(TAK1),p-TAK1,c-Jun N-terminal kinase(JNK)and p-JNK was detected by Western blot method.The results showed that compared with C group,the levels of 24 h UTP,MCP-1,IL-1βand TNF-αwere significantly increased(P<0.001),a large number of CD68 and iNOS protein deposits were seen in renal interstitium,and the expression of p-TAK1 and p-JNK protein was significantly increased in M group(P<0.001).Compared with M group,the levels of 24 h UTP,MCP-1,IL-1βand TNF-αwere significantly decreased(P<0.001,P<0.001,P<0.01,P<0.01),the deposition of CD68 and iNOS protein was significantly decreased,and the expression of p-TAK1 and p-JNK protein was significantly decreased in Z group(P<0.05).The results show that Huayu Tongluo herbs can reduce proteinuria and alleviate renal inflammation in DN rats,and its mechanism may be related to its reduction of macrophage infiltration and activation,and release of inflammatory factors in renal tissue via inhibiting TAK1/JNK pathway.