化瘀通络中药全方及拆方对DN大鼠血液相关指标及肾脏足细胞ZO-1表达的干预作用

Effect of Combined Prescription and Separate Prescription of Huayu Tongluo Traditional Chinese Medicine on Blood Correlation Index and the Expression of ZO-1 in Renal Podocytes of DN Rats

通过化瘀通络中药全方及拆方对糖尿病肾病(DN)大鼠的24 h尿蛋白定量(UTP)、血脂、血小板活化指标及肾脏足细胞ZO-1的调节作用,探讨化瘀通络中药全方、拆方的不同作用特点及防治DN的可能作用机制。将健康雄性SD大鼠60只分为正常组(C组)10只,造模组50只。造模组大鼠选择高脂高糖饲料喂养联合腹腔注射链脲佐菌素(STZ,1%)35 mg/kg复制糖尿病(DM)大鼠模型,72 h后检测血糖,将成模大鼠(血糖≥16.7mmol/L)随机分为模型组(M组)、化瘀通络中药全方组(Q组)、通络中药组(T组)、化瘀中药组(H组)。按人/鼠体表面积比率换算等效计量法计算各组大鼠用药量,同时C、M两组大鼠灌以等量饮用水,1次/日,连续16周,分别检测各组大鼠24 h UTP、血脂、血小板活化指标等;采用实时荧光定量(Real-time PCR)、蛋白印迹(Western-Blot)方法检测各组大鼠肾组织足细胞ZO-1的基因及蛋白表达情况。实验结果,与C组比较,其余各组大鼠血糖、24 h UTP明显升高(P<0.01),血脂、血小板活化指标出现明显变化(P<0.05,P<0.01);与M组相比,各中药组24 h UTP均明显减少(P<0.01),血脂、血小板相关指标明显改善(P<0.05,P<0.01);中药各组间比较,Q组部分指标的改善优于T组(P<0.05)。与C组相比,其他各组大鼠足细胞裂孔膜蛋白ZO-1表达水平均明显降低(P<0.01);与M组相比,各中药组大鼠足细胞裂孔膜蛋白ZO-1表达水平均明显升高(P<0.01);与Q组比较,T、H组ZO-1基因和蛋白表达水平均明显降低(P<0.05,P<0.01)。结果表明化瘀通络中药可以降低DN大鼠24 h UTP,其机制可能与调节脂质代谢紊乱,改善血凝状态,上调足细胞裂孔膜蛋白ZO-1的表达有关,并且,化瘀中药与通络中药联合应用有协同作用,为临床优化中药配伍提供实验依据。

To investigate the effect of combined prescription and separate prescription of Huayu Tongluo traditional Chinese medicine upon 24 h urine protein quantitation,blood lipid,platelet activation index and the expression of ZO-1 in podocytes of rats with diabetic nephropathy. Totally 60 male sprague-dawley rats were randomly selected as the normal control group（ C group,n = 10）,the other 50 rats were injected with 1% streptozotocin（ 35 mg/kg） after feeding high fat and high glucose diet for one month. The diabetic model rats（ blood glucose≥16. 7 mmol/L） were randomly divided into the model group（ M group）,Huayu Tongluo traditional Chinese medicine group（ Q group）,Tongluo group（ Tgroup）,Huayu group（ H group）. According to the body surface area ratio,the dosage of rats in each group was calculated. The rats in C group and M group were fed with the equal volume of drinking water,once daily for 16 weeks. 24 h urine protein quantitation（ 24 h UTP）,blood lipid and platelet activation index of the rats were detected at the end of the16 th week. The protein and mRNA expression of ZO-1 in kidney tissue were observed by Western blotting and Real-time PCR methods. Compared with C group,blood sugar,24 h UTP were significantly increased in other groups（ P〈0. 01）.Blood lipid and platelet activation index were significantly changed（ P〈0. 05,P〈0. 01）. Compared with M group,24 h UTP in each group of Chinese medicine was significantly reduced（ P〈0. 01）. Blood lipid and platelet activation index were improved significantly（ P〈0. 05,P〈0. 01）. Compared with C group,the expression of ZO-1 reduced in other groups（ P〈0. 01）. Compared with M group,the expression of ZO-1 were significantly higher in each group of Chinese medicine（ P〈0. 01）. Compared with Q group,the expression of ZO-1 was significantly reduced in H,T group（ P〈0. 05,P〈0. 01）. The results indicated that Huayu Tongluo Chinese medicine can reduce 24 h UTP in DN rats. The mechanism may be related to the regulation of lipid metabolism disorder,the improvement of blood clotting state and the up-regulation of the expression of ZO-1 and Huayu Chinese medicine and Tongluo Chinese medicine have synergistic effect to provide experimental basis for the compatibility of clinical Chinese medicine.