益气解毒活络方对早期糖尿病肾病大鼠TGF-β/Smad信号通路的影响

Effect of Yiqi Jiedu Huoluo Formula on TGF-β / Smad Pathways in Rats with Early Diabetic Nephropathy

目的:研究益气解毒活络方(YQJDHL)对早期糖尿病肾病(DN)大鼠的治疗作用及对转化生长因子-β(TGF-β)/信号转导蛋白(Smad)信号通路的影响。方法:SPF级健康雄性SD大鼠72只,尾静脉注射链脲佐菌素(STZ)制造糖尿病(DM)模型,再根据大鼠血糖值高低对各治疗组进行随机分组,分别为模型组,YQJDHL预防组(2.4 g·kg-1·d-1),YQJDHL低、高剂量治疗组(2.4,7.2 g·kg^(-1)·d^(-1)),阳性药组(盐酸贝那普利,10 mg·kg^(-1)·d^(-1)),另设正常组,每组12只,YQJDHL预防组在成DM模型后立即给予灌服YQJDHL复方;YQJDHL低、高剂量组及西药组均在成DM模型2周后即成DN模型后灌服YQJDHL复方。在灌服YQJDHL 4周末收集大鼠24 h尿液待测尿微量白蛋白,腹主动脉穿刺取血测定大鼠血糖,留取肾组织标本,苏木素-伊红(HE)染色观察肾组织病理学的变化,逆转录-聚合酶链式反应(RT-PCR)法及蛋白质免疫印迹(Western blot)法观察YQJDHL对早期DN大鼠肾组织转化生长因子-β1(TGF-β1),Smad3,Smad7 mRNA及蛋白表达的影响。结果:与正常组比较,模型组大鼠血糖及尿微量白蛋白均明显升高,TGF-β1及Smad3表达明显升高,Smad7表达明显降低(P<0.01),病理学观察显示肾组织病变较明显;YQJDHL治疗组及预防组均能降低大鼠血糖及尿微量白蛋白,降低大鼠肾组织TGF-β1及Smad3表达,升高Smad7表达(P<0.05,P<0.01),肾组织病变明显改善。结论:YQJDHL能够预防和治疗DN,其作用机制可能与TGF-β/Samd信号通路有关。

Objective： To investigate the treatment effect of Yiqi Jiedu Huoluo formula（ YQJDHL） in rats with early diabetic nephropathy（ DN） and observe its effect on transforining growth factor-β（ TGF-β） / Smad pathways. Method： Totally 72 SPF healthy male SD rats were used to establish diabetes mellitus（ DM） models by tail injection of streptozotocin（ STZ）. Then according to the blood sugar levels,the rats were randomly divided into model group,YQJDHL prevention group（ 2. 4 g·kg^-1·^-1）,YQJDHL low dose group and high dose group（ 2. 4,7. 2 g·kg^-1·^-1）,positive drug group（ benazepril hydrochloride,10 mg·kg^-1·^-1）,another normal group was also set up,n = 12 in each group. In the YQJDHL preventive group,YQJDHL compound formula was immediately given after DM models were made. While in YQJDHL high and low dose groups and positive drug group,the medicine was given two weeks after the DM models were made,that was immediately after DN models were made. Four weeks after intragastrical administration of YQJDHL formula,24 h urine of rats was collected to determine the urine trace albumin, and the blood was taken by abdominal aorta puncture to determine blood glucose in rats; renal tissue samples were collected to observe the renal pathological changes by htoxylin-eosin（ HE） staining method; the effects of YQJDHL TGF-β1,Smad3 /7 mRNA and protein expression levels in the kidney tissues of early DN rats were observed by reverse transcription-polymerase chain reaction（ RT-PCR） and Western blot assay. Result： As compared with the normal group,the blood glucose and urine trace albumin were significantly increased in rats of model group; TGF-β1and Smad3 expression levels were significantly increased;Smad7 expression level was significantly reduced（ P〈0. 01）; and the renal pathological changes were obvious in pathological observation. QJDHL treatment groups and preventive group could reduce the blood glucose and urine trace albumin in rats, reduce TGF-β1and Smad3 expression levels in kidney tissues, and increase Smad7expression（ P〈0. 05, P〈0. 01）; in addition, renal pathological changes were significantly improved.Conclusion： The results showed that the YQJDHL formula had the treatment and prevention effect for the DN,and its mechanism of action may be associated with TGF-β / Smad signaling pathways.