| Objective:To observe the clinical efficacy of Danzhi Jiangtang capsule on patients with diabetic kidney disease(DKD)and its effect on the expression of inflammatory signaling pathway-related proteins and gene expression and intervention,and to investigate the underline mechanism of Danzhi Jiangtang capsule in treating DKD.Methods:1.Clinical observation: 80 selected patients with T2 DKD were divided into2groupes(observation group 40 patients,and control group 40 patients).They all were given diabetes education,control of their diet,exercise therapy,lowering of blood glucose and blood pressure,and regulation of blood lipids.Both groups were given oral valsartan80 mg once/d.In the observation group,on top of valsartan,Danzhi Jiangtang capsules were given at the same time,5 capsules each time,3 times a day,for 12 weeks.Patients’ FPG,2h PG,UAER,NAG,m ALB,BUN and Scr were measured.2.The experiment research: Sixty-five male SPF-grade healthy male SD rats were used as experimental subjects,and all rats were fed with normal chow for one week,10 of which were randomly selected as the normal group,while the rest of the rats were modeled.The control group was fed normal chow and injected with citric acid-sodium citrate buffer(p H 4.2)after one week,while the modeling group was fed normal chow for one week and started to be fed with high sugar and high fat for six weeks,then the modeling rats were injected with 1% STZ solution(35 mg/kg)for the first time.Finally,50 rats were successfully modeled,and they were randomly averaged divided into: model control group,low-dose group(270 mg/kg),medium-dose group(540 mg/kg),high-dose group(1080 mg/kg)and positive group(valsartan);the normal rats without modeling were normal control group.For the rats in the normal and model groups,distilled water was given,while the other dosing groups were fed twice daily for 8 weeks according to the established dosage.During the feeding of the rats,the rats in each group were regularly observed for feeding and water intake,body growth status,and detection of body weight changes.Fasting blood glucose(FPG)was collected from the tail tip of the rats at week 0,week 4 and week 8 of feeding,and aortic blood was collected after the last feeding.The changes of FPG and Hb A1 c were measured by glucometer and automatic biochemical analyzer.After the last week of drug feeding,the blood collected from the aorta of rats was fasted for 12 hours,and indicators such as Scr and BUN were measured;after the rats were anesthetized,the kidneys of rats were removed,while they were dehydrated(different concentrations of alcohol 75%-100%),paraffin-embedded and sectioned to form 4-μm paraffin sections,and stained with three kinds of different ways(HE,TUNEL and PASM).The observation on pathological characteristics of rat kidney tissue sections were made with light microscope.Western blot method was used to detect the expression of relevant proteins in rat kidney tissues,and RT-q PCR was used to study the effect of Danzhi Jiangtang capsule on the expression of genes related to inflammatory signaling pathway and its intervention in rat kidney tissues.Results:1.Clinical study results: Compared with the pre-treatment period,the TCM evidence points of both groups decreased;the blood routine indexes such as FPG,2h PG and Hb A1 c of the treatment group decreased significantly after treatment as compared with them before treatment,and the glucose-lowering effect of the observation group that took oral valsartan and then combined with Danzhi Jiangtang capsule The glucoselowering effect of the observation group combined with valsartan was better than that of the control group taking valsartan alone.After treatment,the urinary albumin excretion rate(UAER),serum creatinine(Scr)and blood urea nitrogen(BUN)in the group treated with Danzhi Jiangtang Capsule decreased significantly,and the degree of reduction was better than that in the control group.2.Animal experiment results: The rats in the model group(after successful modeling)were significantly higher before and after treatment,while compared with the normal group,their blood glucose was significantly higher before and after treatment;compared with the normal group,the FPG of rats in all modeling groups in type 2 diabetes was obviousely higher than normal group;compared with the model group,the blood glucose levels in all drug intervention groups were lower,but the Danzhi Jiangtang capsules were lower in all dose groups than the valsartan group,and the high dose group had the blood glucose level was the lowest.Compared with the model group,Scr and BUN were reduced in all dosing groups,and compared with the valsartan group,Scr and BUN were also reduced in the Danzhi Jiangtang capsule group,and the reduced level increased with the increase of the dose of Danzhi Jiangtang capsule;in addition,the proteinuria level of rats in all dosing groups compared with the model group decreased after treatment,and the rats in the Danzhi Jiangtang capsule intervention group had lower proteinuria level than the rats in the valsartan group.The level of proteinuria was lower in the rats in the intervention group than in the rats in the valsartan group,and the level of proteinuria decreased with the increase of the dose of Danzhi Jiangtang capsules.The data of HE and PASM revealed that the glomerular and tubular lesions were relieved to a certain extent in the drug intervention group,especially in the high dose group of Danzhi Jiangtang Capsules.The difference of renal pathological structures of rats under optical microscope showed that Danzhi Jiangtang capsule could improve renal injury in rats.The results of Western blot assay showed that the drug intervention led to significant expression of β-Actin,P-IκB Alpha,TLR4,caspase-8,My D88,NF-κB and MAP3/TAK1 proteins in the kidney tissues of type 2 diabetic rats;the result of P-IκB Alpha protein relative expression was significantly increased.In contrast,TLR4 mRNA,My D88 mRNA,caspase-8 mRNA,MAP3 mRNA,IKBa mRNA and NF-κB p65 mRNA expression in rat kidney tissues were reduced to different degrees after 8 weeks of administration of Danzhi Jiangtang capsules and valsartan;compared with the model group,there were significant The difference was significant in the model group.It indicates that the inflammatory signaling pathway was activated in the kidney tissues of rats with diabetic nephropathy model,and Danzhi Jiangtang capsules could effectively inhibit the overexpression of inflammatory signaling pathway genes.Conclusions:The clinical observation results showed that Danchuang Hypoglycemic Capsules had a significant effect on improving the symptoms of patients with type 2 diabetic nephropathy in the early stage;animal experiments showed that Danchuang Hypoglycemic Capsules could inhibit the inflammatory process in the kidney tissue of rats with diabetic nephropathy;through the detection of the expression of genes and proteins related to the inflammatory response pathway in rats,it was found that Danchuang Hypoglycemic Capsules could inhibit the expression of inflammatory factors. |
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