A Study On The Mechanism Of Shengfu Xiongze Decoction On Oxidative Stress In Rats With Chronic Heart Failure Based On The Keap1-Nrf2-ARE Pathway

ObjectiveObserve the effects of Shenfu Xiongze Decoction on oxidative stress in rats with chronic heart failure,and explore the effect of Shenfu Xiongze Decoction on oxidative stress in rats with chronic heart failure based on Keapl-Nrf2-ARE pathway.MethodsEstablishment of chronic heart failure rat model by intraperitoneal injection of adriamycin（ADR）.After one week of adaptive feeding,65 SD rats were randomly divided into normal group（n=10）and doxorubicin group（n=55）.The adriamycin group was intraperitoneally injected with a adriamycin 2 mg/ml solution once a week for 4 mg/kg in the first 3 weeks,1 mg/kg in each of the following 3 weeks,and a cumulative dose of 15mg/kg.Normal saline was intraperitoneally injected into the normal group 1 time a week for 6 weeks.After 6 weeks,5 rats were randomly selected from the adriamycin group to observe the general state and the shape of the heart.HE staining and plasma BNP were carried out to confirm whether the model was successful or not.After the success of the model,the normal group and remaining adriamycin group rats were divided into normal control group（N）,model group（MOD）,Vasorel group（V）,Shenfu Xiongze Decoction low dose group（L）,Shenfu Xiongze Decoction middle dose group（M）and Shenfu Xiongze Decoction high dose group（H）.The dosage is calculated according to the formula of the equivalent dose conversion between adult and rat.V group was given trimetazidine hydrochloride as 7mg/kg/d,the concentrations of Shenfu Xiongze Decoction in groups H,M,and L were 6.4g/ml,3.2g/ml,and 1.6g/ml,respectively,and they were intragastrically administered at 5ml/kg/d.N and MOD groups were given equal doses of 0.9%normal saline for 8 weeks.Then,left ventricular end diastolic pressure（LVEDP）,left ventricular end systolic pressure（LVSP）,left ventricular pressure maximum rise rate(+dp/dt_max),and left ventricular pressure maximum descending rate(-dp/dt_max)were recorded by BL-420A-type biological signal collecting and processing system via right common carotid artery intubation.The rats were sacrificed to observe the pathological changes in the myocardium by HE staining.The levels of BNP,ROS were detected by ELISA method.MDA content was determined by TBA method.The activity of SOD enzyme was detected by WST-1 method.Also,the expression of Nrf2 and HO-1 protein were detected by immunohistochemical staining and Western blot method.Results1.Haemodynamics showed that LVEDP in MOD group increased significantly,LVSP and±dp/dt_maxax decreased significantly,and the treatment groups could reduce LVEDP and enhance LVSP and±dp/dt_maxax at different degrees,the differences were statistically significant.2.HE staining showed that the myocardial fibers in N group were arranged neatly,and the cells were regular,without swelling,atrophy and no inflammatory cell infiltration.The cardiac fibers in the MOD group were disordered,broken or disappeared,and a great deal of inflammatory cells infiltrated.There were different degrees of swelling and inflammatory cell infiltration in the myocardial fibers of L group and M group.The myocardial fibers in H group and V group were arranged more neatly,no obvious edema,and occasionally inflammatory cell infiltration.3.The results of ELISA showed that the content of plasma BNP in MOD group was significantly increased,there was significant difference with N group.The content of BNP of Shenfu Xiongze Decoction groups and Vasorel group were lower than those of MOD group,and the differences are obvious.The content of plasma ROS in MOD group was significantly higher than that in N group,while ROS in H and V groups were significantly lower than that in MOD group,and the differences were statistically significant.4.The results of MDA and SOD in plasma showed that compared with N group,the content of MDA in MOD group was significantly increased and the activity of SOD enzyme decreased.Compared with the MOD group,the content of MDA in H and V groups decreased significantly and the activity of SOD enzyme increased,and the differences were statistically significant.Compared with the V group,there was no significant difference in MDA and SOD in M and H groups.5.The results of immunohistochemistry showed that compared with the N group,the expression of Nrf2 and HO-1 decreased in MOD group,L group,M group,H group,Vgroup,and the expression of Nrf2 and HO-1 in each therapy group was significantly higher than that in the MOD group.The expression of Nrf2 and HO-1 in H group was compared with that in V group,the difference was not statistically significant.6.The results of Western blot showed that the expression of Nrf2 in MOD,L and M groups decreased significantly compared with that of N group,and the expression of HO-1protein in each group decreased significantly.Compared with the MOD group,the expressions of Nrf2 and HO-1 protein in each therapy group increased,and the differences were statistically significant.It was shown that Shenfu Qize Decoction can increase the expression of Nrf2 and HO-1 protein and protect myocardium.Conclusions1.Shenfu Xiongze Decoction can improve cardiac function in chronic heart failure rats,exert an antioxidant effect.2.This mechanism may be associated with the up-regulation of the expression of Nrf2in the Keap1-Nrf2-ARE pathway,which activates the antioxidant enzymes.