| BackgroundIn recent years,with the change of people's ways and the improvement of dietary conditions,the incidence of hypertension in China is also increasing year by year,and tends to be younger.The research on etiology,prevention and treatment of hypertension has already become an important topic.Traditional Chinese medicine(TCM)has the advantages of one person,precise treatment and improving the overall symptoms of patients,which is a potential research direction.Sancao Decoction(SCD)is an empirical prescription of Professor Liu Duzhou of Beijing University of Traditional Chinese Medicine for treating hypertensive liver-fire syndrome.It has been clinically proved for decades that the effect of reducing blood pressure is exact.At present,although a large number of literatures have explained its hypotensive mechanism from the perspective of traditional Chinese and Western medicine,they are neglected in superficial,and have not made in-depth study on the molecular mechanism of its hypotensive effect.Modern studies have shown that there is a close relationship between liver-fire syndrome,hypertension and inflammatory response.Therefore,this study intends to explore the specific biological mechanism of SCD in reducing blood pressure from the perspective of regulation of SCD on inflammatory response.ObjectiveThrough the network pharmacology research,the potential core target of SCD is excavated and its hypotensive pathway is predicted.By observing the effects of SCD on blood pressure,TCM syndromes,left ventricular hypertrophy and regulation of NF-kappa B/NLRP3/IL-1beta signaling pathway in myocardial tissue of hypertensive rats with hyperactivity of liver-fire,the pharmacodynamics of SCD,the protective effect of target organs and the specific mechanism of its antihypertensive effect were clarified in order to provide a new idea for the treatment of hypertension with integrated traditional Chinese and Western medicine.Method1.To explore the relationship among liver-fire syndrome,hypertension and inflammation.2.TCMSP database was used to query SCD components and their corresponding targets,NCBI Gene database was used to retrieve hypertension targets,and the target genes were fused to construct protein-protein interaction(PPI)network.The core target is extracted from PPI network,and the pathway enrichment analysis is carried out by DAVID database,and the specific mechanism of SCD hypotension is screened.3.Spontaneous hypertensive rats(SHR)with hyperactivity of liver-fire syndrome were selected and divided into five groups: model group,captopril group(30mg/kg),high-dose SCD group(21.2g/kg),medium-dose SCD group(10.6g/kg),low-dose SCD group(5.3g/kg).WKY rats were used as blank control group(WKY group),with 10 rats in each group for 8 weeks after drug intervention.The changes of blood pressure,open field test,rotation tolerance time,irritability and macroscopical characterization were observed during treatment.After treatment,cardiac mass index and left ventricular mass index were calculated;left ventricular pathological damage was observed by HE staining;myocardial fibrosis was observed by Masson staining;serum levels of TNF-a and IL-1beta were detected by ELISA;expression of NF-kappa B p65,NF-kappa B p-p65,NLRP3,Caspase-1,ASC and IL-1beta protein in myocardium was detected by Western blot.The expression of NF-kappa B,NLRP3,Caspase-1 and IL-1beta in myocardium was detected,and the integral optical density of NF-kappa B,NLRP3 and IL-1beta in myocardium was detected by immunohistochemistry.Results1.Network pharmacology research showed that 123 target proteins of SCD were obtained by searching TCMSP and further screening,and 116 core target proteins of SCD were identified by PPI network map.Finally,through KEGG pathway analysis,the potential regulatory pathway of SCD was identified as NF-kappa B/NLRP3/IL-1beta signaling pathway,and subsequent research was carried out.2.The influence on blood pressure and syndrome of hyperactivity of liver fire: after 8 weeks of treatment,compared with model group,(1)SCD high and middle dose groups can significantly reduce SHR arterial systolic and diastolic blood pressure;(2)heart rate and weight among the treatment groups are not statistically different;(3)the pass times of open-field experiment in middle dose group are significantly reduced,while there is no difference in other groups.There is no difference in the stand times of open-field experiment among the treatment groups.Difference;(4)High dose group and medium dose group can significantly increase the rotation tolerance time of SHR;(5)There is no significant difference in irritability score among the treatment groups;(6)High dose group and medium dose group can significantly increase the macroscopical characterization score of rats.3.The effects on left ventricular hypertrophy:(1)The results of cardiac weight index showed that there was no significant difference among the treatment groups,and the results of left ventricular weight index showed that compared with the model group,the captopril group and the middle dose group had a significant decrease;(2)HE staining results showed that the left ventricular tissue of the model group showed a significant increase in the transverse diameter of myocardium,disordered arrangement of muscle fibers,partial rupture of the heart.The arterial wall of intermuscular small vessels was thickened,accompanied by fibrous tissue proliferation and inflammatory cell infiltration.Compared with the model group,the pathological damage of captopril group and SCD high and low dose group was significantly reduced.(3)Masson staining results showed that the blue collagen fibers in the muscle interstitium of the model group were significantly increased,while the number of collagen fibers in captopril group and SCD high and low dose group appeared different degrees.(4)Collagen volume fraction measurement showed that the integral of captopril group and middle dose group decreased significantly.4.The effects on the signal pathway of NF-kappa B/NLRP3/IL-1beta:(1)The results of serum TNF-a and IL-1beta showed that the levels of TNF-a and IL-1beta in the model group were significantly higher than those in the WKY group;compared with the model group,the levels of TNF-a in the high,middle dose group and captopril group were significantly lower,and the levels of IL-1beta in the high and middle dose group were significantly lower;(2)WB results showed that the levels of NF-kappa B P in the model group were significantly higher than those in the WKY group.The expression of 65,NF-kappa B p-p65,NLRP3,Caspase-1,ASC and IL-1 beta protein increased significantly,while the expression of these proteins decreased in different degrees in each treatment group.The results of QPCR and immunohistochemistry showed a similar trend.Conclusions1.Network pharmacological studies predict that the NF-kappa B/NLRP3/IL-1beta signaling pathway plays an important role in the treatment of hypertension by SCD.2.SCD can improve the symptoms of hyperactivity of liver-fire syndrome while reducing SHR blood pressure.3.SCD can improve left ventricular hypertrophy and protect target organs in SHR.4.SCD can down-regulate the expression of NF-kappa B/NLRP3/IL-1beta signaling pathway-related proteins,thus inhibiting inflammatory response,which may be the molecular mechanism of its antihypertensive and target organ protection. |
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