| Objective: Study on the substantial base for calming the liver and suppressing yang effect of uncaria rhynchophylla,identify its core components and the surrounding components to achieve an accurate description of the mechanism of the core components and the ambiguous description of the mechanism of the surrounding components in vivo and construct the pharmacological map of “Composition-Target-Pathway”.Methods: 1.Hypertension with liver-yang hyperactivity syndrome rat model was produced by spontaneously hypertensive rats with fuzi decoction administration by gavage and intervened with different components of uncaria rhynchophylla.The blood pressure,irritability,pain threshold,rotation time,24 h drinking water as well as the changes of facial temperature were observed throughout the intervention program.After intervention,the serum biochemical indexes and metabolomics indexes of rats in each group were detected and analyzed.According to the results of previous studies and the data of this study,we used partial least-squares regression analysis(PLS)with SIMCA-P11.5 software to identify the core components and surrounding components of uncaria rhynchophylla on its calming the liver and suppressing yang effect according to the correlation coefficient and the variable importance parameters of regression analysis.2.The SHRs with liver-yang hyperactivity syndrome were interved by the core components and surrounding components of uncaria rhynchophylla on its calming the liver and suppressing yang effect and the blood pressure and “syndromes and signs” were observed throughout the intervention program.For the NC group,the DS group and the core composition group: Enzyme-linked immune sorbent assay(ELISA)was applied to detected the rat’ serum biochemical indexes.HE staining was used to observe the pathological changes of the heart,brain tissue and kidney.Scanning electron microscope(SEM)was used to observe the endometrial morphology of renal artery and mesenteric artery.The protein expression of AKT and TGF-β1 were measured by western blot and the gene expression for kallikrein,ACE2 and mas-1 were detected by RT-PCR.For the NC group,the DS group and the surrounding components group: The serum metabolites were detected using metabolomics technique based on UPLC-Q-Exactive/MS to explore the metabolic pattern and potential biomarkers on system biology.After the above experiments,the accurate molecular biology mechanism of the core components and the ambiguous metabolomics mechanism of the peripheral components of uncaria rhynchophylla on its calming liver and suppressing yang effect in vivo were clarified,respectively.3.With the method of network pharmacology,the mechanisms of the core components and the surrounding components of uncaria rhynchophylla on its its calming liver and suppressing yang effect were comprehensively investigated,and the pharmacological map of “Composition-Target-Pathway” was constructed.Results: 1.The component of uncaria rhynchophylla with ursolic acid(UA)or oleanolic acid(OA)knocked out can effectively reduce the blood pressure in hypertensive rats with liver-yang hyperactivity,reduce the rats’ facial temperature and water intake,improve the rats’ pain threshold,extend the rotation time,and reverse the changes of biochemical indexes and the abnormal expression of 11 biomarkers in the serum of hypertension rats with liver-yang hyperactivity.Combined the previous study with the present work to analyze the integrated efficacy,we found that the UA knockout group,the ICO knockout group and the valsartan group present a better treatment effect than the COR knockout group and the OA knockout group,the Rhy knockout group and the IR knockout group showed a poor effect with the IR knockout group to be the worst.2.IR can effectively reduce the blood pressure in hypertensive rats with hyperactivity of liver-yang,improve the quantitative indicators of “symptoms and signs” in rats,reverse the abnormal changes of the biochemical indicators in rat serum and improve the pathological changes of rat heart,brain tissue and kidney and the endometrial morphology of renal artery and mesenteric artery under SEM.Furthermore it can also regulate the aberrant expression of AKT,TGF-β1 proteins and KK,ACE2 and mas-1 m RNA in the rat thoracic aorta.The peripheral compositions can also effectively reduce the blood pressure of hypertensive rats with liver-yang hyperactivity,improve the quantitative indicators of “symptoms and signs” in rats as well as regulate the aberrant expression of thirteen differential metabolites involved in lipid metabolism and energy metabolism.3.Network analysis found that the activeconstituents of uncaria rhynchophylla on its tranquilizing liver and yang effect could regulate 50 target genes including AKT2,CASP,IL4 and IL10,interfering with eight major pathways such as MAPK signaling pathway,T cell receptor signaling pathway,Ras signaling pathway,and neurotrophin signaling pathway and regulating the expression of cytokines,neurotransmitters,and hormones,thus to exert its function.Conclusions: 1.IR was the core composition of uncaria rhynchophylla on its tranquilizing liver and yang effect.2.The mechanism of IR on its treatment on hypertensive rats with hyperactivity of liver-yang was related to the regulation of RAAS and sympathetic nervous system activity by regulating the levels of vascular endothelium contraction and contraction factors including Ang II,ET-1 and NO,as well as by promoting the expression of related signal molecules in ACE-Ang1-7-mas-AKT signaling pathway,enhancing the expression of kallikrein gene and regulating the release of vascular endothelium contraction factors.IR can effectively improve the pathological changes of heart,brain tissue and renal tissue,ameliorate the endometrial morphology of renal artery and mesenteric artery in hypertensive rats with liver-yang hyperactivity,and inhibit the expression of TGFβ-1 protein,thereby improving rat vascular remodeling.The metabolomics mechanism of the peripheral compositions of uncaria rhynchophylla was that it could regulate the abnormal expression of 13 related differential metabolites in lipid metabolism and energy metabolism pathways.3.The activeconstituents of uncaria rhynchophylla on its its tranquilizing liver and yang effect can interfere with eight major pathways such as MAPK signaling pathway,T cell receptor signaling pathway,Ras signaling pathway,and neurotrophin signaling pathway and regulating the expression of cytokines,neurotransmitters,and hormones,thus to exert its function. |
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