| Purposes:The latest epidemiological studies have shown that the incidence of acute myocardial infarction(AMI)still maintains an upward trend and is getting younger in China.Ventricular remodeling(VR)is a compensatory and secondary complex pathophysiological process that occurs after AMI and involves multiple influencing factors.Long-term and continuous VR could change the structure and function of the heart,eventually induce heart failure and death.It is one of the determinants of cardiovascular mortality.Therefore,inhibiting or reversing the process of VR has become a key in the prevention and treatment of cardiovascular disease.Clinical studies have shown that the basic TCM syndrome type of patients with VR after AMI is Qi deficiency and blood stasis syndrome or combined other syndrome types as well.Yiqihuoxue TCM and their effective components as the basic prescription have achieved obvious clinical effects in the treatment of VR,especially the effective component of TCM.Studies at home and abroad have shown that Panax notoginseng saponins(PNS)have pharmacological effects such as dilating blood vessels,improving microcirculation,and reducing myocardial oxygen consumption;tanshinone IIA(Tan IIA)could improve cardiac nerve remodeling and electrophysiological abnormalities by inhibiting myocardial fibrosis cardiomyocyte apoptosis;total ginsenosides(TGS)could enhance myocardial contractility,accelerate lipid metabolism,and significantly reduce the formation of scars in myocardial infarction area.In order to improve the cardiac function and hemodynamic indexes after AMI,the three are the biological active ingredients of the drugs that promote blood circulation and benefit qi,and they could protect the cardiovascular system of myocardial infarction rats when used alone,however,the combination of them to interfere with VR after AMI has not been studiedThe ATF3/MAP2K3/p38 MAPK signaling pathway can affect the inflammatory infiltration of myocardial ischemia by affecting the expression of NFκB and other factors,as well as inhibiting the expression such as TGF-β1 and Collagen I,reducing Runx1 and improving PPARa expression to prevent VR after MI.Based on the tutor’s many years of clinical medication experience and the research foundation of our research group,this study intends to explore the combination of traditional Chinese medicine PNS,TanⅡA and TGS in the Yiqihuoxue group by adjusting ATF3,MAP2K3,p38 MAPK,p-p38 MAPK,TGF-β1,Collagen I,NFκB p65,p-NFκB p65,Runx1,PPARa expressions,thereby activating the ATF3/MAP2K3/p38 MAPK signaling pathway in preventing and treating VR after AMI.To provide a powerful experimental basis for the development of Yiqihuoxue component traditional Chinese medicine to be a new clinical drug for prevention and treatment of VR after AMI.Methods:The left anterior descending(LAD)branch of the heart of healthy SPF grade SD male rats were ligated to construct MI animal models.The rats modeled successfully were randomly divided into six groups:model group(MI),fosinopril group(Fosinopril,FIP,1.2 mg/kg d),panax notoginseng saponins group(PNS,40 mg/kg·d),Tanshinone ⅡA group(Tan Ⅱ A,70mg/kg·d),total ginsenosides(TGS,30 mg/kg-d),component compatibility group(Component compatibility,CC,PNS 40mg/kg·d+Tan ⅡA70mg/kg·d+TGS 30mg/kg·d);rats in the sham-operated group(Sham)were threaded without ligating at the same position of the LAD coronary artery as positive control.The corresponding drugs were given by gavage in each administration group,and rats in the MI group and Sham group were given equal volumes of deionized water for 4 weeks.After 4 weeks,the heart of each group of rats were detected by ultrasound,and the heart weight/body weight(HW/BW)of each group were calculated ELISA method was used to detected the levels of CRP,TNF-α,GDF-15 and MMP9/TIMP1 in rat serum;ATP,ADP,AMP levels in myocardial tissue were detected by high-performance liquid chromatography;Na+-K+-ATPase and Ca2+-Mg2+-ATP enzyme content in myocardial tissue were detected by chemical colorimetry;western blotting,immunohistochemistry and RT-PCR were used to detected the expression of ATF3,MAP2K3,p38 MAPK,p-p38 MAPK,TGF-β1,Collagen Ⅰ,NFκB p65,p-NFκB p65,Runx1,PPARα.Results:1 The effect of huoxueyiqi CC-TCM on cardiac structure and function in MI rats1.1 Echocardiographic testing of cardiac function after 4 weeks of administration and the results showed that the LVIDs,LVIDd was significantly increased,and LVEF and LVFS were significantly reduced in MI group;after 4 weeks of administration,compared with the MI group,the LVIDs and LVIDd were significantly reduced,and LVEF and LVFS were significantly increased in each administration group;the improvement of each index of the CC-TCM was significantly better than that of the single-component Chinese medicine group.1.2 HE staining results showed that after ligation of the LAD coronary artery,compared with the sham-operated group,the model group had disordered myocardial fiber arrangement,widened gap,and uneven staining;myocardial tissues were loose,and the volume of myocardial cells increased;there were inflammatory cells infiltration and hyperplasia of connective tissue,accompanied by the dissolution,rupture,and necrosis of myocardial fibers,indicating that the animal models of VR animals after MI were established successfully.After 4 weeks of administration,compared with the model group,the pathological structure of the myocardium of the rats in each administration group were improved in varying degrees;inflammatory infiltration was slightly reduced,and cell morphology and structure were improved,and the CC group was more obvious than the single component Chinese medicine group1.3 Masson staining results showed that after ligating the LAD coronary artery of the rat for 4 weeks,compared with the sham-operated group,the model group’s heart collagen fibers and connective tissue were significantly proliferated accompanied by increase in myocardial collagen tissue and fibrous scars;after 4 weeks of administration,compared with the model group,the myocardial fibers and extracellular matrix remodeling of rats in each administration group were improved in varying degrees,and collagen deposition decreased.Among them,the CC group improved more obviously than the single component Chinese medicine group.1.4 The results of HW/B W showed that the model group had a significantly higher cardio-body index than the sham-operated group of rats;after 4 weeks of administration,compared with the MI group,the HW/BW of each administration group decreased;compared with the traditional Chinese medicine group of each component,the decrease of the HW/BW in CC group rats were more obvious1.5 ELISA method was used to detected VR markers after 4 weeks of administration.Compared with the sham-operated group,the levels of CRP,TNF-α,GDF-15 and MMP9/TIMP1 in MI group were elevated significantly;after 4 weeks of administration,compared with the MI group,the levels of CRP,TNF-α,GDF-15 and MMP9/TIMP1 in the serum of rats in each administration group were reduced;the level of markers were decreased in CC group rats significantly.2 The effect of huoxueyiqi CC-TCM on the ATF3/MAP2K3/p38 MAPK signaling pathway2.1 The LAD coronary artery of the rats were ligated after four weeks,and the results of immunohistochemistry,western blotting and RT-PCR showed that the expression levels of TGF-β1 and collagen Ⅰ were significantly increased;after 4 weeks of treatment with CC-TCM,compared with the MI group,the huoxueyiqi CC-TCM could reduce collagen Ⅰ and TGF-β1 expression to improve myocardial fibrosis and extracellular matrix remodeling after MI.2.2 The LAD coronary artery of the rats were ligated after four weeks,and the results of immunohistochemistry,western blotting and RT-PCR showed that ATF3 protein in the myocardial tissue of the MI group was reduced,the expression of MAP2K3 and p38 MAPK increased significantly compared with the sham-operated group;after 4 weeks of treatment,the results showed that huoxueyiqi CC-TCM could increase ATF3 protein expression in MI rats and reduce the expression levels of MAP2K3 and p38 MAPK.2.3 The LAD coronary artery of the rats were ligated after four weeks,the results of immunohistochemistry,western blotting and RT-PCR showed that the expression of NFκB and p-IκBα were significantly increased compared with the sham-operated group;after 4 weeks of treatment with huoxueyiqi CC-TCM,the results showed that huoxueyiqi CC-TCM could reduce the expression of NFκB and p-IκBα in MI rats compared with the MI rats.3 The effect of huoxueyiqi CC-TCM on the energy metabolism of the MI rat3.1 The high-performance liquid chromatography detection of ATP,ADP,AMP levels in myocardial tissues after administration four weeks showed that compared with rats in the sham-operated group,the ATP,ADP levels in the model group myocardial tissue were significantly reduced,AMP levels were significantly increased;after 4 weeks of treatment with CC-TCM,the results showed that the huoxueyiqi CC-TCM could increase ATP and ADP in the myocardial tissue of MI rats compared with the model group.3.2 The chemical colorimetric method detected the contents of Na+-K+-ATPase and Ca2+-Mg2+-ATPase in the myocardial tissue of rats four weeks after the administration showed that the content of Na+-K+-ATPase and Ca2+-Mg2+-ATPase decreased significantly in MI group compared with the rats in the sham operation group;after 4 weeks of treatment with huoxueyiqi CC-TCM,the results showed that huoxueyiqi CC-TCM could increase the levels of Na+-K+-ATPase and Ca2+-Mg1+-ATPase in MI rats.3.3 The ELISA method detected the level of free fatty acid in myocardial tissues of rats after four weeks administration.Compared with the sham-operated group,the free fatty acid levels in the myocardial tissues of the model group were significantly reduced;the huoxueyiqi CC-TCM could increase the level of free fatty acid in MI rats.3.4 The LAD coronary artery of the rat model was ligated after four weeks,the results of immunohistochemistry,Western blotting and RT-PCR showed that the expression of Runx1 were increased and PPARa were significantly decreased;after 4 weeks of treatment,huoxueyiqi CC-TCM could reduce the expression of Runx1 and improve the expression of PPARα to improve the energy metabolism of the MI rats.Conclusion:1 The CC-TCM of huoxueyiqi could improve the heart function and structure after MI,and the combination of CC-TCM of Huoxueyiqi shows obvious synergistic effect.2 The CC-TCM of huoxueyiqi could reduce the level of myocardial tissue fibrosis and myocardial extracellular matrix remodeling after MI by inhibiting the expression levels of Collagen Ⅰ and TGF-β13 The CC-TCM of huoxueyiqi could activate the ATF3/MAP2K3/p38 MAPK signaling pathway in MI rats,and then blocks the NFκB/IκB signaling pathway to reduce inflammation and inhibit myocardial fibrosis and extracellular excess precipitation of the matrix,and the CC-TCM of huoxueyiqi could reduce the expression of Runx1 and improve the expression of PPARα,the downstream of ATF3/MAP2K3/p38 MAPK in VR of key factors of energy metabolism,which may be one of the important molecular mechanisms for preventing and treating VR. |
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