Mechanism Of Regional Remodeling Of Myocardial Energy Metabolism In Patients With Heart Failure After Myocardial Infarction And The Intervention Effect Of Qili Qiangxin Capsul

As the key pathological link of heart failure(HF),cardiac metabolic remodeling takes a crucial role to the prognosis of the disease.However,the exact metabolic transformation of myocardial substrates after HF has yet to be determined.The previous results showed that the changes of glucose metabolism in the infarcted border area and the remote area were different,indicating that the heart might go through a metabolic remodeling in a regional manner after HF,which brought a new insight to explain the current controversies.Natriuretic peptide(NPs)increase sharply in the course of HF,regulating neurohormone and water-liquid balance.In recent years,the metabolic effects of NPs have been revealed,on the basis of a strong regulatory potential on glucose and fatty acid metabolism of peripheral tissues and organs.However,whether they participate in the regional metabolic remodeling of the heart after HF needs further exploration.As the promising treatments for HF,metabolic interventions have not been widely applied at present,which due to the controversies in the treatment strategies.Forcing the heart to rely on one substrate may lead to an extreme metabolic pattern,accompanied by the disuse of another substrate,resulting in the loss of metabolic flexibility finally.Hence,promoting the flexibility of cardiac metabolism and encouraging heart to choose the suitable metabolic phenotype flexibly according to the local environment is more conducive to myocardial energy production.Qiliqiangxin capsule(QLQX)has unique metabolic benefits,which can promote glucose and fatty acid metabolism simultaneously,so it is of great potential that QLQX could retain the availability of two substrates of failing myocardium and restore the flexibility.In order to explore the metabolic remodeling in different areas after HF and the regulatory role of NPs,and to clarify the interventional mechanism of QLQX in improving the metabolic flexibility of HF,the high performance liquid chromatography was performed to detect the energetic changes in the border area and the remote area in rats of HF after myocardial infarction(MI),and the PET/CT was conducted to observe the myocardial glucose and fatty acid metabolism in different areas.Furthermore,related enzymes of three metabolic phenotypes,glucose oxidation,glycolysis and fatty acid oxidation in distinct areas were evaluated.In addition,the regional changes of NPs and their receptors as well as the metabolic responses of cardiomyocytes to the NPs stimulation were also observed in vivo and vitro.On this basis,the effects of QLQX on the above indexes were then determined.The study of metabolic patterns in different areas after HF and the improvement on metabolic flexibility of Traditional Chinese medicine have important theoretical significance and clinical value for HF metabolic remodeling researches.Objectives:1.To reveal the regional properties of myocardial energy metabolism disorders and substrate remodeling and to elucidate the pathological mechanism of metabolic inflexibility in failing myocardium,via observing the changes of energetics and substrate metabolic enzymes in the border area and the remote area in rats of HF after MI;2.To clarify the efficacy advantages and potential mechanism of QLQX on improvements in the metabolic flexibility and energetic efficiency after HF,by observing its regulatory characteristics on myocardial metabolism in distinct areas.3.To preliminarily explore the mechanism of NPs and their receptors mediating the metabolic changes in distinct areas and the treatment on myocardial metabolic remodeling of QLQX,by evaluating the metabolic changes of cardiomyocytes stimulated with NPs in vitro and determining the regulation of QLQX on the NPs signals.Methods:1.The rat model of HF after MI was established by ligation of anterior descending branch of left coronary artery.The rats were randomly divided into sham,model,QLQX and enalapril group and the drug was given by gavage 24 h after operation for 8 weeks.H&E staining,echocardiography and treadmill exercise test were conducted to evaluate the cardiac pathological structures,systolic function and left ventricular remodeling and the exercise tolerance,respectively.The contents of adenosine triphosphate(ATP),adenosine diphosphate(ADP),adenosine monophosphate(AMP)and phosphocreatine(PCr)in the remote area and the border area were determined via the high performance liquid chromatography,and the total adenine nucleotide(TAN),energy charge(EC)and the ratio of PCr to ATP were calculated.2.18F-FDG PET/CT was used to observe the myocardial glucose uptake and metabolism level in the remote area and the border area of rats in each group.The protein expressions of glucose transporter 4(GLUT4),glucose transporter 1(GLUT1),phosphorylated pyruvate dehydrogenase(p-PDH),pyruvate dehydrogenase(PDH)and pyruvate dehydrogenase kinase 4(PDK4)in different areas were examined by Western blot and immunohistochemical staining,and the ratios of sarcolemmal to cytosolic expression of GLUT4 and GLUT1 in different areas were detected by immunofluorescence staining.phosphorylated3.The protein expressions of lactate dehydrogenase A(LDHA)in the remote area and the border area of rats in each group were examined by Western blot and immunohistochemical staining.The level of lactic acids(LA)in the infarcted border area myocardium was evaluated by lactate dehydrogenase method,and Na+/K+-ATPase and Ca2+-ATPase activities were detected by phosphorus determination method.4.18F-FTHA PET/CT was conducted to observe the myocardial fatty acid uptake and metabolism level in the remote area and the border area of rats in each group;Western blot and immunohistochemistry were used to detect the protein expressions of CD36 and CPT1A in different areas.5.The level of LA in plasma was measured by lactate dehydrogenase method and blood glucose(GLU),serum cholesterol(TC),triglyceride(TG),high density lipoprotein(HDL)and low density lipoprotein(LDL)by biochemical method.6.The cardiac expressions of B-type natriuretic peptide(BNP),A-type natriuretic peptide(ANP),natriuretic peptide receptor-A(NPR-A)and natriuretic peptide receptor-C(NPRC)protein in the remote area and the border area of rats in each group were detected by Western blot Establishing the model of rat embryonic cardiomyocytes H9c2 stimulated by BNP and ANP respectively,and the expressions of glucose and fatty acid metabolic enzymes were detected by Western blot.The ligand-receptor molecular docking technology was conducted to predict the affinity of the components of QLQX binding to NPR-A.Results:1.Compared with the sham group,it was confirmed that in the model group,there were slightly disordered myofibers in the remote area,and obvious cardiomyocytes necrosis,myofibrillar fragmentation and disordered arrangement in the border area,both of which could be alleviated in the QLQX group.The ejection fraction(EF)and fractional shortening(FS)were significantly lower,the left ventricular end-systolic internal diameter(LVIDs),left ventricular end-diastolic internal diameter(LVIDd),left ventricular end-systolic volume(LVESV)and left ventricular end-diastolic volume(LVEDV)were much higher and the left ventricular end-systolic anterior wall thickness(LVAWs),left ventricular enddiastolic anterior wall thickness(LVAWd),left ventricular end-systolic posterior wall thickness(LVPWs)and left ventricular end-diastolic posterior wall thickness(LVPWd)were remarkably thinner in the model group than in the sham group(P<0.05 or P<0.01);QLQX obviously elevated EF and FS,decreased LVIDs,LVIDd,LVESV and LVEDV,and thickened LVAWs,LVAWd,LVPWs and LVPWd(P<0.05 or P<0.01).Compared with the sham group,a significantly decrease of the time and distance of the exhaustive exercise(P<0.05)whereas no change of the maximum speed(P>0.05)were observed in the model group;there was a tendency to increase of the exercise time and distance in the QLQX group(P>0.05).2.Compared with the sham group,HF did not induce marked changes in ATP,AMP,TAN and EC but led to a notable decline of ADP only in the border area(P<0.05),a decrease of PCr in both areas(P<0.01 or P<0.001)and a reduction of the ratio of PCr/ATP only in the border area(P<0.05).It was found that the ADP in the border area was significantly lower than in the remote area(P<0.05).3.The fasted sham rats showed a minimal cardiac 18F-FDG uptake in the remote and the border areas.In comparison,a substantial increase of the SUVmean,SUVmax and SUVmin of 18F-FDG in the border area(P<0.001)and no obvious change in the remote area were found in the MI group;the 18F-FDG uptake in the border area was much higher than that in the remote area(P<0.001).In the remote area,the SUVmean,SUVmax and SUVmin of 18F-FDG were not changed by QLQX;however,in the border area,a rather higher 18FFDG uptake than that in model rats was observed in the QLQX group according to the SUVmean and SUVmax(P<0.05),while SUVmin was unchanged(P>0.05).4.Compared with the sham group,MI led to a significant attenuation of GLUT4 by 10.31%and 16.98%but an augmentation of GLUT1 by 137.53%and 162.29%and a enhancement of GLUT1/GLUT4 ratio by 162.13%and 221.95%in the remote area and the border area respectively;the ratio of sarcolemmal to cytosolic GLUT4 in the remote and the border area were dampened by 15.68%and 25.00%,whereas that of GLUT1 was augmented by 14.34%and 19.85%respectively(P<0.05 or P<0.01).Following the treatment with QLQX,the protein content and the sarcolemmal/cytosolic ratio of GLUT4 remained unchanged in the remote area but increased in the border area(P<0.05 or P<0.01),concomitant with a great suppression of that of GLUT1 in both areas(P<0.01 or P<0.001).5.Compared with the sham group,the ratio of p-PDH to t-PDH in the remote area and the border area were decreased by 48.07%and 64.01%as well as the contents of PDK4 were downregulated by 18.23%and 48.55%respectively in the MI group(P<0.05 or P<0.01);however,only the reduction in the border area(P<0.05 or P<0.01)rather than in the remote area was prevented by QLQX.6.Compared with the sham group,the infarcted hearts were characterized by a marked decline in LDHA protein expression by 38.11%and 58.22%in the remote area and the border area,respectively(P<0.05 or P<0.001);LDHA content was induced in the border area(P<0.05)but remained unchanged in the remote area after QLQX treatment.In addition,the content of LA in the border area decreased in the MI group,but QLQX had no effect on it.No difference of Na+/K+-ATPase and Ca2+-ATPase in the border area myocardium was found among the sham,model and QLQX group.7.Compared with the sham group,a remarkable decrease of the SUVmean,SUVmax and SUVmin of 18F-FTHA in both areas were observed after MI;the border area was higher than that in the remote area(P<0.01 or P<0.001).In the remote area,the average value of 18FFTHA SUVmean,SUVmax and SUVmin was augmented by QLQX without a statistical significance(P>0.05);however,no difference was observed in the border area.The infarcted heart showed a great reduction in the protein expression of CD36 by 50.42%and 26.06%and CPT-1A by 23.83%、30.88%in the remote area and the border area,respectively(P<0.05 or P<0.01).QLQX significantly increased the expressions of CD36 and CPT-1A in the remote area(P<0.05 or P<0.01),but no change was detectable between QLQX group and MI group in the border area.8.Compared with the sham group,the levels of LA in plasma and GLU,FFA,TC,HDL and LDL in serum of the model group were unchanged,whereas serum TG content was significantly reduced(P<0.05).9.Compared with the sham group,MI led to no change in the remote area but a notable suppression in the border area of BNP protein expression(P<0.01).The ANP contents were induced enormously by 135.51%and 486.79%in the remote area and the border area respectively(P<0.05 or P<0.001).QLQX treatment upregulated the BNP expression in the border area(P<0.05)but not the remote area.In contrast,QLQX downregulated the ANP contents in the remote area(P<0.01)rather than the border area.Compared with the control group,the protein expressions of GLUT1,GLUT4,LDHA and CPT-1A of H9c2 in BNP group did not reach an obvious change,but CD36 was significantly increased(P<0.05).The expressions of GLUT1 and CD36 were significantly upregulated and GLUT4 and LDHA were significantly downregulated in the ANP group(P<0.05),but there was no difference of CPT-1A between ANP and control group.10.The NPR-A was lower by 51.07%in the remote area and much higher by 135.79%in the border area in the model group than that in the sham rats;the contents of NPR-C were enlarged by 61.01%and 324.81%as well as the ratio of NPR-A/NPR-C was suppressed by 56.29%and 46.78%in the remote area and the border area respectively(P<0.05 or P<0.01).Contrast to the MI rats,QLQX group stimulated the NPR-A only in the remote area(P<0.05)and decreased the NPR-C protein expression in both areas(P<0.05).In addition,the binding energy affinity of astragaloside IV,a component in QLQX,was lower than ANP,and the binding affinities of a variety of components in QLQX were similar to ANP while lower than BNP.Conclusions:1.The energy metabolic remodeling in the remote area and the border area was characterized by regional diversity in HF rats after MI.The energy reserve capacity was significantly reduced in the remote area,while both the energy reserve and instant energy fell simultaneously in the border area.There also was a heterogeneous substrate remodeling in distinct areas,the total glucose metabolism remained unchanged in the remote area after HF,although glulcose oxidation was elevated and glycolysis was weakened,moderately;however,the glucose metabolism strongly enhanced in border areas,with a sharply increased glucose oxidation and a deficiency in the glycolysis.The cardiac glucose transporters subjected to a transformation from adult subtype GLUT4 to fetal subtype GLUT1.The fatty acid metabolism in both remote and border areas dropped,and the disuse of fatty acid especially the its transportation dysfunction were more obvious in the remote area.In brief,a stronger difficiency of fatty acid metabolism in the remote area and a severer abnormality of glucose metabolism in border aera occurred in the infarcted heart.2.QLQX could correct the disordered fatty acid metabolism in the remote area and glucose utilization in the border area respectively.QLQX further enhancing the total glucose metabolism in the border area via promoting GLUT4 expression and fixing the GLUT4/GLUT1 imbalance;it reduced the abnormally augmented glucose oxidation and improved anaerobic glycolysis in the border area.In addition,QLQX enlarged the fatty acid metabolism in the remote area rather than the border area.The metabolic modulation could optimize the overall cardiac energy efficiency and restore the myocardial metabolic flexibility.3.BNP and ANP could regulate fatty acid and glucose metabolism in cardiomyocytes,and NPs and their receptors expressed regionally in the HF myocardium,indicating that NPs signals might modulate the pathology of cardiac metabolic inflexibility.QLQX exhibited a heterogenous manipulation on NPs and the receptors in different areas,meanwhile,the binding affinity of various effective components especially yiqi components in QLQX with NPR-A was similar to or even more than that of ANP and BNP,suggesting that QLQX may play the role of metabolic regulation by correcting the NPR-A/NPR-C unbalance or binding with NPR-A directly.