Study Of PPARγ Partial Agontists Telmisartan Inhibiting Myocardial Interstitial Remodeling In Spontaneous Hypertensive Rats

BackgroundHypertension is one of the most common cardiovascular diseases at present,and also the risk factor for coronary artery disease,cerebral apoplexy,heart function failure and kidney function failure.The cause of death of most patients with high blood pressure is the damage to target organs induced by hypertension.So it is important equally to prevent and treat the damage to target organs.Myocardial remodeling is one of the most common changes in primary hypertension.The two aspects of pathological changes are brought forward,one is myocardial cell's hypertrophy and the other is myocardial interstitial remodeling.Myocardial remodeling reduces the function of both diastole and contraction,impacts the myocardial metabolism and function,and then impairs the ventricular function,and induces the heart failure at last.As an independent cardiovascular risk factor,it is closely linked to the prognosis of hypertension.Therefore,how to prevent and reverse the hypertensive myocardial remodeling is becoming a key study.The major cause of hypertensive myocardial remodeling is the unbalance between the synthesis and degradation of the myocardial interstitial collagen,and the change in proportion of different collagens(mainly the proportion of CollagenⅠto CollagenⅢ).The synthesis and degradation of collagen are controlled by the gene and other factors.The factors stimulating the synthesis of collagen effect the fibroblast,and then induce expression of the gene AP-1 and NF-κB,after that bring about a series of events of transcriptional control,result in changes of the gene expression and the cellular function at last.Matrix metalloproteinase(MMPs)is a group of main enzyme system,which could degrade the composition of extracellular matrix.The tissue inhibitor of metalloproteinase(TIMPs)combines with MMPs, and forms a stable and irreversible complex,MMP-TIMP complex,to block the combination of MMPs with its substrate,and thus inhibit its hydrolysis effect. Peroxisome proliferator-activated receptors(PPARs)are a family of at least 3 nuclear receptors(α,β/δ,γ).PPARγexpresses in cardiac highly,involved in fat production and glucose metabolism.Telmisartan,as a high-selective angiotensinⅡandⅠreceptor antagonist,could block the combination of AngⅡwith AT1,to reverse the myocardial interstitial remodeling.However,telmisartan as the PPARγpartial agonist,the research on the mechanism of its effect on the myocardial interstitial remodeling is less.In recent years,it is reported that PPARγinhibited AP-1 and NF-κB activity,to improve the left ventricular hypertrophy caused by pressure overload.On the basis of advance in myocardial interstitial remodeling and the cellular signal transduction pathway,we hypothesize that temisartan regulates the expression of PPARγmRNA up,and enhances its activity of transforming;then inhibits the signal transduction pathway of AP-1 and NF-κB via stimulating PPARγ, then control expression of MMP-9 and TIMP-1,thus reduce the myocardial interstitial remodeling in SHR.Therefore,we observe the intervention by telmisartan in SHR myocardial remodeling,try to clarify the possible mechanism and find the new target point of therapy,and to provide the foundation of theory and practice for reversing remodeling and heart failure by conversing of myocardial interstitial remodeling and further for getting new areas of application of drug to intervention myocardial interstitial remodeling of hypertension.Objective(1)To observe the effect of telmisartan on myocardial remodeling in spontaneous hypertensive rat(SHR). (2)To probe into the mechanism of telmisartan on myocardial interstitial remodeling of hypertension.Methods16 male 8-week-old SHR were randomly divided into two groups: SHR-controlled group(SHR group,n=8)and Telmisartan group(n=8).The same week-old Wistar-kyoto(WKY)rats was also randomly selected as normal contuol (WKY group,n=8).The animals in Telmisartan group were dosed orally with Telmisartan(60mg.kg^-1.d^-1)and the other group animals were given with same volume of saline every day for 8 weeks.The detection contents were listed below. (1)All animals were weighed every week and the blood pressure through arteria caudilis was measured every two weeks.(2)The common echocardiographic indexes were detected before and at the end of the experiment.(3)After the rats were killed,we get left ventricle mass(LVM)and calculate the left ventricular relative mass(LVM/BM).(4)Ultrastructral and histopathological changes of myocardial tissue morphology myocardial with HE staining were observed.(5)The ultrastructure of myocardial cell was observed with transmission electron microscope. (6)The content of collagen was quantified by Masson staining.(7)The protein levels of C-Fos,C-Jun were detected by immunohistochemistry.(8)The mRNA expression of CollagenⅠ,CollagenⅢ,MMP-9,TIMP-1,PPARγ,C-Jun and C-Fos were detected by quantification real-time RT-PCR,the proportion of MMP-9 to TIMP-1 was calculated.(9)C-Jun/C-Fos heterodimers and ezpression level of C-Jun preotein were detected by Co-immunoprecipitation-Western-blot.Result1.Blood pressureDuring the whole experiment,compared with the WKY group,the blood pressure in SHR group were significantly increased(P＜0.01);compared with SHR group,the blood pressure in telmisartan group were significantly improved(P＜0.01).2.left ventricular relative mass(LVM/BM)At the end of the experiment,Compared with SHR group,the LVM/BM telmisartan group were significantly decreased(P＜0.01). 3.Echocardiographic detectionThe common echocardiographic indicators:Before the experiment,compared with WKY group,the LVPWTd and IVSTd in SHR group and telmisartan group were significantly increased(P＜0.01);at the end of the experiment,compared with SHR group,the LVPWTd and IVSTd in telmisartan group were significantly improved (P＜0.01).Before the experiment,the E peak,A peak,E/A ratio,E/DT and IVRT were insignificantly different from the WKY group,the Em/Am in SHR group and telmisartan group were lower than that in WKY group.At the end of the experiment, compared with SHR group,the E/A ratio in telmisartan group was obviously increased(P＜0.01),the Em/Am was also increased(P＜0.05)and IVRT was shorten (P＜0.01).Integrated backscatter detection:Before the experiment,there was no significant difference of the integrated backscatter percent(IBS%)and cyclic variation of IBS (CVIB)in left ventricular posterior wall(LVPW)and interventricular septa(IVS) among the three groups(P＞0.05).At the end of the experiment,Compared with SHR group,IBS%of the telmisartan group was obviously decreased(P＜0.05)and CVIB of telmisartan group was obviously increased(P＜0.05)in IVS,and IBS%of the telmisartan group was obviously decreased(P＜0.01)and CVIB of telmisartan group was obviously increased(P＜0.05)in LVPW.4.The changes of myocardial tissue morphology with HE stainingIn WKY group,heart cells neat and uniform size nuclear,cytoplasmic staining uniform.In SHR group myocytes disarranged,the size of irregular nuclear, cytopLasmic staining better,visibility myocardial cells within the fiber breaKage.In Telmisartan group,myocytes with more rows,the nuclear occasionally irregular size, and cells myocardial fiber marked improvement.5.Ultrastructural changes of myocytes observed by transmission electron microscopy.The left ventricular myocytes from the WKY group arranged regularly.The pericellular membrane was uninterrupted and intact.The thick and thin myofilament arranged regularly.The sarcomere and light dark band were clear.The sarcomere and light dark band were clear.The uniformly sized mitochondrial was abundant and showed round or oval shape.A little fibrobLast and collagenous fibers distributed in extracellular matrix.The left ventricular myocytes from SHR group arranged irregularly.The pericellular membrane was interrupted and unclear.The local myofibril was disintegrated.The myofilament was distorted and interrupted. The sarcomere was in a bad position.The swelling mitochondrial increased and accumulated.A lot of collagenous fibers distributed in extracellular matrix. Compared with SHR group,the ultrastructural change of Telmisartan group was obviously improved.The myocytes from Telmisartan group arranged more regularly than SHR group. The phenomenon that the local myofibril was disintegrated was decreased;the mitochondrial was more regular than SHR group.Then unclear shape was more regular than SHR group.The collagenous fibers in extracellular matrix decreased obviously.6.The content of collagen was quantified by Masson-staining.The myocyte was red or yellow and the collagen was green or blue.The collagen tissue was appropriate arranged among cardiomyocytes in WKY group, however,collagen tissue in creased markedly,and disrupted in sarcomere in SHR group.The collagen tissue decreased and arranged regularly in Telmisartan group.7.Immunohistochemistry detection(1)C-Fos,C-Jun:The positive reactions of C-Fos,C-Jun protein were stained brown and absentin myocardial cell nucleus.There was little distribution of brown granule in WKY group.There was thick brown granule in SHR group(P＜0.01).The granule of Telmisartan group was weaker than SHR group(P＜0.01),but still higher than WKY group(P＜0.01).(2)NF-κB:The positive reaction of NF-κB protein was stained brown and absentin myocardial Cytoplasm.There was little distribution of brown granule in WKY group.There was thick brown granule in SHR group(P＜0.01).The granule of Telmisartan group was weaker than SHR group(P＜0.01).8.The mRNA expression of C-Fos,C-Jun were detected by quantification real-time RT-PCRThe mRNA expression of C-Fos,C-Jun,CollagenⅠ,CollagenⅢ,NF-κB in SHR group were higher than that of was in WKY group(P＜0.01),its in Telmisartan group were lower than SHR group(P＜0.01).The mRNA expression of PPARγin SHR group was lower than that of was in WKY group(P＜0.01),its in Telmisartan group was higher than SHR group(P＜0.01).The mRNA expression of MMP-9 in SHR group was lower than that of was in WKY group(P＜0.01),its in Telmisartan group was higher than SHR group(P＜0.01). The mRNA expression of TIMP-1 in SHR group was higher than that of was in WKY group(P＜0.01),its in Telmisartan group was lower than SHR group(P＜0.01). MMP-9/TIMP-1 in SHR group was lower than that of was in WKY group(P＜0.01), its in Telmisartan group was higher than SHR group(P＜0.01).9.The protein levels of C-Fos/C-Jun,C-Jun were detected by Western blot.(1)C-Fos/C-Jun:The expression of C-Fos/C-Jun protein in SHR group was higher than that of was in WKY group(P＜0.01),its in Telmisartan group was lower than SHR group(P＜0.01).(2)C-Jun:The expression of C-Jun protein in SHR group was higher than that of was in WKY group(P＜0.01),its in Telmisartan group was lower than SHR group (P＜0.01).Conclusion1.Sixteen week-old SHR have obvious myocardial myocardial remodeling.2.Telmisartan has a protective effect on myocardial remodeling in the SHR.3.Telmisartan partly actives PPARγreceptor,inhibits the AP-1 and NF-κB signal transduction pathway and the formation of C-Fos/C-Jun heterodimers resulting inhibiting the transcription of its downstream gene,which may be the mechanism of above 2.