Effects Of Matrix Metalloproteinase-9/ Tissue Inhibitor Metalloproteinase-1 On Ventricular Remodeling In Normal Rat

BackgroundVentricular remodeling is one of pathologic mechanisms of chronic cardiac insufficiency,which is the outcome of many cardiovascular disease(such as hypertension,valvular heart diseases,and hypertrophic cardiomyopathy). Ventricular remodeling include myocardial remodeling and myocardial interstitium remodeling.The pathological change of extracellar matrix(ECM)is one of important components of myocardial interstitium remodeling,it would influence on proliferation and arrangement of myocardial cell and chang the compliance of ventricular wall,fanally,lead to heart failure.Matrix metalloproteinases(MMPs) are a family of zinc-dependent enzymes, each capable of degrading several ECM and non-ECM substrates. The MMPs is one of the major proteolysis system,it may play an important role in ECM remodeling by changing the content of ECM collage. Matrix metalloproteinases 9(MMP-9) is a member of the MMPs family and a key MMPs in myocardial interstitium. It was reported that MMP-9 and its endogenous inhibitor called tissue inhibitors of metalloproteinases 1(TIMP-1) may participate in the process of ventricular remodeling through regulating the contents of collagen and elastic fiber and acting on myocardial cell arrangement and morphology. Because TIMP-1 and MMP-9 are a couple of contradictive factors, we hypothesis their"net effect"may be the real key to ECM remodeling.But the previous researches showed their effects were inconsistent even in the same disease in different studies. It could not be excluded that these changes resulted from the influence of disease itself. Furthermore, it is not clear whether the changes of correlation between MMP-9, TIMP-1 with ventricular remodeling were causal relationship or accompanying relationship.To identify their real role,it is necessary to investigate the effect of MMP-9,TIMP-1 and MMP-9/TIMP-1 on ventricular remodeling without the effect of disease itself. We so designed the present experiments to study the effects of Matrix Metalloproteinase-9(MMP-9)/ Tissue Inhibitor Metalloproteinase-1(TIMP-1)on left ventricular structure ,function and cardiac pathology through injecting a seris of ratio solution of rat recombinant MMP-9 (rMMP-9)and TIMP-1(rTIMP-1) in normal Wistar rat (in vivo) and cultivating left ventricular tissue in same ratio culture solution of rMMP-9 and rTIMP-1 (in vitro).Objectiveto investigate the role of MMP-9, TIMP-1 and MMP-9/TIMP-1 in left ventricular remodeling so as to elucidate ventricular remodeling mechanism and search a new target for the treatment of ventricular remodeling furtherly.Methods1. In vivo experiment Thirty healthy male and thirty female Wistar rats weighing 120-200g in eight weeks period were matched according sex and weight and randomly divided into six groups:control group(n=l0) and interventional groups: TIMP-1 group(n=l0),MMP-9group(n=l0), 1:0.5group(n=l0),1:1group(n=l0), 1:2group(n=l0).The interventional groups were given rTIMP-1=30ng, rMMP-9=30ng, rMMP-9:rTIMP-1= 30ng,:15ng, rMMP-9:rTIMP-1=30ng:30ng, rMMP-9:rTIMP-1 =30ng:60ng respectively, and the control group were given the same dose of 0.9% sodium chloride solution through intraperitoneal injection, one time per day×14 days.The blood pressure were measured using noninvasive tail artery sphygmomanometer in the day before experiment ,7th day and 14th day after experiment.In 21th day after experiment,the changes of left ventricular structure[interventricular septal thickness at end diastole (IVSd), left ventricular internal dimension at end diastole (LVIDd), left ventricular posterior wall thickness at end diastole (LVPWd)]and function[left ventricular fractional shortening rate(LVFS),left ventricular ejection fraction(LVEF),stroke volume(SV),end-systolic left ventricular volume(LVESV) ,end-diastolic left ventricular volume(LVEDV),Cardiac Output (CO), ratio of mitral E peak velocity to A peak velocity (E/A)] were measured by echocardiography. And then the changes of the hearts histopathology were observed, collagen volume fraction(CVF) of left ventricular tissue and the index of left ventricular weight(LVWI= left ventricular weight(mg)/ body weight(g)) were measured.2. In vitro experiment Left ventricular explants from a healthy 8 weeks male (weight=146g)and a female (weight=125g)Wistar rat were cultivated 96 hours dividing into six groups respectively,ie control group and interventional groups( TIMP-1 group,MMP-9 group, 1:0.5 group,1:1 group, 1:2 group). The explants in interventional group given rTIMP-1=30ng only, rMMP-9=30ng only , rMMP-9:rTIMP-1=30ng:15ng suspl, rMMP-9:rTIMP-1=30ng:30ng suspl , rMMP-9:rTIMP-1=30ng:60ng suspl respectively and those in control group given the same dose of 0.9% sodium chloride solution were cultivated 96 hours. And then the pathobiologieal changes of the hearts were observed and CVF were measured.Results1. In vivo1) Blood pressure: Systolic blood pressure were significant higher in interventional groups than in controls(p<0.01),but no difference in diastolic blood pressure among groups.2) LVIDd: LVIDd of interventional groups were increased obviously than that of controls(p <0.01).In MMP-9/TIMP-1 combined groups LVIDd were increased accompany with the increasing doses of TIMP-1(p<0.01).3) LVWI: LVWI of interventional groups were increased obviously than that of controls(p <0.01).In MMP-9/TIMP-1 combined groups LVWI were increased accompany with the increasing doses of TIMP-1(p<0.01). 4) Histological examination:①Cardic myocyte proliferation, hypertrophy, ,disorder arrangement and interstitial hyperplasia predominantly occurred in interventional groups.②CVF of interventional groups were increased obviously than that of controls(p <0.01).In MMP-9/TIMP-1 combined groups CVF were increased accompany with the increasing doses of TIMP-1(p<0.01).5) The administration of exogenous MMP-9 can increase the level of serum TIMP-1, and vice versa.6) The ralationship between surem MMP-9,TIMP-1 and ventricular remodeling:①Serum MMP-9 was positively related to LVIDd, LVWI and CVF(r=0.911,p<0.001;r=0.729,p<0.001;r=0.87,p<0.001;respectively).②Serum TIMP-1 was positively related to LVIDd,LVWI and CVF(r=0.527,p<0.001;r=0.695,p<0.001;r=0.392,p<0.001;respectively).③Serum MMP-9/TIMP-1 was negatively related to LVWI(r=-0.287,p<0.001).2. In vitroCompare with control group, CVF was descreased in MMP-9 group and 1:0.5 group and increased in TIMP-1 group and 1:2 group. There was CVF increased from MMP-9 group,1:0.5 group,1:1 group to 1:2 group gradually, but no CVF difference between TIMP-1 group and 1:2 group and no CVF difference between control group and 1:1 group. Conclusions1. MMP-9 and TIMP-1 participated in the initial and developmental process of extracellular matrix remodeling. Their possible mechanisms are: 1) MMP-9 can low concentrations of collagen in ECM by degrading collagen and induce cardiac hypertrophy. 2) TIMP-1 can upregulate concentrations of collagen in ECM by inhibitting the activity of MMP-9. 3) The administration of exogenous MMP-9 can increase the level of serum TIMP-1, vice versa.2. Results in vitro showed the net effect of MMP-9 and TIMP-1 play a crucial role in ventrivular remodeling; when the ratio of MMP-9 and TIMP-1 was 1, the effect on ventricular remodeling is the lightest. This imply that adjusting the ratio of MMP-9 and TIMP-1 appropriately could extenuate ventricular remodeling.