RhoA/ROCK Signaling Pathway Joins In The Cardiac Hypertrophy Induced By Iso In Rats And The Intervention Of Fasudil

Objective:To investigate the role of RhoA/ROCK signaling pathway in cardiac hypertrophy(CH)induced by isoproterenol(Iso)in rats and the intervention effects of fasudil(Fas).Methods: CH models of rats were established by subcutaneous injection of Iso.48 Sprague-Dawley rats weighing 180 g ~ 200 g,were randomly divided into four groups: Normal control group,CH model group,Fas with low-dose group(L-Fas,5mg / kg / d),Fas with high-dose group(H-Fas,20 mg / kg / d),12 in each group.Except for the normal control group,the rest three groups rats were injected subcutaneously with Iso for setting up CH model,L-Fas group was given Iso(s.c,5mg / kg / d)+ Fas(i.p,5mg / kg / d),H-Fas group was given Iso(s.c,5mg / kg / d)+ Fas(i.p,20 mg / kg / d),normal control group and Iso model were given the same volume of normal saline.Fasadministered for 8 weeks.After the treatment,the following index were detected:(1)Ejection fraction(EF),fractional shortening(FS),left ventricular wall thickness(LVPW)and septal thickness(IVS)of M-mode echocardiography was measures in six rats randomly selected in each group;(2)Six rats in each group were randomly selected,after weighed,systolic pressure(LVSP),left ventricular late diastolic(LVEDP),left ventricular systolic maximum rate(+ dp/dtmax)and left ventricular diastolic maximum rate(-dp/dtmax)and heart rate(HR)of hemodynamic were detected;(3)After hemodynamic,blood taken from common carotid artery and nitric oxide synthase(eNO),glutathione peroxidase(GSH-PX)activity,the level of nitric oxide(NO)were measured in serum,after the heart was removed,we detected myocardial tissue superoxide dismutase(SOD)activity and malondialdehyde(MDA)level;(4)After the blood was completed,we quickly removed the heart,cleared the blood and water,weighed heart weight(HW)and left ventricular weight(LVW),to calculate heart weight index(HWI)and left ventricular mass index(LVWI);(5)Myocardial tissue specimens,select 6 hearts in each group embedded in paraffin,HE and Masson staining were performed for observing the histopathological changes,while using immunohistochemical methods for observing the protein expression of RhoA,ROCK1,ERK1 / 2 and c-FLIP in myocardial tissue.(6)Another cardiac tissue(the left 6hearts)to use semi-quantitative RT-PCR method for detecting the expression of RhoA,ROCK1,ERK1 / 2 and c-FLIP mRNA in the myocardial tissue of rats.Results:(1)Echocardiography.Compared with normal control group,the LVPW(P <0.05)and IVS(P <0.01)increased significantly,while the EF and FS was significantly decreased(P <0.01)in CH group.Compared with the CH group,IVS decreased significantly(P <0.05),while EF was significantly higher(P <0.05),LVPW and FS was no significant difference in L-Fas group;LVPW(P <0.05),IVS(P <0.01)decreased significantly,EF(P <0.01),FS(P <0.05)were significantly increased in H-Fas group.(2)Hemodynamic changes.Compared with normal control group,HR(P <0.05)and LVEDP(P <0.01)increased significantly,while LVSP and ± dp / dtmax decreased significantly(P <0.01)in CH group.Comparison with the CH group,LVSP group was significantly higher(P<0.05)in L-Fas;HR and LVEDP decreased significantly(P <0.05),while LVSP and ± dp / dtmax increased significantly(P <0.05)in H-Fas group.(3)Biochemical parameters of blood and myocardial tissue.Compared to normal control group,NO content and NOS,SOD,GSH-PX activity(P <0.01)significantly lower in CH group,but MDA content was significantly increased(P <0.01).Compared to the CH group,NO(P<0.05)content and NOS(P <0.05),SOD(P <0.01),GSH-PX(P <0.01)activity was significantly increased in L-Fas group,MDA content was significantly lower(P <0.01);NO(P <0.05)content and NOS(P <0.05),SOD(P <0.01),GSH-PX(P <0.01)activity was significantly increased,MDA content was significantly decreased(P < 0.01)in H-Fas group.(4)Heart weight index.Compared to normal control group,HW,LVW,HWI,LVWI significantly increased(P <0.01)in CH group;Compared to the CH group,HW,LVW,HWI,LVWI decreased significantly(P <0.05)in L-Fas group;HW,LVW,HWI,LVWI decreased significantly(P <0.01)in H-Fas group.(5)Myocardial histopathology and immunohistochemistry.After HE and Masson staining,compared with the control group,myocardial cell volume increased,the cell gap increased,cell arranged disorder,severe myocardial interstitial fibrosis was present in CH group;After Fas treatment,cell volume was reduced,cell gap becomes small,cells arranged in neat rows,fibrosis significantly reduced.In immunohistochemistry,protein expression of RhoA,ROCK1 and ERK1/2 were found a significant increasing(P <0.01),the expression of c-FLIP protein were decreased significantly(P <0.01).Compared with the CH group,protein expression of RhoA,ROCK1 and ERK1 / 2 were decreased significantly(P <0.01),protein expression of c-FLIP was increased significantly(P <0.01)in L-Fas group;protein expression of RhoA,ROCK1 and ERK1 / 2 was significantly decreased(P <0.01),andprotein expression of c-FLIP was significant increased(P <0.01)in H-Fas group.(6)The expression of RhoA,ROCK1,c-FLIP and ERK1 / 2 mRNA in cardiac tissue.Compared with normal control group,the expression of RhoA,ROCK1 and ERK1 / 2 mRNA was significantly increased(P<0.01),the expression of c-FLIP mRNA significantly decreased(P <0.01)in CH group.Compared with the CH group,the expression of RhoA(P<0.05),ROCK1 and ERK1 / 2 mRNA was significantly lower(P <0.01),the expression of c-FLIP mRNA was significantly increased(P <0.01)in L-Fas group;The expression of RhoA,ROCK1 and ERK1 / 2 mRNA significantly reduced(P <0.01),and the c-FLIP mRNA expression(P<0.01)was significant increased in H-Fas group.Conclusion: RhoA / ROCK and ERK1 / 2 signaling pathway have joined in Iso induced CH,Fas protects against the CH of rats induced by Iso,which mechanisms may be related with blocking RhoA / ROCK and ERK1/2 signaling pathway,regulating the protein expression of RhoA,ROCK1,c-FLIP and ERK1/2,scavenging oxygen free radicals,inhibiting lipid peroxidation and promoting the release of NO.