The Effects Of G-CSF On Pressure Overload-induced Ventricular Remodeling And Heart Failure In Mice

Accumulating evidences indicate that cardiac hypertrophy is a fundamental process of adaptation to an increased workload due to hemodynamic overload. Development of cardiac hypertrophy is initially beneficial since it augments the number of contractile units and reduces ventricular wall stress to normal levels according to the law of Laplace.However,the adaptation has its limits and heart failure may ensue.Furthermore,arrhythmias and ischaemic heart disease may develop which increase the risk of sudden death.And the structural basis of the progression from well-compensated hypertrophy caused by mechanical overload to heart failure(HF)is still largely unknown.It is evident that cardiac remodeling,defined as "genome expression,molecular, cellular,and interstitial changes that are manifested clinically as changes in size,shape and function of the heart after injury" occurs in the chronically pressure-overloaded heart.However,the correlation between morphological alterations and cardiac function status during the different phases of transition to HF has not yet been described in the animal model.Granulocyte colony-stimulating factor(G-CSF) is a hematopoietic cytokine that promotes proliferation and differentiation of neutrophil progenitors.It was recently reported that G-CSF mobilizes bone marrow stem cells(BMSCs) into the infarcted hearts and accelerates the differentiation into vascular cells and cardiac myocytes.In animal models,G-CSF shows protective effects of preventing left ventricular remodeling and dysfunction after acute myocardial infarction.Although it is controversial whether BMSCs mobilized by G-CSF can differentiate into cardiac myocytes,G-CSF-induced angiogenesis is indeed recognized in infarcted heart.The cardioprotective effects of G-CSF are recognized even in isolated perfused heart.In addition,G-CSF activates various signaling pathways such as Akt,extracellular signal-regulated kinase, and Janus kinase 2/signal transducer and activator of transcription 3 through G-CSF receptors in cardiac myocytes.These observations suggest that G-CSF not only induces mobilization of stem cells and progenitor cells but also acts directly on cardiomyocytes.Therefore,G-CSF may be utilized as a novel agent to have protective effects on non-ischemic injured myocardium.In the present study,we established a mice model of pressure overload induced by transverse aorta constriction(TAC) after intramyocardial transgene and investigated the effects of G-CSF on pressure overload-induced ventricular remodeling and heart failure in mice and its mechanism. PartⅠBiological characteristics of mice model of intramyocardial transgene after pressure overloadAim:This study was aimed to establish a mice model of pressure overload induced by transverse aorta constriction(TAC) after intramyocardial transgene and to primarily obtain a profile knowledge of biological characteristics of this model,including body weight,survival rate and cardiac function status.Methods:Forty-four C57BL/6 male mice were randomly divided into transverse aorta constricted group(operated group) and sham-operated group(control group).The operated group was divided into LacZ and PBS group after left ventricular injection of Ad-LacZ or PBS with temporarily blocking aortic blood flow for 5,10,20 and 30 s,respectively.The weight,carotid blood pressure and cardiac coefficient were measured during 4 weeks after TAC.The thickness of left ventricular wall,heart function and left ventricular inner diameter were measured by echocardiography.The morphology of left ventricle and the degree of cardiac blue staining were observed by Hematoxylin-eosin and X-gal staining.Results:The overall survival rate was 88.8%.Compared with sham-operated group,the carotid blood pressure in constricted group was increased markedly after TAC (P＜0.01).Left ventricular hypertrophy appeared in 2 weeks and heart failure in 4 weeks.The heart tissue appeared blue in models with aortic blood flow blocking for more than 5 s,whereas there was no difference among 10 s,20 s and 30 s.Conclusions:It is a practical and reproducible model of pressure overload after intramyocardial transgene.The efficacy of gene expression may be better in mice with aortic blood flow blocking for more than 10 s after left ventricular injection of Ad-LacZ.PartⅡEffects of G-CSF on pressure overload-induced ventricular remodeling and heart failure in mice Aim:This study was aimed to explore the effects of granulocyte colony-stimulating factor(G-CSF) on pressure overload induced ventricular remodeling and heart failure in mice and its mechanism.Methods:Eighty six 10～12-week-old male mice were randomly assigned into seven groups:PBS group(PBS,n=14),telmisartan group(ST,n=12),G-CSF(A) group(GA, n=12),G-CSF(B) group(GB,n=12),G-CSF combined with telmisartan group (GT,n=12),telmisartan combined with G-CSF group(TG,n=12) and sham-operated group(Sham,n=12).The transverse ascending aorta constriction(TAC) in six groups were performed in order to make heart failure model.Mice in each group were injected with G-CSF or/and telmisartan subcutaneously at different time respectively.Cardiac function was assessed by echocardiography every week after TAC.Animals were sacrificed in batches at 1,2 and 4 weeks postoperatively after blood pressure of right carotid artery was measured.Morphological change in left ventricle were detected by HE staining,Masson staining.The expression of vascular endothelial growth factor(VEGF) in myocardium was detected by Western blotting.The mRNA expression of p53 and hypoxia-inducible factor 1(HIF-1) in myocardium were detected by semi-quantitative RT-PCR.Results:(1) The end-systolic thickness of left ventricular walI(LVWTs) in PBS group increased gradually and reached peak at 14 days,then decreased gradually with left ventricular ejection fraction(LVEF) from 14 to 28 days.LVWTs increased gradually in mice injected with G-CSF during 14 days,but there was no significant change with LVEF from 14 to 28 days(P＜0.05).(2) Compared with PBS group,the expression of VEGF protein and HIF-1mRNA were significantly higher in mice injected with G-CSF or/and telmisartan(P＜0.05).The expression of p53 mRNA,myocardial fibrosis and mortality were significantly lower in mice injected with G-CSF or/ and telmisartan(P＜0.05).Conclusions:G-CSF can delay the progression of pressure overload induced ventricular remodeling and heart failure in mice.The regulation of HIF-1 maybe an important underlying mechanism. Conclusions1.Using the method of transverse aorta constriction after left ventricular direct injection of Ad-LacZ can quickly establish a transgened mice model of pressure overload.The efficacy of gene expression may be better in mice with aortic blood flow blocking for more than 10 s after left ventricular injection of Ad-LacZ.2.G-CSF can delay the progression of pressure overload induced ventricular remodeling and heart failure in mice.The regulation of HIF-1 maybe an important underlying mechanism.The potential application and novelty of this project1.We established a transgened mice model by successfully transferring target gene into cardiomyocyte through adenovirus vector for the first time.2.Using the method of transverse aorta constriction after left ventricular direct injection of Ad-LacZ,we successfully simulated the whole pathophsiological process from cardiac hypertrophy to heart failure in mice,thereby providing a practical experimental model for the research of molecular mechanism of cardiac hypertrophy.3.We found that G-CSF can delay the progression of pressure overload induced ventricular remodeling and heart failure in mice,and the regulation of HIF-1 maybe an important underlying mechanism.