The Efficiency And Safety Of Recombinant Adeno-associated Virus Serotype 9 Transfected By Different Methods In Rat Myocardium

Objective Recombinant adeno-associated virus serotype 9(r AAV9)can express the target gene continuously and highly as a new virus vector.There are several approaches to transfer r AAV9 into rat myocardium.Among these approaches,the transfection efficiency and safety are different.This study was aimed to explore the transfection efficiency and safety using r AAV9 by multi ways.Methods One hundred male Sprague-Dawley rats were randomly and equally divided into four groups : Tail vein injection(TVI)group,Tail vein pumping(TVP)group,multi-point intramyocardial injection(IMI)group and intracoronary injection(ICI)group.Then each group was sub-divived five groups:negative control(NC)group(n=5),transfected groups that duration for 7days,14 days,21 days and 28 days respectively.The rats in all transfected groups were injected with 1×1011 vector genome of AAV9 carrying labeled enhanced green fluorescence protein(e GFP)gene,while the rats of all the NC subgroups were injected with the same volume of normal saline.The optimal time for transfection and the transfection efficiency were determined by assessing the e GFP expression in rat myocardium,and the safety was decided by both measurements of the levels of creatine kinase MB isoenzyme(CK-MB)and cardiac troponin T(c Tn T)and evaluations of cardiac function by echocardiogram and by electrocardiogram.Results r AAV9 can be effectively transferred to rat myocardium with these methods mentioned above.The expressions of e GFP in all tranfection groups reached highest 14 days after transfection and then declined slightly.In these groups,transfection efficiency from high to low was:ICI group,TVP group,IM group,TVI in turns.In each group,there is no significant difference both in the levels of CK-MB and c Tn T and in cardiac function between transfected subgroups and NC subgroups.For all transfacted subgroups,the levels of CK-MB and c Tn T in the two groups which were transferred with r AAV9 by tail vein were lower than IMI and ICI groups.Compared to TVI and TVP groups,the cardiac function of IMI and ICI groups decreased markly.Severe cardiac arrhythmia(include ventricular tachycardia,ventricular fibrillation,cardiac arrest)did not occurred in the TVI and TVP groups,while itoccurred in IMI and ICI groups.There were nothing abnormal detected in heart samples from TVI and TVP groups,but scar formation and adhesion between chest wall and heart-lung in IMI and ICI groups were detected.Conclusion r AAV9 can be transfected into rat myocardium safely and efficiently.Compared to intracoronary injection,transferring into rat myocardium by tail vein pumping has a slightly lower tranfection efficiency and a higher safety.