In Vivo Retroviral Transduction of Cardiac Myofibroblasts Using Intramyocardial Injection Immediately Post-myocardial...

Following myocardial infarction (MI), myocardial cells undergo cell death, and the necrotic region is replaced by extracellular matrix (ECM) proteins such as collagens. Myofibroblasts are responsible for producing these ECM proteins. Cardiac myofibroblasts are differentiated from resident fibroblasts in response to inflammation. To date, genetically modified mice driven by the Periostin promoter and adeno-associated virus 9 (AAV9) carrying the Periostin promoter have been used for gene transfer into cardiac myofibroblasts. However, these methods require multiple steps and are time-consuming and expensive. Therefore, we developed a method for delivering genes into cardiac myofibroblasts using retroviruses. Specifically, the DNA of the target gene was transfected into Plat-E cells, which are packaging cells, to generate retroviruses. The virus-containing supernatant was then harvested, and the viruses were pelleted by centrifugation and suspended in PBS-containing polybrene. Subsequently, permanent occlusion of the left coronary artery was performed, and 20 μL of viral solution was immediately administered using a 29G needle at a position 1-2 mm below the ligation site in the heart of mice maintained in an open chest state. Using this method, we were able to introduce genes into the myofibroblasts of interest surrounding the MI site. Key features • Retroviruses are taken up only by proliferating cells, enabling highly specific gene transfer into myofibroblasts. • Any gene incorporated into the genome by retroviruses will continue to be expressed over the long term, providing chronic in vivo evaluation. • Myocardial injection targeting the infarct area of the left ventricle shows high infection efficiency in myofibroblasts. • This protocol employs a very small-scale and simple virus concentration method.