Heart immunoengineering by lentiviral vector-mediated genetic modification during normothermic ex vivo perfusion

• Published in: Frontiers in immunology 2024-06, Vol.15, p.1404668
• Main Authors: Schmalkuche, Katharina, Rother, Tamina, Burgmann, Jonathan M, Voß, Henrike, Höffler, Klaus, Dogan, Günes, Ruhparwar, Arjang, Schmitto, Jan D, Blasczyk, Rainer, Figueiredo, Constanca
• Format: Article
• Language: English
• Subjects: Animals; beta 2-Microglobulin - genetics; Cytokines - metabolism; Endothelial Cells - immunology; Endothelial Cells - metabolism; Genetic Engineering; genetic modification; Genetic Vectors - genetics; Graft Survival - genetics; Graft Survival - immunology; heart immunoengineering; Heart Transplantation - methods; Histocompatibility Antigens Class I - genetics; Histocompatibility Antigens Class I - immunology; Histocompatibility Antigens Class I - metabolism; Histocompatibility Antigens Class II - genetics; Histocompatibility Antigens Class II - immunology; Histocompatibility Antigens Class II - metabolism; Humans; Immunology; lentiviral vector; Lentivirus - genetics; Myocytes, Cardiac - immunology; Myocytes, Cardiac - metabolism; normothermic ex vivo perfusion; Nuclear Proteins; Perfusion - methods; RNA, Small Interfering - genetics; Swine; Trans-Activators; transplantation
• ISSN: 1664-3224; 1664-3224
• DOI: 10.3389/fimmu.2024.1404668

Heart transplantation is associated with major hurdles, including the limited number of available organs for transplantation, the risk of rejection due to genetic discrepancies, and the burden of immunosuppression. In this study, we demonstrated the feasibility of permanent genetic engineering of the heart during perfusion. Lentiviral vectors encoding for short hairpin RNAs targeting beta2-microglobulin (shβ2m) and class II transactivator (shCIITA) were delivered to the graft during two hours of normothermic EVHP. Highly efficient genetic engineering was indicated by stable reporter gene expression in endothelial cells and cardiomyocytes. Remarkably, swine leucocyte antigen (SLA) class I and SLA class II expression levels were decreased by 66% and 76%, respectively, in the vascular endothelium. Evaluation of lactate, troponin T, and LDH levels in the perfusate and histological analysis showed no additional cell injury or tissue damage caused by lentiviral vectors. Moreover, cytokine secretion profiles (IL-6, IL-8, and TNF-α) of non-transduced and lentiviral vector-transduced hearts were comparable. This study demonstrated the generation of genetically engineered hearts without compromising tissue integrity. Downregulation of SLA expression may contribute to reduce the immunogenicity of the heart and support graft survival after allogeneic or xenogeneic transplantation.