Grafting of Connexin43 (Cx43) Overexpressing Cardiac Fibroblasts Protects from Postinfarct Arrhythmia

Objectives: In the past, we could show a reduced incidence of postinfarct ventricular tachycardia (VT) following lentiviral transduction of resident fibroblasts (FB) with the gap junction protein Cx43. However transduction efficacy was very low and the underlying mechanisms of protection were therefore also not completely understood.

Methods: In order to further prove that Cx43 overexpression in (myo) fibroblasts is critical for the anti-VT effect and to potentially enhance the numbers of Cx43 + FB within the scar we have lentivirally (LV) transduced embryonic cardiac FB (cFB) with Cx43 ex vivo. After LV transduction with either the Cx43-IRES-eGFP or IRES-eGFP construct, 2 × 105 cFB were loaded with magnetic nanoparticles overnight and injected under magnetic attraction into cryolesioned hearts. At 2 weeks postoperation cardiac function was evaluated by echocardiography and electrophysiological analysis in vivo. Grafted cells were identified based on their eGFP fluorescence, the Cx43 content of the scar was determined by western blot (WB) analysis and morphological changes investigated by histology.

Results: Overnight incubation with lv constructs resulted in a transduction efficacy of ca. 25% of the cFB and significantly increased protein expression of eGFP and Cx43, as shown by WB. Two weeks after the procedure transmural engraftment of ÉGFP + cFB could be identified within the lesioned hearts of both animal groups. Echocardiography revealed significantly increased anterior wall thickening (33.7%; n = 12 vs. 23.7% n = 18) and fractional shortening after transplantation of Cx43 expressing cFB compared to control virus cFB transplanted animals (29.9%, n = 12 vs. 25.4%, n = 18). Electrophysiology testing in vivo showed a significantly reduced VT incidence in Cx43-Fib engrafted mice compared to controls (40%, n = 15 vs. 89.5%, n = 19). WB analysis of cut out infarct areas revealed significantly increased protein levels for Cx43 (50.2 vs. 0%, n = 3), as well as for aSMA (38.6%, n = 4 vs. 10.2%, n = 3). Interestingly during histological examination, also resident native fibroblasts in vicinity to the implanted Cx43 +/eGFP + Cx43 expressing cFB displayed hints of myofibroblast transdifferentiation.

Conclusion: Taken together, Cx43 expressing cFB engraft into lesioned hearts and improve heart function by reducing post-infarct VT incidence and to our surprise also the pump function. The cellular fate of the grafted cells and their impact on other cells is currently being investigated more in detail.