Abstract Body: Background: Conventional MSCs and iiPSC based clinical trials revealed inconsistent outcomes, and minimal improvement in heart function following a MI. Endogenous left ventricular stem cells (LVSC) are superior in cardiac regeneration (compared to other sources) due to their lineage specificity and intrinsic cardiac origin, despite the very limited attempts in translational arena.
Hypothesis: rLVSCs are optimally suited to accelerate healing at the peri-infarct zone by replenishing cardiomyocyte (CM)-like cells to promote survival responses.
Aim: To determine the regenerative potential of LVSC.
Approach: We screened LGALS1+ rLVSCs harvested from MI-swine model using scRNA-seq, rLVSCs were differentiated to CM-like cells by manipulating Wnt signaling, assessing metabolic modulation by mitochondrial health status and regenerative mechanism was determined by knocking out LGALS1.
Results: We defined regenerative phenotype of LVSCs (rLVSCs), featured with the upregulation of the cardioprotective protein LGALS1 (Galactin-1) at single cell resolution. rLVSCs demonstrated enhanced differentiation potential into CM-like cells as evident by the formation of cardiac embryoid bodies (EBs) and decreased resistance to senescence upon ischemia challenge. Also, rLVSCs were differentiated to CM-lineage upon manipulating the Wnt signaling resulted in tube like cell morphology, statistically significant upregulation of CM-specific genes including GATA4, Nkx2.5, Cx43 at transcript and protein levels, increased K+ efflux, and non-synchronized contractile movements. Knocking out and silencing LGALS1 in rLVSCs resulted in the concomitant downregulation of key CM-specific genes, including GATA4, Nkx2.5, IRX5, and TBX5, confirming its critical role in cardiac regeneration by differentiation into CM-like cells. We observed an increased density of electron-rich mitochondria in the ischemic rLVSCs and in the post-differentiation CM-like cells. Also, the CM-like cells exhibited increased oxygen consumption and decreased anaerobic glycolysis suggestive of a switch to oxidative phosphorylation for energy requirements representing mature CMs as evident from Seahorse assay.
Conclusion: The protective events in rLVSCs mirror their ability to modulate metabolism supporting the regeneration of cardiac function. Overall, rLVSCs represent an ideal sub-population for myocardial regeneration demonstrating immense translational promise in MI management.