Defined factors to reactivate cell cycle activity in adult mouse cardiomyocytes

Abstract

Adult mammalian cardiomyocytes exit the cell cycle during the neonatal period, commensurate with the loss of regenerative capacity in adult mammalian hearts. We established conditions for long-term culture of adult mouse cardiomyocytes that are genetically labeled with fluorescence. This technique permits reliable analyses of proliferation of pre-existing cardiomyocytes without complications from cardiomyocyte marker expression loss due to dedifferentiation or significant contribution from cardiac progenitor cell expansion and differentiation in culture. Using this system, we took a candidate gene approach to screen for fetal-specific proliferative gene programs that can induce proliferation of adult mouse cardiomyocytes. Using pooled gene delivery and subtractive gene elimination, we identified a novel functional interaction between E2f Transcription Factor 2 (E2f2) and Brain Expressed X-Linked (Bex)/Transcription elongation factor Alike (Tceal) superfamily members Bex1 and Tceal8. Specifically, Bex1 and Tceal8 both preserved cell viability during E2f2-induced cell cycle re-entry. Although Tceal8 inhibited E2f2-induced S-phase re-entry, Bex1 facilitated DNA synthesis while inhibiting cell death. In sum, our study provides a valuable method for adult cardiomyocyte proliferation research and suggests that Bex family proteins may function in modulating cell proliferation and death decisions during cardiomyocyte development and maturation.

Transcriptional Regulation of Postnatal Cardiomyocyte Maturation and Regeneration

“…For example, AKT-mediated nuclear localization of the forkhead box transcription factor FoxM1 promotes cardiomyocyte proliferation in the developing heart [ 69 ], but FoxM1 expression decreases during the postnatal period [ 19 ] ( Figure 1 A). Likewise, the transcription factors E2f2 and E2f4 promote embryonic cardiomyocyte mitotic activity and are downregulated postnatally ( Figure 1 A); however, forced expression of E2f2/4 in adult cardiomyocytes leads to massive cell death [ 70 , 71 , 72 ]. Finally, Isl1 promotes embryonic cardiomyocyte proliferation by activating Fgfs and Bmps [ 73 ], which elicit a downstream proliferative response, and cooperate with Gata4 to express Hand2, another transcription factor that activates proliferative genes [ 74 , 75 ].…”

Reawakening the Intrinsic Cardiac Regenerative Potential: Molecular Strategies to Boost Dedifferentiation and Proliferation of Endogenous Cardiomyocytes

“…Similar to E2F1, E2F3 overexpression was also associated with cell death (215). Instead, contrasting data have been obtained for the apoptotic response induced by E2F2 and E2F4, with initial studies demonstrating a reduction in cell death upon E2F2 and E2F4 overexpression in cultured neonatal cardiomyocytes (215), and more recent studies describing an increase in apoptosis of cultured adult mammalian cardiomyocytes (217). Interestingly, co-expression of E2F2 and BEX1 [Brain Expressed X-Linked (Bex)] was demonstrated as a strategy to induce cardiomyocyte cell cycle activity, without cell death (217).…”

Engineering Extracellular Matrix Proteins to Enhance Cardiac Regeneration After Myocardial Infarction

“…Recent studies reported that binding of agrin to αDG could contribute to enhanced CM proliferation. To expedite such changes, modulating the stiffness of microenvironments could synergistically initiate CM proliferation via dedifferentiation–proliferation–redifferentiation of CMs (Yahalom-Ronen et al, 2015 ; Wang et al, 2017 ; Judd et al, 2019 ).…”