Abstract 14217: Myofibroblasts Enhance the Intrinsic Sinus Rhythm at Low Cell-Cell Coupling

Abstract

Heart rhythm is initiated by the sinoatrial node (SAN). Compared to the chamber myocardium, the healthy SAN exhibits an exceedingly high content of fibroblasts (FBs). Developmental, aging and pathophysiological cues can activate FBs to myofibroblasts (MFBs) which exhibit depolarized membrane potential and stronger electrotonic coupling compared to FBs. We hypothesized that MFBs, at a low content, can paradoxically increase the pacing frequency of pacemaker cells (PMs) by contributing to the spontaneous depolarization slope.Monolayers and scaffold-free 3D spheroids of TBX18-induced PMs (iPMs) were generated as in vitro models of SAN by somatic gene transfer of TBX18 to neonatal rat cardiomyocytes. Retrograde-perfused, adult rat heart was employed to study intact SAN ex vivo. In silico modeling was performed to simulate pacing frequencies of PMs coupled to MFBs in 2D and 3D pacemaker tissues. Upon reducing electrotonic coupling with carbenoxolone (CBX, 5μM), pacing frequency of iPM monolayers either increased from 66 ± 51 to 168 ± 92 bpm (n=4/13) or decreased from 125 ± 47 to 98 ± 28 bpm (n=9/13). GFP-transduced control NRVM monolayers (n=8) showed only occasional activity, which was not affected by CBX treatment (|ΔBR| = 57 ± 69 in iPM monolayers vs. 11± 11 bpm in control, GFP monolayers, p<0.05). In culture, FBs are known to undergo spontaneous activation to MFBs via TGFβ signaling. Treatment of iPM spheroids with a TGFβ inhibitor (SB431542, 10μM) slowed their automaticity to 124 ± 89 bpm compared to 214 ± 18 bpm in untreated iPM spheroids. Sinus rhythm of the rat heart ex vivo increased upon CBX treatment from 199 ± 64 to 242 ± 80 bpm (n=3, p=0.07). Immunostaining confirmed presence of αSMA+ MFBs in both iPMs and the SAN. Computational study demonstrated that pacing frequencies of 2D and 3D PMs rose and fell with increasing gap junction conductance when coupled to MFBs (Vrest = -25 mV, g = 0 to 4.0 nS) but not to FBs (Vrest = -50 mV, g = 0 to 0.4 nS). Dynamic range of cardiac pacing was optimal when the ratio of MFB to PM was no more than one.Our data demonstrate that MFBs positively contribute to the pacing frequency and the safety factor of the sinus rhythm. We identify fibroblast activation state as a potential target to rejuvenate heart rhythm under aging and pathological conditions.