Abstract 18646: Low Metabolic Demand of Induced Pacemaker Cells Enhances Their Survival and Automaticity

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Abstract

The automaticity of cardiac pacemaker cells is central to sinus rhythm generation. As such, metabolism of the pacemaker cells may be optimized to support their survival. However, the scarcity of native sinoatrial node pacemaker cells has impeded gaining insights. We previously demonstrated creation of induced pacemaker cells (iPMs) with singular expression of a transcription factor, TBX18. We hypothesized that the iPMs are similar to native PMs in their mitochondrial architecture, but are distinct from chamber cardiomyocytes in their metabolic signature. The iPMs were created by reprogramming neonatal rat cardiomyocytes (NRVMs) with TBX18. Live-cell, super-resolution imaging illustrated that the mitochondria are smaller and globular in the iPMs and native PMs in contrast to large, tubular networks in chamber cardiomyocytes. This suggested lower Ox-Phos capacity in pacemaker cells relative to chamber cardiomyocytes. Indeed, both basal and maximal oxygen consumption rates were 51±0.3% and 29%±1.4% lower in the iPMs compared to control (n=6). The glycogen storage and secreted lactate contents were also lower in the iPMs (48±17% and 60±3.9%, respectively) compared to control. Upon 1h of 1% O2 hypoxia, control NRVMs exhibited glycogen breakdown (0.04±0.007μg/μg protein) and increased lactate secretion (18±4nmol/0.1M cells), whereas the iPMs did not show significant differences. The iPMs were more resistant to metabolic stress, showing lower cell death upon inhibition of glycolysis (NRVMs: 32±6% vs iPMs: 15±2%) and 1% O2 hypoxia (NRVMs: 52±5% vs iPMs: 7±2%). Mass spectrometry coupled with two-dimensional liquid chromatography (2D-LC-MS) revealed that among >7000 proteins, Ox-Phos and glycolysis metabolism-related proteins were downregulated in the iPMs compared to control, including a mitochondrial fusogenic molecule, Opa1. Further knockdown of Opa1 by siRNA significantly increased the number of spontaneous and synchronous action potentials in the iPMs compared to the iPMs with no Opa1 siRNA knockdown. Taken together, our data demonstrate lower Ox-Phos and total energy demand in the cardiac pacemaker cells compared to the working cardiomyocytes. The lower metabolic load renders positive impact on the iPMs’ automaticity and survival.

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