Sustained Activation of AMPK Enhances Differentiation of Human iPSC-Derived Cardiomyocytes via Sirtuin Activation

Abstract

Summary Recent studies suggest that metabolic regulation may improve differentiation of cardiomyocytes derived from induced pluripotent stem cells (iPSCs). AMP-activated protein kinase (AMPK) is a master regulator of metabolic activities. We investigated whether AMPK participates in iPSC-derived cardiomyocyte differentiation. We observed that AMPK phosphorylation at Thr172 increased at day 9 but then decreased after day 11 of differentiation to cardiomyocytes. Inhibition of AMPK with compound C significantly reduced mRNA and protein expression of cardiac troponins TNNT2 and TNNI3. Moreover, sustained AMPK activation using AICAR from days 9 to 14 of differentiation increased mRNA and protein expression of both TNNT2 and TNNI3. AICAR decreased acetylation of histone 3 at Lys9 and 56 and histone 4 at Lys16 (known target sites for nuclear-localized sirtuins [SIRT1, SIRT6]), suggesting that AMPK activation enhances sirtuin activity. Sustained AMPK activation during days 9–14 of differentiation induces sirtuin-mediated histone deacetylation and may enhance cardiomyocyte differentiation from iPSCs.

Activation of AMPK Promotes Maturation of Cardiomyocytes Derived From Human Induced Pluripotent Stem Cells

“…AMPK is energy-sensing protein kinase that plays a key role in the regulation of cellular energy metabolism. Several studies have shown that relatively mature hiPSC-CMs have highly phosphorylated AMPK ( Yang et al, 2019 ; Sarikhani et al, 2020 ). AMPK has been shown to play a protective role in various cardiac pathophysiological processes to restore energy balance ( Dolinsky and Dyck, 2006 ).…”

“…Studies have reported that the activation of AMPK increases fatty acid uptake and inhibits fatty acid synthesis by promoting fatty acid transport proteins in the cell membrane and increasing fatty acid β-oxidation ( Arad et al, 2007 ). Sustained AMPK activation led to increased glucose and fatty acid uptake and an increase in the mRNA and protein expression of GLUT4 and CD36 concomitant to an increased in oxygen consumption rate (OCR) ( Sarikhani et al, 2020 ). Previous studies demonstrated that fatty acid treatment of hPSC-CMs induced phosphorylation of AMPK, which ultimately increased cardiomyocyte maturation ( Horikoshi et al, 2019 ; Yang et al, 2019 ).…”

Mitochondria and metabolic transitions in cardiomyocytes: lessons from development for stem cell-derived cardiomyocytes

“…Skeletal myoblasts with depletion of mitochondrial DNA or inhibition of mitochondrial complex III with antimycin A had reduced expression of Kir2.1 leading to membrane depolarization, suggesting that impaired oxidative phosphorylation can have direct adverse effects on resting membrane potential [290]. AMPK activation downregulates Kir2.1 [291], and we recently found that sustained activation can enhance maturation of contractile and metabolic features of PSC-CMs but also reduces expression of KCNJ2 [292]. In contrast, in neurons, mTOR signaling increases expression of HCN channels [293].…”

AMPKβ1 and AMPKβ2 define an isoform-specific gene signature in human pluripotent stem cells, differentially mediating cardiac lineage specification

“…Along this line, it was shown recently that pharmacological activation of AMPK in cardiac progenitor cells under differentiation is neither sufficient to alter gene expression of differentiation and CM markers, nor to promote PPARGC1A transcription and mitochondrial biogenesis (70). A recent publication suggest increase in AMPK activation during cardiac differentiation of hiPSCs, reaching highest levels of phosphorylation at day 9 and 11 followed by a drop of protein activation at day 14 (71). Our data does not confirm this finding, which might be due to differences in the differentiation protocol.…”