Acquisition of a Quantitative, Stoichiometrically Conserved Ratiometric Marker of Maturation Status in Stem Cell-Derived Cardiac Myocytes

• The TNNI gene switch is a quantitative maturation signal for hiPSC-CMs
• TnI isoform ratio is necessary, but not sufficient, to establish the mature state
• TNNI protein isoform switching is stalled in hiPSC-CMs
• Gene transfer enables acquisition of the mature TNNI signature in hiPSC-CMs

SummaryThere is no consensus in the stem cell field as to what constitutes the mature cardiac myocyte. Thus, helping formalize a molecular signature for cardiac myocyte maturation would advance the field. In the mammalian heart, inactivation of the “fetal” TNNI gene, TNNI1 (ssTnI), together in temporal concert with its stoichiometric replacement by the adult TNNI gene product, TNNI3 (cTnI), represents a quantifiable ratiometric maturation signature. We examined the TNNI isoform transition in human induced pluripotent stem cell (iPSC) cardiac myocytes (hiPSC-CMs) and found the fetal TNNI signature, even during long-term culture. Rodent stem cell-derived and primary myocytes, however, transitioned to the adult TnI profile. Acute genetic engineering of hiPSC-CMs enabled a rapid conversion toward the mature TnI profile. While there is no single marker to denote the mature cardiac myocyte, we propose that tracking the cTnI:ssTnI protein isoform ratio provides a valuable maturation signature to quantify myocyte maturation status across laboratories.

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