Affect of iPSC Passage Number on Neuronal Differentiation

Induced pluripotent stem cells (iPSCs) are a valuable resource for neurological disease-modeling and drug discovery, due to their ability to differentiate into neurons reflecting the genetics of the patient from which they are derived. iPSC-derived cultures, however, are highly variable due to differences in culture conditions. We investigated the effect of iPSC passage number on differentiation to optimize the generation of functional, mature sensory neurons (iPSC-dSNs). Three iPSC lines were differentiated into iPSC-dSNs at passage numbers within each of the following ranges: low (LP; 5-10), middle (MP; 20-26), and high (HP; 30-38). Morphology and pluripotency of the parent iPSCs were assessed prior to differentiation at each passage number. iPSC-dSNs were evaluated based on electrophysiological properties and expression of key neuronal markers. All iPSC lines displayed the same morphology and were similarly pluripotent across passage numbers. iPSC-dSNs were also morphologically comparable across passage numbers. However, the expression levels of neuronal markers and an analysis of sodium channel function indicated greater maturity in LP iPSC-dSNs. Our results demonstrate that lower passage numbers may be better suited for differentiation into peripheral sensory neurons. Further studies are warranted to elucidate factors that may contribute to the variability associated with iPSC passage number. Expression of pluripotency markers in three different iPSC lines, as well as expression of neuronal markers in the three lines after differentiation, at three different passage numbers each