Self-renewal of human embryonic stem cells on defined synthetic electrospun nanofibers

Human embryonic stem cells (hESCs) are conventionally expanded and maintained in vitro on biological substrates. Synthetic electrospun polymer nanofibers have the potential to act as non-biological substrates in the culture of hESCs. Three synthetic, FDA approved polymers: poly-ɛ-caprolactone (PCL), poly-L-lactic acid (PLLA) and poly lactic-co-glycolic acid (PLGA) were electrospun as nanofibers (random or aligned conformations) on glass coverslips and their supportive role in hESC culture examined. Clonogenicity experiments demonstrated that nanofibrous scaffolds (PCL aligned and random, PLLA aligned and PLGA aligned) supported hESC adhesion and expansion. A significantly greater number of colonies were observed on PCL-aligned nanofibrous scaffolds in comparison to PLLA-aligned and PLGA-aligned substrates (p < 0.05). hESC colonies were significantly larger on PCL aligned nanofibrous substrates when compared to other polymer substrates (p < 0.05-0.001), where fiber diameter played a pivotal role in support of hESC clonogenicity (on PCL). Retention of pluripotentiality was confirmed by expression of Alkaline phosphatase, OCT-3/4 and Nanog expression on PCL scaffolds and the expression of transcripts representative of mesoderm (ACTC1), ectoderm (SOX1) and endoderm (AFP) during subsequent spontaneous in vitro differentiation. These results demonstrate the potential of nanofibers as xeno-free scaffolds supportive of hESC adhesion, self-renewal and differentiation in in vitro culture conditions.