ENGINEERING NOVEL SURFACES FOR THE LONG-TERM PROPAGATION OF HUMAN PLURIPOTENT STEM CELLS.

Abstract

Defined substrate that can maintain pluripotent stem cells long term is critical for its future regenerative medicine applications. This thesis aims to engineer a tuned heparan sulfate (HS) substrate that is able to bind and present sufficient vitronectin (VN) for the long-term propagation of human embryonic stem cells (hESCs). Results showed that passive adsorption of VN onto tissue-culture treated polystyrene surfaces is inefficient (250 ng/cm2). Therefore by using affinity chromatography technique, this thesis isolated a VN-binding fraction of HS that is able to be immobilized onto allylamine polymerized TCPS by electrostatic interactions which in turn bind and present sufficient VN on surface (150 ng/cm2). The hESCs are able to proliferate on this engineered substrate and continued to retain its pluripotency (morphology, genetic stability, differentiation) for more than 8 passages. This lower VN surface density required on the HS substrate has resulted in a significant savings of ~ 30 %.