hESC expansion and stemness are independent of connexin forty-three-mediated intercellular communication between hESCs and hASC feeder cells.

Abstract

BACKGROUND: Human embryonic stem cells (hESCs) are a promising and powerful source of cells for applications in regenerative medicine, tissue engineering, cell-based therapies, and drug discovery. Many researchers have employed conventional culture techniques using feeder cells to expand hESCs in significant numbers, although feeder-free culture techniques have recently been developed. In regard to stem cell expansion, gap junctional intercellular communication (GJIC) is thought to play an important role in hESC survival and differentiation. Indeed, it has been reported that hESC-hESC communication through connexin 43 (Cx43, one of the major gap junctional proteins) is crucial for the maintenance of hESC stemness during expansion. However, the role of GJIC between hESCs and feeder cells is unclear and has not yet been reported. METHODOLOGY/PRINCIPAL FINDINGS: This study therefore examined whether a direct Cx43-mediated interaction between hESCs and human adipose-derived stem cells (hASCs) influences the maintenance of hESC stemness. Over 10 passages, hESCs cultured on a layer of Cx43-downregulated hASC feeder cells showed normal morphology, proliferation (colony growth), and stemness, as assessed by alkaline phosphatase (AP), OCT4 (POU5F1-Human gene Nomenclature Database), SOX2, and NANOG expression. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that Cx43-mediated GJIC between hESCs and hASC feeder cells is not an important factor for the conservation of hESC stemness and expansion.