Large-scale expansion of Wharton's jelly-derived mesenchymal stem cells on gelatin microbeads, with retention of self-renewal and multipotency characteristics and the capacity for enhancing skin wound healing

Abstract

Introduction: Successful stem cell therapy relies on large-scale generation of stem cells and their maintenance in a proliferative multipotent state. This study aimed to establish a three-dimension culture system for large-scale generation of hWJ-MSC and investigated the self-renewal activity, genomic stability and multi-lineage differentiation potential of such hWJ-MSC in enhancing skin wound healing. Methods: hWJ-MSC were seeded on gelatin microbeads and cultured in spinning bottles (3D). Cell proliferation, karyotype analysis, surface marker expression, multipotent differentiation (adipogenic, chondrogenic, and osteogenic potentials), and expression of core transcription factors (OCT4, SOX2, NANOG, and C-MYC), as well as their efficacy in accelerating skin wound healing, were investigated and compared with those of hWJ-MSC derived from plate cultres (2D), using in vivo and in vitro experiments. Results: hWJ-MSC attached to and proliferated on gelatin microbeads in 3D cultures reaching a maximum of 1.1-1.30∈×∈107cells on 0.5 g of microbeads by days 8-14; in contrast, hWJ-MSC derived from 2D cultures reached a maximum of 6.5 -11.5∈×∈105 cells per well in a 24-well plate by days 6-10. hWJ-MSC derived by 3D culture incorporated significantly more EdU (P∈