Robust expansion of human pluripotent stem cells: integration of bioprocess design with transcriptomic and metabolomic characterization

ABSTRACT: Human embryonic stem cells (hESC) have an enormous potential as a source for cell replacement therapies, tissue engineering and in vitro toxicology applications. The lack of standardized and robust bioprocesses for hESC expansion in relevant quantities while maintaining their pluripotency has hindered the application of hESC and their derivatives in clinical setting. Here, we developed a scalable and well-characterized bioprocess for hESC expansion under fully-defined conditions and explored the potential of transcriptomic and metabolomic tools to evaluate the impact of culture system on hESC phenotype. Two different hESC lines (feeder-dependent and feeder-free lines) were efficiently expanded on xeno-free microcarriers in stirred culture systems. Moreover, both hESC lines maintained the expression of stemness markers such as Oct-4, Nanog, SSEA-4 and TRA1-60, and the ability to spontaneously differentiate into the three germ layers. Whole-genome transcriptome profiling revealed a phenotypic convergence between both hESC lines along the expansion process in stirred-tank bioreactor cultures, providing strong evidence on the robustness of the cultivation process to homogenize cellular phenotype. Under low oxygen tensions, results showed a metabolic rearrangement with the up-regulation of the glycolytic machinery favoring an anaerobic glycolysis Warburg-effect like phenotype, with no evidence of hypoxic stress response, in contrast to 2-dimensional culture. Overall, we report a scalable and fully-defined bioprocess for the propagation of hESC while guaranteeing product quality. Furthermore, the “omics” tools herein used provided relevant findings on the physiological/metabolic changes during the hESC expansion in environmentally-controlled stirred-tank bioreactors, which can contribute for more standardized production systems.

ORGANISM(S): Homo sapiens

PROVIDER: GSE63192 | GEO | 2014/11/13

SECONDARY ACCESSION(S): PRJNA266969

REPOSITORIES: GEO