Baffled-flow culture system enables the mass production of megakaryocytes from human embryonic stem cells by enhancing mitochondrial function

Human embryonic stem cells (hESCs) have become an ideal cell source for the ex vivo generation of megakaryocyte (MK) and platelet products for clinical applications. However, an ongoing challenge is to establish scalable culture systems to maximize the yield of stem cell-derived MKs that release platelets. We defined a specific dynamic 3D manufacturing system that could remarkably facilitate megakaryopoiesis and increase the yield of platelet-producing MKs from hESCs within a 12-day induction period. Additionally, an increased number of > 16N ploidy MKs, proplatelets, and platelets were generated from induced cells harvested on day 12 using the specific dynamic culture method. The specific dynamic culture method significantly enhanced endothelium-to-hematopoietic transition and early hematopoiesis. More importantly, MK fate was significantly facilitated in a specific dynamic manner during early hematopoiesis. Mechanistically, this dynamic culture significantly enhanced mitochondrial function via the oxidative phosphorylation pathway and caused differentiation skewing of hESCs toward megakaryopoiesis. We defined a specific dynamic culture system in a baffled-flow manner that enabled the mass production of MKs from hESCs within 12 days. This study can aid in the automatic and scalable production of MKs from stem cells using baffled-flow bioreactors and assist in the manufacturing of hESC-derived MK and platelet products. Overall design: We performed RNA-Seq and assay to analyse the molecular mechanism of promoting the production of megakaryocytes from human embryonic stem cells in Baffled-flow culture system. RC were manipulated in baffled flasks with agitation. SC, as controls, were manipulated in baffled flasks without agitation.