(25e) Shear Stress in a Spinner Flask Bioreactor Alters Energy Metabolism and Replicative Senescence of Human Mesenchymal Stem Cells

Human mesenchymal stem cells (hMSCs) have been coveted in regenerative medicine for their multipotent differentiation abilities, secretome, and immunoprivilege. As a direct result, the number of clinical trials and publications has grown exponentially over the last decade highlighting the need for in vitro systems capable of producing large quantities of therapeutically potent cells in order to meet growing demands. However, hMSCs are highly sensitive to microenvironment and biophysical conditions, including shear stress caused by dynamic agitation. In this study, hMSCs grown on plastic plus microcarriers were subjected to varying shear stresses by increasing agitation rates (50 rpm and 80 rpm) in a spinner flask bioreactor system. Our results demonstrated a reduction in proliferation at 80 rpm compared to lower agitation. Real-time polymerase chain reaction analysis revealed increased mRNA expression of genes associated with cell cycle regulation, nicotinamide adenine dinucleotide (NAD+) metabolism, glycolysis, and the pentose phosphate pathway in spinner flask culture as well as altered cytokine secretion (e.g., PGE2 and CXCL10) when simulating an inflammatory microenvironment via interferon-gamma. These results indicate that the behaviors of replicative senescence of the hMSCs in bioreactors may be different from conventional planar cultures, showing the biophysical effects of culture environment on homeostasis of hMSCs. The reduced pro-inflammatory and the enhanced anti-inflammatory properties demonstrate the advantages of bioreactor systems in bio-manufacturing of hMSCs. This study provides advanced understanding of hMSC sensitivity to the bioreactor microenvironment while scaling production for clinical trials and bio-manufacturing.

This study is supported by National Science Foundation (NSF 1743426)-CBET: Advanced Biomanufacturing program.