Comparison of Immunomodulatory Functions in Human Umbilical Cord Mesenchymal Stem Cells after Long-Term Expansion
CHANG Cheng1, LIU Mengting2, ZHANG Quan2, RAO Wei2, CHEN Lian1, HE Jing1, HAN Bing2, LI Changyong1, WU Dongcheng1,2*
The purpose of this study was to systematically compare different passages of human umbilical cord mesenchymal stem cells, explore the effect of long-term subculture on their immune regulatory function and possible mechanisms. The cells were cultured to 5th, 10th and 15th passage, and their differentiation potential and surface markers were determined. Cell senescence and apoptosis were detected by β-galactosidase staining and flow cytometry. Co-cultured with each passages of umbilical cord mesenchymal stem cells under different conditions to compare lymphocytes proliferation, cell cycle changes and T cell subsets in human peripheral blood lymphocytes. ELISA was used to detect the concentration of TNF-α, IFN-γ, IL-β and IL-6 in the co-culture system. Western blot was used to detect the expression of PD-L1 and IDO in hUC-MSCs at different passages. The study indicated that hUC-MSCs were cultured to the 15th passage with no significant change in biological characteristics of stem cell, but the number of β-galactosidase staining positive cells was significantly increased. Three passages of hUC-MSCs could regulate cell cycle in activated lymphocytes, effectively suppressed proliferation and Th1/Th17 cell subsets, affected the expression of inflammatory factors. The 15th passage cells showed deregulation of Treg subsets and IL-1β. hUC-MSCs could maintain their multi-directional differentiation potential and satisfactory immunomodulatory activity after long-term subculture; however, it could not be ignored that the disability on regulation of Tregs subsets and anti-inflammatory in high-passages cells which were aging. In clinical applications, hUC-MSCs within 10th passage should be used to ensure the safety and efficacy in treatment.
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