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A2706 - Aging Mesenchymal Stem Cells Have Diminished Effects in Modulating Macrophages and Producing Extracellular Vesicles
Author Block: R. Huang1, J. Wang1, G. Zheng2, G. Qiu2, H. Tao1, Y. Hu1, M. Ge2, Q. Shu1, J. Xu3; 1The Children’s Hospital of Zhejiang University School of Medicine, Hangzhou, China, 2
Shaoxing Second Hospital, 
Shaoxing, China, 3Shaoxing Second Hospital, The First Affiliated Hospital of Zhejiang University School of Medicine, Zhejiang, China.
Rationale: Adipose-derived stem cells (ASCs) possess potent immunosuppressive and anti-inflammatory properties. Previously, our team has demonstrated that ASCs-treated macrophages (ASCs-M) were polarized towards M2 macrophages and mimicked ASCs in alleviating LPS-induced systemic response. The goals of this study are to determine whether aging and young ASCs have the same effects in modulating macrophages and investigate whether the discrepancy between young and aging ASCs could be accounted for by their capability in generating extracellular vesicles (EVs).Methods: For this study in vitro, peritoneal macrophages were cultured alone or co-cultured with young or aging ASCs for 48h via Transwell, and then were incubated with or without LPS for 24h. Cells were collected for phenotype analysis via flow cytometry. For the study in vivo, C57BL/6 mice were randomly assigned to five groups: PBS control, LPS, LPS + untreated macrophages, LPS + young ASCs-M, LPS + aging ASCs-M. LPS was administered to mice intraperitoneally at 5 mg/Kg. Immediately after PBS or LPS treatment, 1 × 106 cells or PBS were injected via the tail veins of the mice. Lung, serum and bronchoalveolar lavage fluid (BAL) were acquired after 6h, 24h and 48h for examination. To determine the mechanisms of the discrepancy, EVs derived from young or aging ASCs were collected to compare the quantity, quality, and the uptake by macrophages in vitro. Results: In vitro, young ASCs significantly elevated M2 phenotype, while aging ASCs lost the effect on M2 polarization. Young but not aging ASCs-M directly ameliorated LPS-induced lung injury in vivo evidenced by improved histological change, reduced weight loss, lower wet/dry ratio, and decreased albumin concentration in BAL. The changes of pro-inflammatory and anti-inflammatory cytokines levels also confirmed the protective effect of young ASCs-M. While young and aging ASCs-derived EVs exhibited similar characteristics in size, surface makers, and morphology, the aging ASCs generated significantly less quantity of EVs as determined by EVs number and protein level. Furthermore, macrophages uptake significantly less amount of aging ASCs-derived EVs as compared with young counterpart.Conclusion: Adoptive transfer of young but not aging ASC-M alleviates LPS-induced lung injury and systemic response. Aging ASCs have reduced ability in polarizing M2 macrophages, which may be the result of diminished ability in generating EVs.