Small sized mesenchymal stem-cells primed with hypoxia attenuate graft-versus-host disease [DNA methylome dataset]

Mesenchymal stem-cells (MSCs) are of particular interest for treating immune-related diseases due to their immunosuppressive capacities. Here, we show that Small sized MSCs primed with Hypoxia and Calcium ion (SHC-MSCs) exhibit the enhanced functions regarding stemness and immunomodulation for treating allogeneic conflicts. Compared with naïve cultured human umbilical cord-blood MSCs, SHC-MSCs were resistant to the passage dependent cellular senescence mediated by MCP-1 and p53/p21 cascade and highly secreted the pro-angiogenic and immune-modulatory factors, resulting in suppression of T-cell proliferation. Genome-wide DNA methylome and transcriptome analysis indicate that SHC-MSCs characteristically up-regulated immune-modulation, cell adhesion and cell-cycle related genes. As downstream factors, PLK1, ZNF143, DHRS3, and FOG2 proteins played a key role on the beneficial effects of SHC-MSCs, evidenced by the promoted self-renewal, migration, pro-angiogenic, anti-inflammatory, and T cell suppression capacities in their-over-expressing MSCs. Importantly, administration of SHC-MSCs or PLK1-over-expressing cells (PLK1-MSCs) significantly reduced the symptoms of graft-versus-host disease (GVHD) in a humanized mouse model which led to significantly improved survival, less weight loss, and less histopathologic injuries of GVHD target organs compared with naive MSC-infused mice. Collectively, our study suggests that small-sized MSCs primed with hypoxia could advance the therapeutic strategy for the clinical treatment of allogeneic conflicts including GVHD. Human umbilical cord blood derived mesenchymal stem cells (UCB-MSCs) were normally maintained (NT) or cultivated by SHC procedure (SHC) which enriched small sized cells and at the same time primed with hypoxia ( 3 % oxygen) and calcium ion (1.8 mM). Genomic DNA (gDNA) were isolated using the DNeasy Blood & Tissue Kit (Qiagen Inc, Valencia, CA, USA). 500 ng of gDNA were used in bisulfite conversion with Zymo EZ DNA methylation kit (Zymo Research, Irvine, CA) and the DNA methylome of naïve and SHC mesenchymal stem cells (MSCs) was analyzed using on a Infinium HumanMethylation450K BeadChip assay (Illumina Inc, San Diego, CA), in accordance with the manufacturer’s instructions.