Abstract: Objective To analyse the change of DNA methylation with 5-Azac injection in acute graft-versus-host disease (aGVHD) mouse model, which received allogeneic bone marrow transplantation, and explore the immunomodulatory effects of 5-Azac. Methods Male C57BL/6 (H-2b） and female BALB/c (H-2d) mice were selected as donor and recipient of complete allotransplantation. BABL/c mice were divided into two groups, transplantation control group and 5-Azac experi⁃ mental group. At 1-7, 14, 21 and 28-day after transplantion, 5-Azac 0.25 mg/kg (0.3 mL/time) was injected by tail vein in experimental group, while the control group were injected with sterile water 0.3 mL/time. Peripheral blood DNA samples were collected from three control mice and three experimental mice, then mixed with equal amount respectively. The MeDIPseq method was selected to detect methylation changes in mice, and the differential DNA methylation in the biological pathways was analyzed. Results The survival time was prolonged, and the rejection reaction was decreased in 5-Azac experi⁃ mental group, which suggested immune hyporesponsiveness post aGVHD. The MeDIP-seq result showed that 369 different DNA methylation located in the promoter regions, including 239 up-regulated genes and 130 down-regulated genes. There were 184 differential DNA methylation genes located in the exon regions, including 113 up-regulated genes and 71 downregulated genes. Differential DNA methylation genes involved in 10 immunological signaling pathways, respectively. Among them, TGF-β, GSK-3β, SYK, PI3K, NFAT, CD28 and α4β7 were closely related to the development of aGVHD. Conclusion 5-Azac can effectively induce immune hyporesponsiveness post aGVHD by changing the gene methylation status.

Key words: graft vs host disease, methylation, 5-azacytidine, MeDIP-seq, immune hyporesponsiveness