The different gene methylation levels of rat Schwann cells detected by MeDIP-Seq / 天津医药

ABSTRACT

Objective To explore the difference of DNA methylation levels between normal Schwann cells (NSCs) and activated Schwann cells (ASCs) in rats. Methods The adult Wistar rats were received sciatic nerve ligation and fed for 7 days. The ASCs and NSCs were separated from ligated sciatic nerves and brachial plexus respectively. Immunocytochemical staining of S-100 antibody was used to identify the cells. The growth condition of cells was detected by CCK-8 method. Methylated DNA immunoprecipitation sequencing (MeDIP-Seq) was applied to filter the differentially methylated regions in ASCs and NSCs. The distribution of differentially methylated genes related with axonal regeneration in chromosome was analyzed, and Gene ontology(GO)and PATHWAY analysis were also conducted. Results High purity of ASCs and NSCs were obtained successfully, which were both positive for S-100 antibody. In the same culture condition, ASCs showed a faster proliferation than that of NSCs. A total of 177 176 differentially methylated regions were found by MeDIP-Seq. Among them, 1 097 were located in the promoter (≤1 kb), 1 136 in the promoter (1-2 kb) and 567 on the CpG. After functional annotation of differentially methylated genes, 214 differentially methylated genes related with axonal regeneration were found in ASCs and NSCs. Compared with NSCs, 191 genes were up-regulated and 23 genes were down-regulated in ASCs. These genes were located on different chromosomes, most of which on chromosome 12 (22 genes) and the least on chromosomes M (2 genes). GO analysis indicated that the differential methylated genes were involved in axon growth, axon formation, axon elongation and axon guidance. The MAPK, cell adhesion molecules, Ras signaling pathway may be related with the differential methylated genes. Conclusion The methylation levels between ASCs and NSCs are significantly different, which are probably related with axon regeneration.