Preliminary Study on the Abnormal DNA Methylation in T Cells from the Patients with Immune Related Pancytopenia

Objective To explore the global DNA methylation and the expression of regulatory genes for methylation in CD4 ⁺ T cells of the patients with immune related pancytopenia (IRP) and explore the role of methylation in pathogenesis of IRP.

Methods Thirty IRP patients (untreated, n=15; remission, n=15) and 15 healthy donors as controls were enrolled from December 2012 to December 2013. CD4⁺ T cells were sorted by immunomagnetic separation. The global DNA methylation was tested with enzyme-linked immunosorbent assay (ELISA). The mRNA levels of DNA methylation-related regulating genes, DNA methyltransferases (DNMTs) and methylated CpG binding proteins (MBDs), were measured by real-time quantitative polymerase chain reaction (RT-PCR).

Results The level of global DNA methylation in peripheral blood CD4⁺ T cells of untreated IRP patients (3.525%±2.046%)and remission patients (4.790%±1.471%) were significantly lower than that of normal controls (10.101%±3.449%) respectively (both P<0.05). DNMT3b mRNA level of untreated IRP patients (1.332±0.785) was significantly lower than that of normal controls (2.077±1.059) in CD4⁺T cells (P<0.05). The mRNA expression of MBD2 was significantly higher in CD4⁺ T cells from untreated and remission IRP patients (2.999±1.601, 2.055±1.576) than that in controls (0.581±0.247) (both P<0.05). The MBD4 mRNA level was significantly higher in CD4⁺ T cells from untreated IRP patients (2.736±2.719) compared to that in normal controls (1.167±1.006) (p<0.05). DNMT3b mRNA expression and CD4⁺ T cell DNA methylation to be positive correlated within IRP patients (r=0.569, p<0.01). The MBD2 mRNA expression negatively correlated with CD4⁺ T cell DNA methylation and the ratio of Th1/Th2 (r=-0.763, p<0.001; r = -0.652, p<0.05). The global methylation of CD4⁺ T cells negatively related to the ratio of CD5⁺ B cells (r= -0.439, p<0.05).

Conclusions The globe DNA hypomethylation and abnormal expression of DNA methylation-related enzymes in peripheral blood CD4⁺ T cells may be related with the pathogenesis of IRP.