Background/Purpose

Recent studies have uncovered 5-hydroxymethylcytosine (5hmC) as the sixth base of the genome, and that the Ten-eleven translocation (TET) family proteins is responsible for the generation of 5hmC from 5mC in mammalian cells. 5hmC may function as another epigenetic mark by altering chromatin structure or contributing to the recruitment or exclusion of other DNA-binding proteins that affect transcription. However, the report about DNA hydroxymethylation in CD4+ T cell or SLE is poor, though DNA hypomethylation has been confirmed to contribute to SLE. In this study, we will investigate DNA hydroxymethylation in genome-wide in lupus CD4+ T cells.

Methods

36 SLE patients and 36 healthy controls were recruited. CD4+ T cells were isolated by magnetic beads. The age and sex are matched. All patients fulfilled at least 4 of the SLE classification criteria of ACR. The content of 5hmC in genome was detected by dot blot. hMeDIP-NimbleGen Human 3x720K Promoter Plus CpG Island Arrays was completed for identification of genes with different 5hmC modifications between SLE CD4+ T cells and normal controls. mRNA expression levels were measured by real-time RT-PCR. Gene ontology (GO) was analyzed by the Database for Annotation, Visualization and Integrated Discovery (DAVID). Student’s t-test for equality of means was used to compare values. P-values < 0.05 were considered as significant.

Results

Compared with normal controls, the content of 5hmC was increased significantly in lupus CD4+ T cells according to the result of dot blot. hMeDIP-chip and data analysis showed 2753 genes with increased 5hmC and 50 genes with decreased 5hmC within their promoter regions in lupus CD4+ T cells. Through the integrated analysis of DNA hydroxymethylation data with DNA methylation data from our previous study, we found 404 genes with increased 5hmC and decreased 5mC in promoter regions in lupus CD4+ T cells, such as UBE2A, IRF1, JUND. In addition, the integrated analysis of DNA hydroxymethylation data with gene expression data from our previous study showed 211 genes with increased promoter hydroxymethylation and expression in lupus CD4+ T cells, such as TNFRSF4, IL15RA and NR2F6, in which some significant GO enrichments, including T cell proliferation and cytokine-mediated signaling pathway, were observed. Furthermore, the results from real-time PCR showed that mRNA expression levels of TET2, TET3, UBE2A and JUND were up-regulated signficantly in CD4+T cells of SLE patients compared with normal controls. The positive correlation was observed between TET2 or TET3 and UBE2A or JUND expression levels.

Conclusion

The increased 5hmC may contribute to DNA demethylation and overexpression of some genes in lupus CD4+ T cells, which suggest DNA hydroxymethylation play an important role in the aberrant epigenetic mechanisms of SLE.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/dna-hydroxymethylation-changes-in-cd4t-cells-from-patients-with-systemic-lupus-erythematosus/