Hypomethylation of proximal CpG motif of interleukin-lO promoter regulates its expression in human rheumatoid arthritis

• Published in: 中国药理学报：英文版 Vol. 32; no. 11; pp. 1373 - 1380
• Main Author: Li-hong FU Chun-ling MA Bin CONG Shu-jin LI Hai-ying CHEN Jing-ge ZHANG
• Format: Journal Article
• Language: English
• Published: 2011
• Subjects: CpG基序; 人白细胞介素; 去甲基化; 启动子; 外周血单个核细胞; 外周血单核细胞; 染色质免疫沉淀; 类风湿关节炎
• ISSN: 1671-4083; 1745-7254

Aim： The promoter of human interleukin-lO （ILIO）, a cytokine crucial for suppressing inflammation and regulating immune responses, contains an interspecies-conserved sequence with CpG motifs. The aim of this study was to investigate whether methylation of CpG motifs could regulate the expression of ILIO in rheumatoid arthritis （RA）. Methods： Bioinformatic analysis was conducted to identify the interspecies-conserved sequence in human, macaque and mouse ILIO genes. Peripheral blood mononuclear cells （PBMCs） from 20 RA patients and 20 health controls were collected. The PBMCs from 6 patients were cultured in the presence or absence of 5-azacytidine （5 pmol/L）. The mRNA and protein levels of ILIO were examined using RT-PCR and ELISA, respectively. The methylation of CpGs in the ILIO promoter was determined by pyrosequencing. Chromatin immunoprecipitation （CHIP） assays were performed to detect the cyclic AMP response element-binding protein （CREB）-DNA interactions. Results： One interspecies-conserved sequence was found within the ILIO promoter. The upstream CpGs at -408, -387, -385, and -355 bp were hypermethylated in PBMCs from both the RA patients and healthy controls. In contrast, the proximal CpG at -145 was hypomethylated to much more extent in the RA patients than in the healthy controls （P=0.016）, which was correlated with higher ILIO mRNA and serum levels. In the 5-azacytidine-treated PBMCs, the CpG motifs were demethylated, and the expression levels of ILIO mRNA and protein was significantly increased. CHIP assays revealed increased phospho-CREB binding to the ILIO promoter. Conclusion： The methylation of the proximal CpGs in the ILIO promoter may regulate gene transcription in RA.