AIM: The promoter of human interleukin-10 (IL10), 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 IL10 in rheumatoid arthritis (RA). METHODS: Bioinformatic analysis was conducted to identify the interspecies-conserved sequence in human, macaque and mouse IL10 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 μmol/L). The mRNA and protein levels of IL10 were examined using RT-PCR and ELISA, respectively. The methylation of CpGs in the IL10 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 IL10 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 IL10 mRNA and serum levels. In the 5-azacytidine-treated PBMCs, the CpG motifs were demethylated, and the expression levels of IL10 mRNA and protein was significantly increased. CHIP assays revealed increased phospho-CREB binding to the IL10 promoter. CONCLUSION: The methylation of the proximal CpGs in the IL10 promoter may regulate gene transcription in RA.
AIM: The promoter of humaninterleukin-10 (IL10), 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 IL10 in rheumatoid arthritis (RA). METHODS: Bioinformatic analysis was conducted to identify the interspecies-conserved sequence in human, macaque and mouseIL10 genes. Peripheral blood mononuclear cells (PBMCs) from 20 RApatients and 20 health controls were collected. The PBMCs from 6 patients were cultured in the presence or absence of 5-azacytidine (5 μmol/L). The mRNA and protein levels of IL10 were examined using RT-PCR and ELISA, respectively. The methylation of CpGs in the IL10 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 IL10 promoter. The upstream CpGs at -408, -387, -385, and -355 bp were hypermethylated in PBMCs from both the RApatients and healthy controls. In contrast, the proximal CpG at -145 was hypomethylated to much more extent in the RApatients than in the healthy controls (P=0.016), which was correlated with higher IL10 mRNA and serum levels. In the 5-azacytidine-treated PBMCs, the CpG motifs were demethylated, and the expression levels of IL10 mRNA and protein was significantly increased. CHIP assays revealed increased phospho-CREB binding to the IL10 promoter. CONCLUSION: The methylation of the proximal CpGs in the IL10 promoter may regulate gene transcription in RA.
Authors: José Miguel Sempere-Ortells; Vicente Pérez-García; Gema Marín-Alberca; Alejandra Peris-Pertusa; José Miguel Benito; Francisco Manuel Marco; José Jacobo Zubcoff; Francisco Javier Navarro-Blasco Journal: Autoimmunity Date: 2009 Impact factor: 2.815
Authors: J P Issa; P M Vertino; J Wu; S Sazawal; P Celano; B D Nelkin; S R Hamilton; S B Baylin Journal: J Natl Cancer Inst Date: 1993-08-04 Impact factor: 13.506
Authors: John R Glossop; Richard D Emes; Nicola B Nixon; Kim E Haworth; Jon C Packham; Peter T Dawes; Anthony A Fryer; Derek L Mattey; William E Farrell Journal: Epigenetics Date: 2014-07-07 Impact factor: 4.528