OBJECTIVE: To study the effect of DNMT1 on CD4(+) T cells in the peripheral blood of systemic lupus erythematosus (SLE) patients. METHODS: To investigate the differential expression of DNMT1 in CD4(+) T cells of SLE patients and healthy individuals, a DNMT1 lentiviral plasmid (pLenti6.3/V5-DNMT1) and a control plasmid (pLenti6.3/V5-GW/LacZ) were constructed and transfected into CD4(+) T cells from the peripheral blood of SLE patients. The transcriptional and translational expression of DNMT1, global genomic DNA methylation, and the production of IgG antibody in the CD4(+) T cells in the peripheral blood of SLE patients were assessed using qPCR analysis, western blotting, flow cytometry, and ELISA, respectively. RESULTS: The expression level of DNMT1 in SLE patients was significantly lower than that in normal humans. The expression of DNMT1 was found to be positively correlated with the methylation level of genomic DNA and negatively correlated with the IgG titration level. DNA sequencing results confirmed that the DNMT1 lentiviral plasmid was successfully constructed. After the CD4(+) T cells from the peripheral blood of SLE patients were transfected with the pLenti6.3/V5-DNMT1 plasmid, the transcription level of the DNMT1 gene was upregulated and abundance of DNMT1 protein significantly increased. Global genomic DNA methylation was enhanced, while the production of IgG antibody was reduced. CONCLUSION: DNMT1 can inhibit the autoimmune response in SLE patients by reversing the abnormally low DNA methylation level in the CD4(+) T cells.
OBJECTIVE: To study the effect of DNMT1 on CD4(+) T cells in the peripheral blood of systemic lupus erythematosus (SLE) patients. METHODS: To investigate the differential expression of DNMT1 in CD4(+) T cells of SLEpatients and healthy individuals, a DNMT1 lentiviral plasmid (pLenti6.3/V5-DNMT1) and a control plasmid (pLenti6.3/V5-GW/LacZ) were constructed and transfected into CD4(+) T cells from the peripheral blood of SLEpatients. The transcriptional and translational expression of DNMT1, global genomic DNA methylation, and the production of IgG antibody in the CD4(+) T cells in the peripheral blood of SLEpatients were assessed using qPCR analysis, western blotting, flow cytometry, and ELISA, respectively. RESULTS: The expression level of DNMT1 in SLEpatients was significantly lower than that in normal humans. The expression of DNMT1 was found to be positively correlated with the methylation level of genomic DNA and negatively correlated with the IgG titration level. DNA sequencing results confirmed that the DNMT1 lentiviral plasmid was successfully constructed. After the CD4(+) T cells from the peripheral blood of SLEpatients were transfected with the pLenti6.3/V5-DNMT1 plasmid, the transcription level of the DNMT1 gene was upregulated and abundance of DNMT1 protein significantly increased. Global genomic DNA methylation was enhanced, while the production of IgG antibody was reduced. CONCLUSION:DNMT1 can inhibit the autoimmune response in SLEpatients by reversing the abnormally low DNA methylation level in the CD4(+) T cells.
Entities:
Keywords:
CD4+ T cells; Lentivirus; SLE; methylation; pLenti6.3/V5-DNMT1
Authors: B C Richardson; J R Strahler; T S Pivirotto; J Quddus; G E Bayliss; L A Gross; K S O'Rourke; D Powers; S M Hanash; M A Johnson Journal: Arthritis Rheum Date: 1992-06