25 May 2015 : Laboratory Research
Down-Regulation of miR-101 Contributes to Rheumatic Heart Disease Through Up-Regulating TLR2
Hai DongBCDE, Yongmei SunADG, Feng ShanBCF, Qiang SunCDF, Bingxin YangDFDOI: 10.12659/MSM.893540
Med Sci Monit 2015; 21:1500-1506
Abstract
BACKGROUND: RHD is an autoimmune disease that arises following infection by S. pyogenes and imposes a heavy burden on public health.
MATERIAL AND METHODS: We detected 11 selected miRNAs expressed in the cardiac tissues of 11 RHD patients and 11 controls. By employing dual-luciferase assay and Western blot, we identified the relationship between TLR2 and miR-101 and miR-101. We used ELISA to test the concentration of TNF-α, IL-1β, and IL-6.
RESULTS: In cardiac tissue of RHD patients, miR-101 was significantly down-regulated (p=0.011). Ectopically expressed miR-101 repressed the luciferase activity by 27% through targeting TLR2 3’UTR. Combined with the results of Western blot, we confirmed that TLR2 is a direct target gene of miR-101. miR-101 knock-down is related to over-stimulated immune response in PGN-activated THP-1 cells. We detected a significantly higher concentration of TNF-α (p=0.0017), IL-1β (p=0.015), and IL-6 (p=0.014) in serum samples. TLR2 had a higher expression in patients in the protein level rather than the mRNA level, indicating that post-transcriptional regulation factors play a crucial role in regulating TLR2 expression.
CONCLUSIONS: The present study confirmed that miR-101 targets TLR2 3’UTR and represses TLR2 expression. This work also found an association between down-regulated miR-101 and rheumatic heart disease.
Keywords: Enzyme-Linked Immunosorbent Assay, Blotting, Western, Gene Expression Regulation - physiology, Interleukin-1beta - blood, Interleukin-6 - blood, Luciferases, Myocardium - metabolism, Rheumatic Heart Disease - etiology, Toll-Like Receptor 2 - metabolism, Tumor Necrosis Factor-alpha - blood
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