MiR-223 Regulates TNF- α Induced FDC - SP Gene Expression in Human Gingival Epithelial Cells

Follicular dendritic cell-secreted protein（FDC-SP）is expressed in tonsils, follicular dendritic cells, junctional epithelium and periodontal ligament. Functions of microRNAs （miRNAs）are post-transcriptional gene regulation and RNA silencing through binding to the 3’-untranslated region（3’-UTR）of mRNA. To analyze the effects of miR-223 on the expression of FDC-SP with or without stimulation by TNF-α in human gingival epithelial Ca9-22 cells, we conducted real-time PCR, luciferase（LUC）assays using -948FDC-SP+3’-UTR constructs that were inserted by two kinds of miR-223 binding sites of FDC-SP 3’-UTR in the downstream of the LUC gene, and Western blotting. Stimulation of Ca9-22 cells with TNF-α increased miR-223 expression, and transfection of Ca9-22 cells with a miR-223 expression plasmid also increased miR-223 expression. FDC-SP mRNA and protein levels increased by TNF-α were inhibited by overexpression of miR-223, and FDC-SP mRNA level was further increased by miR-223 inhibitor. Inhibitor of κB kinase α（IKKα）and mitogen-activated protein kinase phosphatase 5（MKP-5）mRNA and protein levels were increased by TNF-α, and then decreased by mir-223 overexpression. LUC activities of -948FDC-SP+exon 4-5 and -948FDC-SP+1-984 were increased by TNF-α and almost completely inhibited by transfection with a miR-223 expression plasmid. U0126（MEK1/2 inhibitor）, BMS345541（IKKα and IKKβ inhibitor）and IMD0354（IKKβ inhibitor）inhibited TNF-α induced LUC activities of - 948FDC - SP+exon 4 - 5. These studies demonstrated that miR–223 suppresses TNF-α- induced FDC-SP expression by targeting to FDC-SP, IKKα and MKP5 in human gingival epithelial cells.