RNA interference suppression MyD88 expression on murine bone marrow dendritic cells influence the biological activity of.doc

RNA interference suppression MyD88 expression on murine bone marrow dendritic cells influence the biological activity of Author: Chen Jun, Zong-Quan, Dong Nian States, Su Gang, Liu Chao, Liu Jinping, Deng Zhi [Keywords:] dendritic cell;, MyD88;, RNA interference;, lipopolysaccharide Inhibitory effects of RNA interference on MyD88 expression and biological activity in murine myeloid dendritic cells [Abstract] AIM: To investigate inhibitory effects of RNA interference on MyD88 expression in murine myeloid dendritic cells (DCs) and detect the biological activity of DCs. METHODS: Three pairs of myeloid differentiation factor 88 (MyD88) siRNA were synthesized and transfected into DCs by RNAimate. The mRNA and protein expression of MyD88 were analyzed by semiquantified RTPCR and Western blot. Mouse DCs were divided into control group and RNA interference group. One of the highest effective siRNA was transfected into RNA interference group. 12 hours later, LPS of the final concentrations of 10 mg / L was added in two groups and continued to culture for 3 days. The phenotype and functional properties of DCs were detected by flow cytometry and mixed lymphocyte reaction (MLR). The concentration of TNFα , IFNγ and IL12 in the supernatant was detected by ELISA, and the concentration of NFκ B was detected by immunochemistry. RESULTS: mRNA and protein expression were reduced 90% and 85% in sequence2 siRNA, 92% and 88% in sequence3 siRNA respectively but no change was found in other groups. LPS stimulation increased the expression of CD80, CD86 and MHC Ⅱ in the cytomembrane of DCs, the concentration of TNFα , IFNγ and IL12 in the supernatant in control group. Besides, LPS stimulation promoted the shift of NFκ B to karyon and the proliferation of allogeneic T cells in control group. RNA interference inhibited these effects induced by LPS. CONCLUSION: RNA interference can reduce MyD88 expression in murine myeloid dendritic cells and inhibit the maturation of DCs, which may pr