ABSTRACT
Objective To investigate the regulatory effects of miR-146a on inflammation factors production in C ryptococcus neoformans treated THP-1 cells.Methods Cryptcoo ccus neoformans strains were heat killed .Fluorescence quantitative RTP-CR and ELISA were used to detect the levels of IL -1βand TNF-αin THP-1 cells treated with heat-killed Cryptococcus neoformans.In addition, the production of IL-1βand TNF-αwere analyzed before and after Dectin-1or TLR 4 blocked .THP-1 cell lines that were respectively transfected with miR-146 a mimics and inhibitors were constructed and the production of IL-1βand TNF-αin these cell lines were analyzed after interfered with Cryptococcus neoformans.Results With the interference of heat-killed Cryptococcus neoformans, the expression of miR-146a was up-regulated in THP-1 cells, but was down-regulated after Dectin-1 or TLR4 blocked.The expressions of IL-1βand TNF-αinduced by heat-killed Cryptococcus neoformans were enhanced in miR-146a mimics transfected THP-1 cells, but was inhibi-ted in inhibitors transfected THP-1 cells.Conclusion Heat-killed Cryptococcus neoformans could up-regu-late miR-146a expression in THP-1 cells via Dectin-1 and TLR.miR-146a could negatively regulate the ex-pressions of IL-1βand TNF-αinduced by Cryptococcus neoformans.
ABSTRACT
Objective To study the mechanism of mesenchymal stem cells immunosupression lym- phocyte proliferation via B7-H1/PD-1 pathway upregnlated by IFN-γ. Methods Bone marrow derived mes-enchymal stem cells (MSC) were isolated and purified by repeat adherent passage and detected them multi-potential differentiation in conditioned culture medium. Then MSC were cocuhured with lymphocyte prolifera-tion and assayed the level using γ H-thymidine incorporation. Meanwhile, ELISA measured IFN-γ, TGF-β, TNF-α and IL-10 in the cocuhured supernaatant and analyzed variation of B7-H1 molecular profile in MSC by flow cytometry. At last siRNA technology was deploied to interfere B7-H1 expression and analyzed MSC im-munosuppression on lymphocyte proliferation. Results In vitro the isolated MSC become homogeneous spi-die-shaped adherent cells after five passages, and in conditioned culture medium they could differentiate into adipocytes, osteocytes and chondrocytes. In the eocuhure of MSC with mixed lymphocyte, lymphocyte prolif- eration stimulated by Con A or by anti-CD3/CD28 antibody. The cpm value of the proliferation detected by 3H thymidine incorporation showed MSC suppressed the proliferation significantly (P = 0. 0167, 0. 0081,<0.0001 ) and the suppressive potential in a dose-dependent fashion. In the coeuhured supernatant cyto-kine IFN-T and TNF-α were detected in high concentration, but TGF-β, IL-10 were undetected. Simultane- ously MSC in the coeuhure upregulated B7-H1 expression from basic expression 7% to higher than 70% ( P < 0.05 ). After interfere B7-H1 expression in MSC by specific siRNA, we detected lymphocyte proliferation and got higher cpm by 3H thymidine incorporation ( P < 0. 05 ). Conclusion MSC upregulated B7-H1 mo-lecular expression upon the stimuli of IFN-γ, and through the B7-H1/PD-1 pathway mediated immunosu-pression on lymphocyte proliferation.