马尔尼菲青霉对巨噬细胞模式识别受体TLR-2、TLR-4、Dectin-1的表达及TNF-α分泌的影响

Effect of Penicillium marneffei on TLR-2,TLR-4,and Dectin-1 expression and TNF-α production in macrophages

目的研究马尔尼菲青霉对巨噬细胞模式识别受体TLR-2、TLR-4、Dectin-1的表达及促炎因子TNF-α分泌的影响。方法马尔尼菲青霉酵母相菌液与小鼠腹腔巨噬细胞共培养24h,采用流式细胞技术检测巨噬细胞TLR-2、TLR-4及Dectin-1的平均荧光强度;共聚焦显微镜观察荧光染色的受体;ELISA法测定培养液上清中TNF-α的浓度;Real time PCR检测不同时间段TNF-α的mRNA表达。结果马尔尼菲青霉可使巨噬细胞TLR-2、TLR-4、Dectin-1的平均荧光强度均增高,并激活巨噬细胞产生TNF-α。结论马尔尼菲青霉上调了巨噬细胞模式识别受体TLR-2、TLR-4及Dectin-1的表达,巨噬细胞的激活与TLR-2、TLR-4及Dectin-1的表达上调相关。

Objective To study the effects of heat-killed Penieillium marneffei （PM） on the expressions of toll-like receptor-4 （TLR-4）, toll-like receptor-2 （TLR-2） and dendritic cell associated C-type lectin-1 （Dectin-1）and the production of the proinflammatory cytokine tumor necrosis factor-α （TNF-α. in mouse peritoneal macrophages. Methods Mouse peritoneal macrophages were cultured in the presence of heat-killed yeast-phase PM for 24 h, and the average fluorescence intensity of TLR-2, TLR-4, and Dectin-1 in the macrophages was detected using flow cytometry. Fluorescent staining of the macrophages was performed to observe the fluorescence of TLR-2, TLR-4, and Dectin-1 with confocal microscopy. TNF-α mRNA in the cell culture supernatant was measured with real-time PCR, and TNF-α protein detected using enzyme-linked immunosorbent assay （ELISA）. Results The average fluorescence intensity of TLR-2, TLR-4 and Dectin-1 in the macrophages was increased in response to a 24-h PM stimulation, and the stimulated macrophages produced large amounts of TNF-α. Conclusion PM up-regulates the expression of TLR-2, TLR-4 and Dectin-1 in mouse peritoneal macrophages, and their expressions are directly associated with macrophage activation.