树突状细胞与马尔尼菲青霉酵母体外相互作用的研究

The Interaction Between Dendritic Cells and Penicillium marneffei Yeasts

本文探讨了树突状细胞(DCs)在抗马尔尼菲青霉感染免疫中的作用。用细胞因子rhGM-CSF和rhIL-4诱导人外周血单核细胞分化为树突状细胞,观察DCs的形态,并用流式细胞仪进行DCs的表型测定,ELISA方法检测培养上清液IL-12p70的浓度,混合淋巴细胞反应检测DCs刺激T淋巴细胞的增殖能力,实时荧光定量PCR检测趋化因子受体CCR7、CXCR4的mRNA的表达。倒置显微镜下可见诱导获得的DCs细胞形态不规则,表面伸展出大量树突。与马尔尼菲青霉酵母共同培养24h后DCs的胞内含有大量的酵母细胞;细胞表型CD86、CD83、HLA-DR和CD40的表达明显增高;刺激T淋巴细胞增殖的能力增强;趋化因子受体CCR7和CXCR4的mRNA表达量增加且能够产生IL-12p70但产生的量低于LPS刺激组。DCs能吞噬加热灭活的马尔尼菲青霉酵母,并趋于成熟,抗原呈递能力增加,但是产生IL-12p70的量较低,可能造成宿主抗马尔尼菲青霉酵母的细胞免疫功能的不足。

To assay the influence of dendritic cells （DCs） on the function of anti-infective immunity to Penicillium marneffei. DCs were generated from peripheral blood mononuclear cells （PBMC） and pulsed with Penicillium marneffei yeasts. DCs morphology was observed by the inverted microscope and cell surface markers of DCs were analyzed by flow cytometry. The concentrations of IL-12p70 were detected by ELISA. Mixed lymphocyte reaction was performed to assay the proliferation of T cells. The mRNA of CCR7 and CXCR4 were detected by the Real-time PCR quantifications. The acquired DCs exhibited irregular appearance and numerous long dendrites under light microscope. DCs and Penicillium marneffei yeasts were co-cultured for 24 h, numerous yeasts were observed inside the cells; an enhanced expression of the cell surface markers CD86, CD83, HLA-DR and CD40 were demonstrated; the expression of CCR7 and CXCR4 mRNA were also increased; the improved proliferation of T cells were observed in the mixed lymphocyte reaction. Yeasts-pulsed DCs secreted more IL-12p70 than that of non-pulsed, but less than that of LPS-pulsed DCs. DCs can engulf the Penicilliurn rnarneffei yeasts. When pulsed with Penicillium rnarneffei yeasts, DCs improved their expression of the co-stimulatory molecules and chemokine receptor CCR7, CXCR4, enhanced their capacity to process antigen. DCs play an important role in host defense against Penicillium marneffei infection. But the low level of the IL-12p70 production may lead to deficiency in the cell-mediated immunity against Penicillium marneffei.