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目的:探讨带有甘露糖敏感血凝菌毛的铜绿假单胞菌(Pseudomonas aeruginosawith mannose sensitive hemagglutina-tion pili,PA-MSHA)疫苗处理后急性髓细胞白血病源性树突状细胞(dendritic cells derived from acute myeloid leukemia,AML-DC)对调节性T细胞(regulatory Tcell,Treg)的抑制作用。方法:rhGM-CSF和rhII-4诱导的AML-DC分为对照组、PA-MSHA组和TNF-α组,培养24h后观察3组AML-DC的形态、流式细胞术检测各组AML-DC的表型、MTT法和混合淋巴细胞反应检测AML-DC对淋巴细胞增殖的作用。磁珠法分离健康人外周血CD4+T细胞,加入各组AML-DC中诱导Treg,ELISA法检测各组Treg上清液中IL-10、TGF-β的水平,流式细胞术检测Treg表面CD4、CD25的表达,RT-PCR法检测Treg中Foxp3mRNA的表达水平。结果:PA-MSHA组和TNF-α组AML-DC呈树突状形态,且CDla、CD80、CD83、CD86和HLA-DR表达较对照组明显升高(P<0.05)。PA-MSHA组和TNF-α组AML-DC诱导的T细胞增殖能力显著增强(P<0.05)。PA-MSHA组和TNF-α组AML-DC诱导产生的Treg分泌较低水平的IL-10、TGF-β(P<0.05),CD4、CD25的表达及Foxp3mRNA水平均较对照组明显降低(P<0.05)。上述各指标PA-MSHA组和TNF-α组间均无明显差异。结论:PA-MSHA疫苗可促进AML-DC的成熟,抑制初始T细胞向Treg的分化,增强AML-DC对AML患者Treg的抑制作用。
AIM: To investigate the effect of dendritic cells derived from Pseudomonas aeruginosawith mannose sensitive hemagglutinin pili (PA-MSHA) vaccine from mannose- acute myeloid leukemia (AML-DC) on regulatory T cells (Tregs). Methods: AML-DCs induced by rhGM-CSF and rhII-4 were divided into control group, PA-MSHA group and TNF-α group. The morphology of AML- DC phenotype, MTT assay and mixed lymphocyte reaction to detect the effect of AML-DC on lymphocyte proliferation. Tregs were isolated from AML-DCs of each group by magnetic beads method. The levels of IL-10 and TGF-β in Treg supernatants of each group were detected by ELISA. Flow cytometry was used to detect Treg surface CD4, CD25 expression, Treg Foxp3 mRNA expression levels detected by RT-PCR. Results: The dendritic morphology of AML-DC in PA-MSHA group and TNF-α group was significantly higher than that in control group (P <0.05). The proliferation of T cells induced by AML-DC in PA-MSHA group and TNF-α group was significantly enhanced (P <0.05). Treg secreted by AML-DC induced by PA-MSHA and TNF-α had lower levels of IL-10 and TGF-β (P <0.05), and lower expressions of CD4 and CD25 and Foxp3 mRNA <0.05). There was no significant difference between the above indexes of PA-MSHA and TNF-α groups. Conclusion: PA-MSHA vaccine can promote the maturation of AML-DC, inhibit the differentiation of naive T cells into Tregs and enhance the inhibitory effect of AML-DC on Tregs in AML patients.