B7-H1通过调节肿瘤免疫微环境促进AML肿瘤细胞免疫逃逸的实验研究

B7-H1 promotes immune escape of AML tumor cells by regulating tumor immune microenvironment

目的探究B7-H1在介导急性髓性白血病(AML)肿瘤免疫逃逸中的作用及机制。方法培养M1~M7型AML细胞,采用乳酸脱氢酶(LDH)释放法检测NK细胞对AML细胞的杀伤活性,ELISA法检测IL-2、IFN-γ的含量。通过皮下和尾静脉注射C1498-GFP细胞构建小鼠AML皮下和转移模型,设对照组、IgG抗体组及抗-B7-H1抗体组。对于皮下模型,记录小鼠肿瘤体积变化,第27天取出肿瘤并称重,检测肿瘤中T细胞数量及IFN-γ和TNF-α水平。对于转移模型,每天记录小鼠的体质量,当体质量减少>10%时记为死亡,检测全血、肝、脾和骨髓中AML细胞数量和肝组织中CD4^+T和CD8^+T细胞的水平。结果 NK细胞对M5型AML细胞的杀伤率最低(P<0. 05)。施加抗-B7-H1抗体后,NK细胞对M1、M2、M5、M6型细胞的杀伤率明显增加(P<0. 05),共培养上清液中IL-2和IFN-γ的水平亦升高。在皮下AML模型中,抗-B7-H1抗体组小鼠第27天的肿瘤体积和瘤体质量均显著低于对照组和IgG组(P<0. 01),Treg细胞的数目明显减少,效应T细胞的数目和免疫因子的分泌水平显著提高(P<0. 01)。在转移性AML模型中,抗-B7-H1抗体组小鼠的血液、肝、脾和骨髓中AML细胞含量明显低于IgG组和对照组(P<0. 01),而肝组织中CD4+T和CD8+T细胞的数目明显高于IgG组和对照组(P<0. 01)。结论 B7-H1通过增加肿瘤中抑制性T细胞、减少效应T细胞的数量和降低细胞免疫因子的水平来促进AML肿瘤的进展和免疫逃逸。施加抗-B7-H1抗体可以恢复机体的免疫水平。

Objective To explore the role and mechanism of B7-H1 in tumor immune escape of acute myeloid leukemia(AML).Methods M1-M7 AML cells were cultured.LDH release method was used to detect the killing activity of NK cells on AML cells.The contents of IL-2 and IFN-γwere determined by ELISA kit.Subcutaneous and metastatic models of mouse AML were constructed by subcutaneous and C1498-GFP injection.The control group,IgG antibody group and anti-B7-H1 antibody group were set.For the subcutaneous model,the tumor volume changes of the mice were recorded.The tumor was taken out and weighed on the 27th day,and the number of T cells in the tumor as well as IFN-γand TNF-αlevels were detected.For the metastasis model,the weight of mice was recorded every day.When the weight loss was more than 10%,it was recorded as death.The number of AML cells in the whole blood,liver,spleen and bone marrow and the level of CD4+T and CD8+T cells in liver tissue were measured.Results The killing rate of NK cells against M5 type AML cells was the lowest(P<0.05).After the application of anti-B7-H1 antibody,the killing rate of NK cells against M1,M2,M5 and M6 cells was significantly increased(P<0.05),and the levels of IL-2 and IFN-γin the co-culture supernatant were also increased.In the subcutaneous AML model,the tumor volume and tumor mass of the anti-B7-H1 antibody group on day 27 were significantly lower than those of the control group and IgG group(P<0.01),the number of Treg cells was significantly reduced,and the number of effector T cells and the secretion level of immune factors were significantly increased(P<0.01).In the metastatic AML model,the amount of AML cells in blood,liver,spleen and bone marrow of mice in the anti-B7-H1 antibody group was significantly lower than that in the IgG group and the control group(P<0.01),while the number of CD4+T and CD8+T cells in the liver tissue was significantly higher than that in the IgG group and the control group(P<0.01).Conclusion B7-H1 promotes AML tumor progression and immune escape by increasing inhibitory T cells in tumors,reducing the number of effector T cells,and reducing the level of cellular immune factors.The application of anti-B7-H1 antibody can restore the body s immune level.