下调多巴胺受体D2表达促进Th1细胞分化可抑制乳腺癌EO771细胞移植瘤的生长

Down regulation of dopamine receptor D2 inhibits the growth of EO771 cell-based xenograft breast tumor through promoting Th1 cell differentiation

目的:探讨多巴胺受体D2(dopamine receptor D2,DRD2)在小鼠乳腺癌免疫微环境中的作用及可能的作用机制。方法:采用雌性C57BL/6J小鼠和鼠源性乳腺癌EO771细胞建立乳腺癌移植瘤模型。分别采用腹腔注射DRD2特异性拮抗剂甲硫哒嗪处理移植瘤小鼠(对照组为腹腔注射PBS),以及瘤内注射特异性针对DRD2基因的siRNA(siDRD2)处理移植瘤小鼠[对照组为瘤内注射与DRD2基因无相关性的阴性对照siRNA(siNC)]。采用免疫组织化学法检测肿瘤组织中Ki-67阳性细胞和CD4^(+)T细胞所占百分比,以及FCM法检测肿瘤组织中CD45^(+)CD4^(+)T细胞、Th1细胞(CD45^(+)CD4^(+)IFNγ^(+))和Th17细胞(CD45^(+)CD4^(+)IL-17^(+))所占百分比。结果:与对照组相比,甲硫哒嗪治疗组小鼠的肿瘤体积缩小(P<0.05),肿瘤质量降低(P<0.001),肿瘤组织中Ki-67阳性细胞所占百分比降低(P<0.001),CD45^(+)免疫细胞所占百分比和CD4^(+)T细胞所占百分比(P<0.05和P<0.001)增加,且Th1细胞(CD45^(+)CD4^(+)IFNγ^(+))所占百分比增加(P<0.001),Th17细胞(CD45^(+)CD4^(+)IL-17^(+))所占百分比的变化差异无统计学意义(P>0.05)。与对照组(siNC)相比,siDRD2组小鼠肿瘤体积减小(P<0.05),肿瘤质量降低(P<0.001),肿瘤组织中Ki-67阳性细胞所占百分比降低(P<0.001),肿瘤浸润CD4^(+)T细胞和Th1细胞所占百分比增加(P均<0.001),而Th17细胞所占百分比的变化差异无统计学意义(P>0.05)。结论:在小鼠乳腺癌模型中,下调DRD2表达可促进Th1细胞分化从而抑制乳腺癌细胞的增殖。

Objective:To investigate the role and mechanism of dopamine receptor D2(DRD2)in the immune microenvironment of breast cancer.Methods:The xenograft model of breast cancer is established using female C57BL/6J mice and murine breast cancer EO771 cells.Mice with transplanted tumor were treated by intraperitoneal injection of the specific antagonist of DRD2,thioridazine,while PBS was injected intraperitoneally in the control group.siDRD2 targeting DRD2 gene was intratumorally injected to tumor-bearing mice[negative control siRNA(siNC)was injected to the control group].The percentage of Ki-67 positive cells and CD4+T lymphocytes in tumor tissue was examined by immunohistochemical analysis.The proportion of CD45+CD4+T lymphocytes,CD45+CD4+IFNγ+Th1 cells and CD45+CD4+IL-17+Th17 cells was detected by flow cytometry.Results:Compared with the control group,the tumor volume(P<0.05),tumor mass(P<0.001)and the proportion of Ki-67 positive cells(P<0.001)in tumor tissue were decreased in the thioridazine-treated group.The proportion of CD45+immune cells(P<0.05)and CD4+T lymphocytes(P<0.001)in thioridazine-treated group was higher than that in the control group.The proportion of Th1 cells(CD45+CD4+IFNγ+)increased(P<0.001)in thioridazine-treated group.There was no significant changes in the proportion of Th17 cells(CD45+CD4+IL-17+)(P>0.05).Compared with the control group(siNC),the tumor volume(P<0.05),tumor mass(P<0.001)and the proportion of Ki-67 positive cells(P<0.001)were decreased in siDRD2 group.The proportion of tumor infiltrating CD4+T lymphocytes(P<0.001)and Th1 cells(P<0.001)in the siDRD2 group was higher than that in the siNC group,whereas the change in the proportion of Th17 cells was not statistically significant(P>0.05).Conclusion:Down-regulation of DRD2 could promote the differentiation of Th1 cells and inhibit the proliferation of breast cancer cells in a mouse breast cancer model.