胃癌源性外泌体对HLA-DR^(neg)leukocytes中Dicer1、PTEN基因的调控

The regulatory effects of gastric cancer-secreted exosomes on Dicer1 and PTEN genes in HLA-DR^(neg)leukocytes

目的通过观察胃癌源性的外泌体对HLA-DR^(^(neg))leukocytes中Dicer1、PTEN基因表达的影响,探讨肿瘤源性外泌体调控机体免疫抑制功能的机制。方法提取胃癌患者血清外泌体并鉴定,免疫磁珠法分选HLA-DR^(neg)leukocytes,吖啶橙染色后的外泌体与CFSE染色后的HLA-DR^(neg)leukocytes在37℃,5%CO_2条件下共培养12 h;采用QRT-PCR和Western blot方法分别检测共培养后胃癌患者与健康对照组中Dicer1、PTEN基因和蛋白的表达。结果成功分离胃癌患者外周血外泌体;同时观察到外泌体被HLA-DR^(neg)leukocytes有效摄取。QRT-PCR检测结果显示Dicer1 m RNA、PTEN m RNA的相对表达量分别是健康对照组的0.46±0.19、0.48±0.27倍(P<0.05);Western blot结果显示Dicer1、PTEN蛋白表达量是健康对照组的0.15±0.11、0.33±0.19倍(P<0.05)。结论胃癌来源的外泌体可以抑制HLA-DR^(neg)leukocytes中Dicer1、PTEN基因的表达,为今后进一步研究肿瘤来源的外泌体的免疫调控作用提供线索。

This study aimed to investigate the regulatory effects of gastric cancer-secreted exosomes on Dicer1 and PTEN genes in HLA-DR^neg leukocytes, and explore the function mechanism of tumor derived exosomes in promoting immunosuppression. Exosomes from the serum of gastric cancer patients were extracted and identified;HLA-DR^neg leukocytes were sorted by paramagnetic particle method, then the exosomes acridine orange and the HLA-DR^neg leukocytes stained with CFSE were co-cultured for 12 hours at 37 ℃ and in condition of 5% CO2.QRT-PCR and Western blot were used to detect the protein and m RNA expression of Dicer1 and PTEN in gastric cancer group and healthy control. Data showed that the peripheral blood exosomes were successful purified, and after 12 h co-culture, fluorescence microscope illustrated that exosomes were absorbed by HLA-DR^neg leukocytes significantly. QRT-PCR and Western blot indicated that the m RNA and protein expression of Dicer1 and PTEN in cancer group were inferior to healthy controls（P〈0.05）. In conclusion, gastric cancer-secreted exosomes can inhibit the expression of Dicer1 and PTEN genes in gastric cancer HLA-DR^neg leukocytes, which provide clues for further study on immune regulation of cancer-secreted exosomes.