摘要
目的探讨沉默结肠癌PARG基因后对肿瘤局部免疫状态的影响。方法以PARG-shRNA慢病毒载体转染CT26细胞为实验组,未转染CT26细胞和空载体转染CT26细胞为对照组,分别将各组细胞在BALB/C小鼠脾包膜下接种形成肝转移模型;Western blot法检测脾脏移植瘤中PARG蛋白的表达;免疫荧光双标法检测脾脏中B220+DEC205+DC和CD11c+CD11b+DC,用激光共聚焦显微镜观察上述细胞;免疫荧光单标记法检测脾脏中CD4+T细胞及CD8+T细胞,用激光共聚焦显微镜观察上述细胞;ELISA法检测各组小鼠血清中IL-10和IL-12的表达。结果PARG基因沉默后,脾脏移植瘤中PARG蛋白的表达量明显较对照组降低(P<0.05);脾脏组织中B220+DEC205+DC较对照组显著减少;而CD11c+CD11b+DC较对照组明显增多(P<0.05);脾脏组织中的CD4+T细胞及CD4+/CD8+比值较对照组明显升高(P<0.05);血清中IL-10的表达较对照组明显减少而IL-12的表达较对照组显著增多(P<0.05)。结论沉默结肠癌PARG基因可通过影响DC及T细胞的增殖分化,增强肿瘤局部的免疫反应。
Objective To investigate the effect of silencing PARG on local immune status in colon carcinoma. Methods Lentivirus PARG-shRNA（ Short hairpin RNA） was transfected into CT26 cells as experiment group, the CT26 cells without any treatment or treated with Lentivirus empty vector served as control group. Animal models for liver metastases of colon carcinoma were established by splenic subcapsular inoculation of each group of CT26 cells in Balb/c mice. The expression of PARG protein in spleen transplant tumor was detected by Western blot analysis. Immunofluorescence double labeling assay was used for the numbers detection both of B220 ＋ DEC205 ＋ DC and CDll e ＋ CDll b ＋ DC in the spleen. CD4 ＋ T cells and CD8 ＋ T cells in the spleen were tested by immunofluorescence single staining. All of these cells were observed by confoeal laser scanning microscope. The levels of IL-10 and IL-12 in serum were measured by enzyme-linked immunosorbent assay （ELISA）. Results The expression of PARG in spleen transplant tumor was reduced in PARG-silenced group compared to that in control groups （P 〈 0. 05）. In PARG-sileneed group, the amounts of B220 ＋ DEC205 ＋ DC were less and the numbers of CDllc ＋ CD11 b ＋ DC were more in spleen than that in control groups（ P 〈 0. 05 ）. The numbers of CD4 ＋T cells and the ratio of CD4 ＋/CD8 ＋ in spleen of PARG-silenced group were increased compared to that in control groups（P 〈0. 05）. The serum levels of IL-10 were lower but the serum content of IL-12 were higher in PARG-silenced group than thatin control groups（P 〈 0. 05）. Conclusions Silencing PARG could strengthen the local immune response to colon cancer by effect on the proliferation and differentiation of DC and T cells.
出处
《基础医学与临床》
CSCD
北大核心
2014年第3期391-396,共6页
Basic and Clinical Medicine
