TAMs激活Src-RhoA通路上调免疫抑制因子表达及促进直肠癌细胞增殖和EMT  

TAMs activate Src-RhoA pathway to up-regulate immunosuppressive factor expression and promote the proliferation and EMT of rectal cancer

作　　者：张静[1] 胡哲[1] 居红格[2] ZHANG Jing;HU Zhe;JU Hongge(Department of Basic Medicine,Vocational and Technical College of Baotou Medical College,Inner Mongolia University of Science and Technology,Baotou 014030,China;Department of Pathology,Baotou Medical College,Inner Mongolia University of Science and Technology,Baotou 014101,China)

机构地区：[1]内蒙古科技大学包头医学院职业技术学院医学基础科,014030 [2]内蒙古科技大学包头医学院病理教研室,014010

出　　处：《免疫学杂志》2021年第7期596-603,共8页Immunological Journal

基　　金：内蒙古自治区自然科学一般项目(NJZY16198)。

摘　　要：目的探讨肿瘤相关巨噬细胞(tumor-associated macrophages,TAMs)对直肠癌增殖和EMT的影响及其作用机制。方法用佛波酯(PMA)及白介素4(interleukin 4,IL-4)诱导人单核巨噬细胞(THP1),使其分化为M2型TAMs,采用流式细胞仪检测CD163和CD16的阳性表达率;TAMs与SW837细胞共培养48 h,通过CCK-8法检测SW837细胞活力,流式细胞仪检测细胞凋亡率,RT-qPCR检测IL-10和TGF-β1的表达量,Western blot分析E-cadherin、N-cadherin、IL-10、TGF-β1蛋白的表达以及Src、RhoA蛋白的磷酸化水平;Src通路抑制剂SU6656作用细胞后,分别用CCK-8和流式细胞仪检测SW837细胞活力和细胞总死亡率,Western blot检测细胞中IL-10、TGF-β1、E-cadherin和N-cadherin的表达量。结果THP1分化诱导为TAMs后,CD163的阳性表达率明显高于CD16的阳性表达率(P<0.01);和SW837、SW837+THP1+PMA、SW837+THP1+IL-4细胞相比,SW837与TAMs共培养后使SW837细胞活力明显升高(P<0.05),SW837细胞总死亡率明显降低(P<0.01),N-cadherin表达量明显升高,Ecadherin表达量明显降低,IL-10和TGF-β1蛋白表达量明显升高;TAMs激活直肠癌SW837细胞中Src-RhoA通路,使Src、RhoA蛋白的磷酸化水平明显升高;和SW837+TAMs细胞相比,SW837+TAMs+SU6656细胞活力明显降低,细胞总死亡率明显升高,E-cadherin蛋白表达量明显升高,N-cadherin蛋白表达量明显降低,IL-10和TGF-β1蛋白表达量明显降低。结论肿瘤相关巨噬细胞通过激活Src-RhoA通路产生免疫抑制因子、促进了直肠癌的增殖和EMT。This study was designed to investigate the effect of tumor-related macrophages(TAMs)on the proliferation and EMT of rectal cancer and its mechanism.PMA and IL-4 were used to induce human mononuclear macrophage line THP1 differentiation into TAMs,and the frequency of CD163 and CD16 were detected by flow cytometry.TAMs and SW837 cells were co-cultured for 48 h,then the cell viability was detected by CCK-8 method,the mortality rate of SW837 cells was detected by flow cytometry,and the expression levels of IL-10 and TGF-β1 were detected by RT-qPCR.Western blot was used to detect the expression levels of E-cadherin,N-cadherin,IL-10,TGF-β1 and the phosphorylation level of Src and RhoA.After the cells were treated with SU6656,CCK-8 and flow cytometry were used to detected the cell viability and the mortality rate of SW837 cells respectively,while Western blot was used to detect the expression levels of IL-10,TGF-β1,E-cadherin and N-cadherin.Data showed that after THP1 differentiation into TAMs,the frequency of CD163 was significantly higher than that of CD16(P<0.01);Compared with SW837,SW837+THP1+PMA,SW837+THP1+IL-4 cells,co-culture of SW837 and TAMs significantly increased the expression of N-cadherin,the protein levels of IL-10 and TGF-β1,and theactivity of SW837 cells(P<0.05),while significantly decreased the mortality rate of SW837 cells(P<0.01),theexpression of E-cadherin.TAMs activated the Src-Rho A pathway in SW837 cells,and promotes thephosphorylation level of Src and Rho A significantly.Compared with SW837+TAMs,the cell viability of SW837+TAMs+SU6656 was significantly reduced,the mortality rate was significantly increased,the expression of E-cadherin protein was significantly increased,the expression of N-cadherin protein was significantly decreased,andthe expression of IL-10 and TGF-β1 protein was significantly decreased.Taken together,TAMs activate the Src-RhoA pathway to produce immunosuppressive factors and promote the proliferation and EMT of rectal cancer.

关 键 词：肿瘤相关巨噬细胞 Src-RhoA 免疫抑制因子 直肠癌 

分 类 号：R735.34[医药卫生—肿瘤]