胆酸诱导肿瘤相关巨噬细胞促进肠腺瘤癌变机制探讨  

Cholic acid induced carcinogenesis of intestinal adenoma by altering phenotype of tumor associated macrophage

作　　者：达彬琳 曹海龙[1] 王斯南[1] 刘天宇[1] 董文逍 王丹[1] 王邦茂[1] DA Bin-lin;CAO Hai-long;WANG Si-nan;LIU Tian-yu;DONG Wen-xiao;WANG Dan;WANG Bang-mao(Department of Gastroenterology and Hepatology,General Hospital,Tianjin Medical University,Tianjin 300052,P.R.China)

机构地区：[1]天津医科大学总医院消化科

出　　处：《中华肿瘤防治杂志》2019年第21期1579-1586,共8页Chinese Journal of Cancer Prevention and Treatment

基　　金：国家自然科学基金（81470796;81570478）;天津市应用基础与前沿技术研究计划（15JCZDJC36600）

摘　　要：目的持续暴露胆酸可诱导肠腺瘤癌变,机制尚不清楚,本研究旨在探讨肿瘤相关巨噬细胞表型转化在胆酸诱导小鼠肠腺瘤癌变中的作用。方法 20只4周龄APCmin/+小鼠随机分为对照组和胆酸组,每组10只。对照组常规饮食,胆酸组在常规饮食基础上添加0.4%胆酸。给药12周后处死,观察两组肠道肿瘤数目、大小及癌变率。HE染色评价肠道肿瘤病理类型,Ki-67免疫组织化学染色评价肿瘤细胞增殖水平。实时荧光定量PCR及免疫组织化学染色方法评价单核细胞趋化蛋白1(monocyte chemotactic protein 1,MCP-1)的表达。实时荧光定量PCR检测肠道炎症因子、巨噬细胞表面分子F4/80、M1型巨噬细胞表面分子诱导性一氧化氮合酶(inductible nitric oxide synthase,iNOS)、趋化因子配体10(CXC chemokine ligand-10,CXCL-10)、M2型巨噬细胞表面分子甘露糖受体(mannose receptor,MR)、精氨酸1(argnine-1,Arg-1)和趋化因子配体17(chemokine ligand-17,CCL-17)的表达水平。运用免疫荧光双染方法检测肠道息肉组织中巨噬细胞表面分子F4/80、M1型及M2型巨噬细胞表面分子的表达。结果胆酸组小鼠全肠道腺瘤总数(33.40±1.17)较对照组(22.80±0.93)明显增加,差异有统计学意义,t=7.11,P=0.002。胆酸组小鼠肠道肿瘤Ki-67阳性细胞百分比为(70.60±5.45)%,较对照组的(29.80±1.66)%明显增加,差异有统计学意义,t=7.17,P<0.001。胆酸可导致小鼠肠道低度炎症的产生,实时荧光定量PCR显示胆酸组小鼠肠道IL-1β相对表达量为14.14±2.78,较对照组的1.24±0.14明显增加,差异有统计学意义,t=4.62,P=0.009 9。IL-6(3.40±0.11)和TNF-α(3.99±0.44)相对表达量明显高于对照组的1.45±0.29(t=7.32,P=0.001 8)和1.42±0.24(t=5.07,P=0.007 1)。且胆酸组MCP-1的相对表达量(13.25±1.55)明显高于对照组(1.96±0.68),t=5.07,P=0.002 6。免疫组织化学染色显示,胆酸组小鼠肠道组织间质MCP-1表达(86.67±2.03)高于对照组(43.99±4.73),t=8.49,P=OBJECTIVE To investigate the role of altering the phenotype of tumor associated macrophages(TAM on carcinogenesis of intestinal adenoma promoted by cholic acid(CA)in APCmin/+mice.METHODS Twenty four-weekold APCmin/+mice were divided into two groups randomly:control group(regular diet)and CA group(0.4% CA in regular diet).All mice were sacrificed after 12 weeks.The number,size and location of intestinal adenomas were observed.The pathology was evaluated via hematcxylin-eosin(HE)staining.Ki-67 expression was detected by immunohistochemistry(IHC)to evaluate the cell proliferation.Intestinal inflammatory factors and macrophage surface molecular F4/80 were measured by real-time PCR.Monocyte chemotactic protein 1(MCP-1)was detected by IHC and real-time PCR.Immunofluorescence double staining were employed to detect the impact of cholic acid on altering the phenotype of TAM.RESULTS Compared with control group(22.80±0.93),the number(33.40±1.17,t=7.11,P=0.002)and carcinogenesis of the intestinal tumors.The elevated positive cells of Ki-67 were observed in CA group(70.60±5.45)compared with that of the CA group increased control group(29.80±1.66,t=7.17,P<0.000 1).Although there was no inflammatory infiltration according to HE staining,the relative mRNA expression levels of IL-1β(14.14±2.78),IL-6(3.40±0.11)and TNF-α(3.99±0.44)were dramatically increased in CA group,compared with that of control group,which were1.24±0.14(t=4.62,P=0.009 9),1.45±0.29(t=7.32,P=0.001 8)and 1.42±0.24(t=5.07,P=0.007 1).The relative mRNA expression levels of MCP-1 were increased in CA group(13.25±1.55)compared with that of control group(1.96±0.68,t=5.07,P=0.002 6).Based on IHC,the protein levels of MCP-1 in stroma of intestinal polyps in CA group(86.67±2.03)were significantly increased compared with that of the control group(43.99±4.73,t=8.49,P=0.001 1).CONCLUSION CA induced low-grade intestinal inflammation and upregulated MCP-1,and then shifted macrophage phenotype in tumor microenvironment from M1 to M2 type,and induced the carcino

关 键 词：胆酸 肠腺瘤 癌变 MCP-1 肿瘤相关巨噬细胞 

分 类 号：R73[医药卫生—肿瘤]