肿瘤相关巨噬细胞分泌的抗菌肽cathelicidin促进小鼠结直肠癌生长的作用及机制  

Effect and mechanism of antimicrobial peptide cathelicidin secreted by tumor associated macrophage on the growth of colorectal cancer in mice

作　　者：权文强[1] 潘星昊 吴军录[1] 姚懿雯[1] 李冬[1] Quan Wenqiang, Pan Xinghao, Wu Junlu, Yao Yiwen, Li Dong(Department of Clinical Laboratory, Tongfi Hospital, University of Tongji, Shanghai 200065, China; School of Biomedical Sciences, Huaqiao University, Xiamen 361021, Chin)

机构地区：[1]同济大学附属同济医院检验科,上海200065 [2]华侨大学生物医学学院,厦门361021

出　　处：《中华肿瘤杂志》2018年第3期172-177,共6页Chinese Journal of Oncology

基　　金：国家自然科学基金(81272603、81472179);上海市浦江人才计划(13PJ1407300);上海市第四轮公共卫生体系建设三年行动计划(15GWZK0301)

摘　　要：目的研究肿瘤相关巨噬细胞分泌的抗菌肽cathelicidin促进小鼠结直肠癌生长的作用及机制。方法通过氧化偶氮甲烷/葡聚糖硫酸钠（AOM/DSS）方法建立小鼠肠炎相关的结直肠癌模型。对诱导成瘤的小鼠，同时每隔3 d注射1次小鼠cathelicidin中和抗体、IgG抗体（阳性对照）和PBS（阴性对照），持续1个月，取小鼠结肠，拍照、计数肿瘤数目并进行评价。利用实时荧光定量PCR和Western blot法检测小鼠结肠肿瘤组织、非肿瘤组织以及组织中巨噬细胞cathelicidin的表达；采用免疫组化法检测小鼠结肠肿瘤组织与非肿瘤组织中的cathelicidin、肿瘤增殖抗原Ki-67和CD68的表达；采用原位末端转移酶标记技术（TUNEL）检测小鼠结肠肿瘤组织中细胞的凋亡情况。结果cathelicidin在小鼠结肠肿瘤组织的炎症免疫细胞（巨噬细胞）中呈高表达状态，而在肿瘤细胞中表达较弱。注射cathelicidin中和抗体组小鼠的肿瘤数目和长径分别为（4.50±1.18）个和（1.74±0.18）mm，明显低于注射PBS组小鼠的肿瘤数目[（13.88±1.98）个，P〈0. 01]和肿瘤长径[（3.74±0.38）mm，P〈0. 01]；也明显低于注射IgG抗体小鼠的肿瘤数目[（15.25±1.82）个，P〈0. 01]和肿瘤长径[（3.40±0.36）mm，P〈0.01]。注射cathelicidin中和抗体组肿瘤组织中Ki-67阳性肿瘤细胞百分比为（28.20±3.44）%，低于注射PBS组[（68.20±3.51）%，P〈0.01]和注射IgG抗体组[（69.20±3.41）%，P〈0.01]。免疫组织化学染色检测结果显示，注射cathelicidin中和抗体组小鼠组织中CD68的表达明显低于注射IgG抗体组和注射PBS组。TUNEL染色显示，注射cathelicidin中和抗体对肿瘤细胞的凋亡影响不大。结论肿瘤相关巨噬细胞分泌的cathelicidin具有促进小鼠结直肠癌增殖的作用，中和cathelicidin的表达可以抑制结直肠癌的生长和增殖。cathelicidin通过募集炎症细胞（巨噬细胞）进入肿瘤微�ObjectiveTo investigate the effect and mechanism of the antibacterial peptide cathelicidin secreted by tumor associated macrophages on the growth of colorectal cancer in mice.MethodsAzoxymethane （AOM）/ dextran sodium sulfate （DSS） method was used to establish a mouse model of colitis associated colon cancer. To induce tumor formation, cathelicidin antibody, IgG antibody （positive control） or PBS （negative control） was respectively injected into mice once every 3 days and lasted one month. Then the pictures of mice colon were taken, and the numbers of tumor were counted and evaluated. Expressions of cathelicidin in tumor associated macrophages isolated from tumor and adjacent normal tissues of mice were examined by quantitative RT-PCR （qRT-PCR） and Western blot. Expressions of the tumor proliferating antigen Ki-67, macrophage marker CD68 and cathelicidin in tumor and non-tumor tissues were determined by immunohistochemistry analysis. Apoptosis of cells from tumor tissues was analyzed by using TdT-mediated dUTP nick-end labeling （TUNEL）.ResultsIn colon tumor tissues, cathlicidin strongly expressed in inflammatory cells （macrophages）, but weakly expressed in tumor cells. The tumor number and size in mice injected with cathelicidin neutralizing antibody were 4.50±1.18 and （1.74±0.18） mm, respectively, significantly lower than 13.88±1.98 and （3.74±0.38） mm of mice injected with PBS （t=4.07, t=4.72; P〈 0.01） and 15.25±1.82 and （3.40±0.36） mm of mice injected with IgG antibody （t=4.96, t=4.08; P〈0.01）. The Ki-67 positive rate of cells in tumor tissues of mice injected with cathelicidin neutralizing antibody was （28.20±3.44） %, significantly lower than （68.20±3.51） % of mice injected with PBS （t=8.135, P〈0.01） and （69.20±3.41） % of mice injected with IgG antibody （t=8.461, P〈0.01）. Immunohistochemistry analyses showed that the expression of CD68 in tumor tissues of mice injected with cathelicidin antibody was significantly lower than tha

关 键 词：结直肠肿瘤 CATHELICIDIN 肿瘤微环境 炎症 肿瘤相关巨噬细胞 

分 类 号：R735.34[医药卫生—肿瘤]