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Effect of TLR2 on the proliferation of inflammation-related colorectal cancer and sporadic colorectal cancer
Cancer Cell International ( IF 5.8 ) Pub Date : 2020-03-30 , DOI: 10.1186/s12935-020-01184-0
Shuang Meng 1, 2 , Yingjie Li 1 , Xiaozhen Zang 1 , Zheng Jiang 3 , Huahan Ning 1 , Jing Li 1
Affiliation  

Colitis-associated cancer (CAC) is a complication of inflammatory bowel disease (IBD) with a poor prognosis because it is often diagnosed in advanced stages with local progression or metastasis. Compared with the more common polyp-induced sporadic colorectal cancer (sCRC), CAC has different molecular mechanisms. Toll-like receptor 2 (TLR2) expression is not limited to cells related to inflammation and immune function. High levels of TLR2 expression in tumor tissues of colorectal cancer (CRC) patients have been reported. This report is to investigate the effects of knockout and knockdown of the TLR2 gene on the proliferation of CAC and sCRC. Twelve C57BL/6 J wild-type mice (WT) and 12 TLR2 knockout mice (TLR2-/-) were used to rapidly establish a colitis-associated cancer (CAC) model via the 1,2-dimethylhydrazine-dextran sodium sulfate (DMH-DSS) method and were divided into the normal WT control group (NC), TLR2 knockout control group (KC), normal wild-type tumor modeling group (NT), and TLR2 knockout tumor modeling group (KT), with 6 mice in each group. The general performance of the mice during modeling, the gross changes of the colon and the rectum, and the pathological score of HE staining were used to observe tumor growth. The expression of TLR2 was detected by immunohistochemistry, and tumor proliferation was detected by Ki67 labeling. Lentivirus carrying TLR2-RNAi was used to stably infect colorectal cancer cells (HCT116 and HT29) to knock down TLR2 gene expression. The experimental groups included the uninfected control group, negative control group, and gene knockdown group. After infection, the expression of TLR2 protein was detected by Western blot, and cell proliferation and the cell cycle were detected by the CCK-8 method and fluorescence-activated cell sorting. Western blot was used to detect the expression levels of p- NF-κβ, cyclin D1 and cyclin D3 protein in each group of cells. TLR2 knockout in the CAC model resulted in greater changes in body weight and more severe diarrhea and colorectal hemorrhage. However, knocking out the TLR2 gene reduced the shortening of colorectal length, the number of tumors, and the total tumor volume and inhibited the growth of CAC. Knocking out the TLR2 gene also reduced the pathological score and tumor severity. TLR2 was localized in the cell membrane of the colorectal epithelium of the NC group and of the colorectal tumors of the NT group and was highly expressed in the NT group, while antigen Ki67 was localized in the nucleus of the colorectal tumor cells of the NT group and the KT group, and its expression was reduced in the KT group. In an in vitro sporadic colorectal cancer cell experiment, TLR2 protein in the TLR2 knockdown group was significantly downregulated, and TLR2 knockdown significantly inhibited the proliferation of HCT116 and HT29 colorectal cancer cells, resulting in G1 phase arrest. The expression levels of p-NF-κβ, cyclin D1 and cyclin D3 proteins in TLR2 gene knockdown group cells were significantly reduced. Knockout and knockdown of TLR2 can inhibit the proliferation of inflammation-related colorectal cancer and sporadic colorectal cancer.

中文翻译:

TLR2对炎症相关结直肠癌和散发性结直肠癌增殖的影响

结肠炎相关癌症 (CAC) 是炎症性肠病 (IBD) 的一种并发症,预后较差,因为它通常在具有局部进展或转移的晚期被诊断出来。与更常见的息肉诱发的散发性结直肠癌(sCRC)相比,CAC具有不同的分子机制。Toll 样受体 2 (TLR2) 表达不仅限于与炎症和免疫功能相关的细胞。据报道,结直肠癌 (CRC) 患者的肿瘤组织中存在高水平的 TLR2 表达。本报告旨在研究敲除和敲除TLR2基因对CAC和sCRC增殖的影响。12只C57BL/6 J野生型小鼠(WT)和12只TLR2敲除小鼠(TLR2-/-)通过1、2-二甲基肼-葡聚糖硫酸钠(DMH-DSS)法,分为正常WT对照组(NC)、TLR2敲除对照组(KC)、正常野生型肿瘤模型组(NT)、TLR2敲除肿瘤模型组组(KT),每组 6 只小鼠。通过小鼠造模时的一般表现、结肠和直肠的大体变化、HE染色病理评分观察肿瘤生长情况。免疫组化检测TLR2的表达,Ki67标记检测肿瘤增殖情况。携带 TLR2-RNAi 的慢病毒被用于稳定感染结肠直肠癌细胞(HCT116 和 HT29)以抑制 TLR2 基因表达。实验组包括未感染对照组、阴性对照组和基因敲除组。感染后,Western blot检测TLR2蛋白的表达,CCK-8法和荧光激活细胞分选法检测细胞增殖和细胞周期。Western blot检测各组细胞中p-NF-κβ、cyclin D1和cyclin D3蛋白的表达水平。CAC 模型中的 TLR2 敲除导致体重变化更大,腹泻和结肠直肠出血更严重。然而,敲除 TLR2 基因减少了结直肠长度、肿瘤数量和总肿瘤体积的缩短,并抑制了 CAC 的生长。敲除 TLR2 基因也降低了病理评分和肿瘤严重程度。TLR2定位于NC组大肠上皮和NT组大肠肿瘤的细胞膜,在NT组高表达,而抗原Ki67定位于NT组和KT组结直肠肿瘤细胞核内,KT组表达减少。在体外散发性大肠癌细胞实验中,TLR2敲低组TLR2蛋白显着下调,TLR2敲低显着抑制HCT116和HT29大肠癌细胞增殖,导致G1期阻滞。TLR2基因敲低组细胞中p-NF-κβ、cyclin D1和cyclin D3蛋白的表达水平显着降低。敲除和敲除TLR2可抑制炎症相关大肠癌和散发性大肠癌的增殖。在体外散发性大肠癌细胞实验中,TLR2敲低组TLR2蛋白显着下调,TLR2敲低显着抑制HCT116和HT29大肠癌细胞增殖,导致G1期阻滞。TLR2基因敲低组细胞中p-NF-κβ、cyclin D1和cyclin D3蛋白的表达水平显着降低。敲除和敲除TLR2可抑制炎症相关大肠癌和散发性大肠癌的增殖。在体外散发性大肠癌细胞实验中,TLR2敲低组TLR2蛋白显着下调,TLR2敲低显着抑制HCT116和HT29大肠癌细胞增殖,导致G1期阻滞。TLR2基因敲低组细胞中p-NF-κβ、cyclin D1和cyclin D3蛋白的表达水平显着降低。敲除和敲除TLR2可抑制炎症相关大肠癌和散发性大肠癌的增殖。TLR2基因敲低组细胞中cyclin D1和cyclin D3蛋白明显减少。敲除和敲除TLR2可抑制炎症相关大肠癌和散发性大肠癌的增殖。TLR2基因敲低组细胞中cyclin D1和cyclin D3蛋白明显减少。敲除和敲除TLR2可抑制炎症相关大肠癌和散发性大肠癌的增殖。
更新日期:2020-04-22
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