长链非编码RNA MANCR诱导肿瘤相关成纤维细胞分化对结肠癌细胞的影响  

Effect of long non-coding RNA MANCR-induced differentiation of cancer-associated fibroblasts on colon cancer cells

作　　者：潘妍 朱绍华 卢瑗瑗 PAN Yan;ZHU Shaohua;LU Yuanyuan(Department of Gastroenterology,First Affiliated Hospital of Air Force Medical University,Xi'an 710032,China)

机构地区：[1]空军军医大学第一附属医院消化内科,陕西西安710032

出　　处：《国际消化病杂志》2025年第1期25-31,68,共8页International Journal of Digestive Diseases

基　　金：国家自然科学基金(82273142)。

摘　　要：目的探究长链非编码RNA(lncRNA)MANCR诱导的肿瘤相关成纤维细胞(CAF)对结肠癌细胞增殖、迁移和侵袭的影响。方法将人正常结肠成纤维细胞CCD-18Co分别转染MANCR阴性对照质粒和MANCR pcDNA 3.1过表达质粒,对应设为Control组和MANCR组。将CCD-18Co细胞转染MANCR pcDNA 3.1过表达质粒,然后分别与SCH772984及相同体积的PBS共孵育,对应设为MANCR+SCH772984组和MANCR+PBS组。将人结肠癌细胞HCT116、Caco-2分别与Control组及MANCR组CCD-18Co细胞上清液共孵育,对应设为CCD-18Co组和MANCR CCD-18Co组。采用实时荧光定量PCR法检测各组CCD-18Co细胞中CAF标志物成纤维细胞激活蛋白(FAP)、纤连蛋白(Fibronectin)、基质金属蛋白酶-9(MMP-9)、α-平滑肌动蛋白(α-SMA)mRNA表达水平,采用ELISA法检测各组CCD-18Co细胞上清液中IL-8、IL-1β和IL-6水平。采用CCK8法、细胞划痕实验和Transwell小室法分别检测CCD-18Co组和MANCR CCD-18Co组HCT116、Caco-2细胞的增殖、迁移和侵袭能力。采用蛋白质印迹法检测相关蛋白表达水平,以明确各组CCD-18Co细胞中细胞外信号调节激酶(ERK)信号通路的激活情况,以及CCD-18Co组和MANCR CCD-18Co组HCT116、Caco-2细胞中磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路的激活情况。结果与Control组相比,MANCR组CCD-18Co细胞中FAP、Fibronectin、MMP-9、α-SMA mRNA表达水平均显著升高(P均<0.001),细胞上清液中IL-8、IL-1β、IL-6水平均显著升高(P均<0.05),ERK磷酸化水平显著升高(P<0.001)。与MANCR+PBS组相比,MANCR+SCH772984组CCD-18Co细胞中FAP、Fibronectin、MMP-9、α-SMA mRNA表达水平均显著降低(P均<0.01),细胞上清液中IL-8、IL-1β、IL-6水平均显著降低(P均<0.001)。与CCD-18Co组相比,MANCR CCD-18Co组HCT116、Caco-2细胞的增殖、迁移和侵袭能力均显著增强(P均<0.05),细胞中p-PI3K/PI3K和p-Akt/Akt水平均显著升高(P均<0.001)。结论lncRNA MANCR诱导的CAF分化可激活结肠癌细胞PI3K/Akt信号通路,�Objective This paper aims to explore the effects of long non-coding RNA(lncRNA)MANCR-induced cancer-associated fibroblasts(CAF)on the proliferation,migration,and invasion of colon cancer cells.Methods Normal human colon fibroblast cells CCD-18Co were transfected with either a MANCR negative control plasmid or a MANCR pcDNA 3.1 overexpression plasmid,corresponding to the Control group and the MANCR group.MANCR pcDNA 3.1 overexpression plasmid was also transfected into CCD-18Co cells,which were then co-cultured with SCH772984 or an equal volume of PBS,corresponding to the MANCR+SCH772984 group and the MANCR+PBS group.Human colon cancer cells HCT116 and Caco-2 were co-incubated with the supernatant from CCD-18Co cells in both the Control group and the MANCR group,respectively,referred to as the CCD-18Co group and the MANCR CCD-18Co group,respectively.Real-time fluorescence quantitative PCR was used to detect the mRNA expression levels of CAF markers—fibroblast activation protein(FAP),Fibronectin,matrix metalloproteinase-9(MMP-9),and alpha smooth actin(α-SMA)in CCD-18Co cells from each group.ELISA was used to measure the levels of IL-8,IL-1β,and IL-6 in the supernatant of CCD-18Co cells.The proliferation,migration,and invasion abilities of HCT116 and Caco-2 cells from the CCD-18Co group and the MANCR CCD-18Co group were assessed using CCK8 method,cell scratch assay,and Transwell assays.Western blotting was performed to assess the activation of the ERK signaling pathway in CCD-18Co and the activation of the PI3K/Akt signaling pathway in HCT116 and Caco-2 cells.Results Compared with the Control group,the mRNA expression levels of FAP,Fibronectin,MMP-9,andα-SMA in CCD-18Co cells in the MANCR group are significantly increased(P<0.001),and the levels of IL-8,IL-1β,and IL-6 in the cell supernatant are significantly elevated(P<0.05).The phosphorylation level of ERK is significantly increased(P<0.001).Compared with the MANCR+PBS group,the mRNA expression levels of FAP,Fibronectin,MMP-9,andα-SMA in CCD-18Co cells of th

关 键 词：MANCR 肿瘤相关成纤维细胞 结肠癌细胞 增殖 迁移 侵袭 

分 类 号：R735.35[医药卫生—肿瘤]