摘要
目的探讨多功能蛋白聚糖(VCAN)基因对膀胱尿路上皮癌(BUC)免疫细胞浸润及预后的影响。方法从TCGA数据库下载BUC的高通量测序数据及临床数据,运用R语言Limma包分析VCAN表达水平与BUC患者临床特征的相关性;采用单因素和多因素Cox回归分析VCAN表达情况及患者临床特征(包括性别、年龄、肿瘤分级、肿瘤分期、淋巴转移和远处转移等)对预后的影响;基于肿瘤免疫浸润评分(TIMER)数据库分析VCAN表达水平与免疫细胞浸润的相关性。取T24细胞,设置对照组(未转染任何siRNA)、NC siRNA组(转染阴性对照siRNA)与VCAN siRNA组(转染VCAN siRNA),采用MTT法、细胞侵袭实验和流式细胞术分别检测各组细胞的增殖、侵袭能力和细胞周期比例,Western blotting检测Cyclin E、Cyclin D1、E-cadherin和基质金属蛋白酶-9(MMP-9)的表达水平。结果与正常膀胱组织比较,BUC组织中VCAN mRNA表达水平明显上调,且与肿瘤分级、肿瘤分期及远处转移呈正相关(P<0.01);VCAN高表达是BUC患者预后不良的独立危险因素(P<0.05);TIMER数据库分析结果显示,BUC中VCAN表达水平与巨噬细胞及调节性T细胞的浸润程度呈明显正相关(P<0.001)。与对照组比较,VCAN siRNA组T24细胞的增殖和侵袭能力明显下降,G_(0)/G_(1)期细胞比例明显增高(P<0.05),Cyclin E、Cyclin D1和MMP-9蛋白表达水平明显下调,E-cadherin蛋白表达水平明显上调(P<0.05)。结论VCAN在BUC中呈高表达,且与BUC的免疫浸润及预后相关。沉默VCAN可明显抑制T24细胞的增殖及侵袭,其机制可能与调控Cyclin E、Cyclin D1、E-cadherin及MMP-9蛋白的表达有关。
Objective To explore the effect of versican(VCAN)gene on immunocyte infiltration and prognosis of bladder urothelial carcinoma(BUC).Methods The high-throughput sequencing data and clinical data of BUC were downloaded from TCGA database.The correlation between the expression level of VCAN and clinical characteristics of BUC patients was analyzed using the Limma R package;univariate and multivariate Cox regression analysis were performed to analyze the effect of VCAN expression and clinical characteristics(including gender,age,tumor grade,tumor stage,lymph node metastasis and distant metastasis)on the prognosis of BUC patients;TIMER database was used to analyze the correlation between VCAN expression level and immunocyte infiltration.T24 cells were divided into three groups:control group(cells transfected without any siRNA),NC siRNA group(cells transfected with negative control siRNA),and VCAN siRNA group(cells transfected with VCAN siRNA).After transfection,the proliferation,invasion and cell cycle proportion of T24 cells were determined by MTT assay,Transwell assay and flow cytometry,respectively.The protein expression levels of Cyclin E,Cyclin D1,E-cadherin and matrix metalloproteinase-9(MMP-9)were determined by Western blotting.Results Compared with normal bladder tissues,the expression level of VCAN mRNA in BUC tissues was significantly up-regulated,and was positively correlated with tumor grade,tumor stage and distant metastasis(P<0.01).High VCAN expression was an independent risk factor for poor prognosis in BUC patients(P<0.05).The results of TIMER database analysis showed that the expression level of VCAN was significantly positively correlated with the infiltration degree of macrophages and regulatory T cells in BUC(P<0.001).Compared with control group,the proliferation and invasion ability of T24 cells in VCAN siRNA group decreased significantly and the cell proportion of G_(0)/G_(1) phase increased significantly(P<0.05),the protein expression levels of Cyclin E,Cyclin D1 and MMP-9 were down-regulated si
作者
吕磊
郑福鑫
周高峰
吴维
Lv Lei;Zheng Fu-Xin;Zhou Gao-Feng;Wu Wei(Department of Urology,Wuhan No.1 Hospital,Wuhan,Hubei 430022,China)
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2023年第8期921-928,共8页
Medical Journal of Chinese People's Liberation Army
基金
国家自然科学基金(81902593)
湖北省自然科学基金(2020CFB732)
武汉市科技计划项目(2020020601012325)。
