LRRC4基因表达能降低胶质母细胞瘤细胞系U251的生长和成瘤潜能

背景与目的：LRRC4是作者最近克隆的一个新基因,该基因在原发性脑肿瘤活检标本中明显表达下调。本研究旨在研究LRRC4基因是否具有抑制脑肿瘤生长的能力。方法：LRRC4基因的全长编码区被亚克隆至表达载体pcDNA3．1中,应用脂质体转染的方法将重组的质粒载体导入胶质母细胞瘤细胞系U251,经G418筛选,建立稳定表达LRRC4基因的U251的细胞系。采用细胞增殖实验、软琼脂实验、肿瘤形成实验来考察LRRC4基因表达对于细胞生长和肿瘤形成的影响。结果：经过脂质体转染和筛选,建立了稳定表达LRRC4全长编码区的U251细胞系,用于进一步实验。比较未转染组和转染空白载体组,Northern blot实验证实转染了LRRC4基因的细胞LRRC4 mRNA的表达增强。细胞增殖一定时间后,转染LRRC4基因的细胞较未转染细胞的生长速度明显减慢,克隆形成率明显降低。将这些细胞注射入无胸腺裸鼠体内,40天后处死裸鼠,测量肿瘤大小,结果显示,转染LRRC4基因的细胞形成的肿瘤明显小于对照组。结论：LRRC4基因可转染于人脑胶质母细胞瘤细胞系U251。LRRC4在U251细胞的表达有抑制瘤细胞增殖和抑制裸鼠移植瘤的形成和生长的作用。

Expression of LRRC4 has the potential to decrease the growth rate and tumorigenesis of glioblastoma cell line U251

BACKGROUND & OBJECTIVE: LRRC4 is a novel gene that theauthor has identified recently, which displayed significant downregulation inprimary brain tumor biopsies. This study was designed to investigate if LRRC4has the potential of suppressing brain tumor growth. METHODS: The full-lengthcoding region of LRRC4 gene was subcloned into the expression vectorpcDNA3.1, the recombinant was introduced into the glioblastoma cell line U251by liposome transfection, and the U251 cells stably expressing LRRC4 genewere established by G418 selection. Furthermore, cell proliferation assay, softagar assay, tumorigenesis assay were taken to examine the effect of LRRC4expression on cell growth and tumor formation. RESULTS: U251 cells stablyexpressing full-length coding region of LRRC4 were established bylipofection-mediated transfection and selected for further study. Compared withthe nontransfected and vector-transfected cells, the cells transfected with LRRC4cDNA exhibited a significant increase of expression of LRRC4 mRNA by Northernblot analysis. Further, when cell proliferation was followed over several days, thecells expressing the transfected LRRC4 cDNA grew more slowly thannontransfected cells. Consistently, the cells transfected with LRRC4 exhibitedmarkedly lower colony formation rate. These clones were injected into athymicnude mice who was killed after 40 days and the tumor sizes were evaluated.Tumor volume in mice was significantly smaller in the group of cells stablytransfected with LRRC4 cDNA than in the control. CONCLUSION: LRRC4 genemay be transfected into the human glioblastoma cell line U251. The expression ofLRRC4 in U251 cells may have the potential to suppress tumor cell growth andthe tumorigenesis of U251 cell transplanted in nude mice.