长链非编码RNA MALAT1在骨肉瘤组织中的表达及其对骨肉瘤细胞侵袭和转移的影响

Expression of long non-coding RNA MALAT1 in osteosarcoma and its effect on invasiveness and metastatic potential of osteosarcoma cells

目的观察长链非编码RNA MALAT1与骨肉瘤的相关性，探讨其在骨肉瘤细胞侵袭和转移中的作用机制。方法应用即时荧光定量PCR（qRT－PCR）检测2014年1月至2015年12月就诊于河南省新乡医学院第一附属医院和河南省平顶山市第一人民医院的20例骨肉瘤患者肿瘤组织和配对瘤旁组织中MALAT1的表达水平，应用Mann－Whitney U检验分析其与临床病理参数的关系。应用慢病毒载体、MALAT1 shRNA慢病毒载体转染骨肉瘤U－2OS细胞，分别应用qRT－PCR法检测MALAT1表达情况；四甲基偶氮唑盐（MTT）比色法检测肿瘤细胞的增殖能力；细胞侵袭实验检测肿瘤细胞侵袭能力；免疫荧光和Western blot法检测Wnt/β－catenin信号通路相关蛋白的表达。结果在骨肉瘤患者肿瘤组织中MALAT1的表达水平高于配对瘤旁组织（P〈0.01）；MALAT1表达水平与淋巴结状态、远处转移（肺转移）相关（P〈0.01）；MTT法结果显示，沉默MALAT1可抑制骨肉瘤U－2OS细胞增殖（P〈0.05）；细胞侵袭实验结果显示，抑制MALAT1表达可降低U－2OS细胞的侵袭能力，与正常对照组和慢病毒载体组相比，差异有统计学意义（P〈0.01）；免疫荧光和Western blot结果显示，沉默MALAT1可下调Wnt/β－catenin信号通路相关蛋白β－catenin、基质金属蛋白酶7、c－MYC等的表达。结论MALAT1在骨肉瘤组织中高表达并与肿瘤侵袭密切相关；MALAT1促进骨肉瘤侵袭的机制可能与Wnt/β－catenin信号通路有关。

Objective To investigate the significance of long non-coding RNA MALAT1 expression in osteosarcoma, and the potential mechanism by which MALATI promotes tumor metastasis. Methods Twenty cases of osteosarcoma in the First Affiliated Hospital of Xinxiang Medical University and Ping Ding ShaM First People ＂s Hospital were collected from January 2014 to December 2015. The expression of MALAT1 in osteosarcoma tissue and paired adjacent noncancerous tissue were analyzed by qRT-PCR. Correlation of MALAT1 expression in osteosarcoma with clinical pathologic features was performed by the Mann-Whitney U test. U-2OS cells were transfected with lenti-virus carrying MALATI-shRNA and nonspecific shRNA （LV-vector）. The expression of MALAT1 was detected by qRT-PCR. The cell activity was evaluated by MTr asssy. The impact of MALATI-shRNA on invasion in U-20S cells were determined by transwell migration assay. The expression of Wnt/β-catenin signal pathway related proteins were detected by Immunofluorescence stain and Western blot. Results The expression level of MALAT1 in osteosarcoma tissue was higher than that in paired adjacent noncancerous tissue and correlated significantly with nodal and pulmonary metastasis（P 〈 0. 01 ）. MTT assay showed that knockdown of MALAT1 with lenti virus-MALAT1 shRNA inhibited the growth of U-2OS cells, along with marked decrease of invasive ability of U-2OS cells in the transwell migration assay. By immunofluorescence stain and Western blot assay, MALAT1 significantly reduced the expression of β-catenin, MMP7, and c-MYC in U-2OS ceils. Conclusions The expression of MALAT1 is high in osteosarcoma and correlates with tumor metastasis. MALAT1 promotes invasion and metastasis of osteosarcoma ceils likely thought the Wnt/β-catenin signal pathway.