Inhibition of EBV-encoded LMP-1 by DNA-based RNA interference affects metastatic ability of nasopharyngeal carcinoma cells

Abstract

OBJECTIVE: To evaluate the application of DNA vector-based RNA interference for silencing LMP-1 expression in EBV-positive nasopharyngeal cancer (NPC) C611 cells and analyze the inhibiting effects of LMP-1 silencing on metastatic property of NPC cells. METHODS: DNA vectors with LMP-1 shRNA template were transfected into C611 cells. After G418 selection, a LMP-1 silenced sub-clone named C611L was isolated. Effect of LMP-1 inhibition on its metastatic properties was evaluated by cell adhesion, invasion and wound healing assays. RESULTS: shRNA vector mediated stable RNAi against LMP-1 in C611 cells. Loss of LMP-1 significantly altered cell motility, substratum adhesion and transmembrane invasion ability. CONCLUSION: Stable suppression of LMP-1 induced by shRNA vector can inhibit metastatic properties of NPC cells.

目的探討應用DNA載體介導RNA干擾技術,抑制鼻咽癌細胞中EB病毒潛伏膜蛋白1(LMP-1)表達的可能性,觀察LMP-1基因沉默對鼻咽癌細胞轉移能力的影響。方法將能轉錄出干擾RNA的DNA模板插入pAVU6+27質粒,構建能夠介導RNA干擾的發夾狀干擾RNA (shRNA)表達載體。采用磷酸鈣共沉淀法,將質粒導入EB病毒(+)的鼻咽癌細胞C611,篩選LMP-1基因沉默的克隆,通過觀察其貼壁生長能力、基底膜穿透能力和移動能力的變化,分析LMP-1基因沉默對鼻咽癌細胞轉移能力的影響。結果在shRNA表達載體有效轉染的C611細胞中,LMP-1基因的表達受到穩定抑制。腫瘤細胞轉移實驗表明,LMP-1基因沉默的C611細胞,貼壁生長能力提高,基底膜穿透能力和細胞移動能力均明顯下降。結論shRNA表達載體能夠在鼻咽癌細胞內介導RNA干擾,并獲得對LMP-1基因穩定的抑制。LMP-1基因可能通過貼壁生長能力、基底膜穿透能力和細胞移動能力等,影響鼻咽癌細胞的轉移。