凝溶胶蛋白表达稳定下调的小鼠肝癌高淋巴道转移力Hca-F细胞株的构建  

The construction of mouse hepatocarcinoma Hca-F cell line with high lymphatic metastasis potentials in which gelsolin is down-expressed stably

作　　者：王绍清[1] 唐建武[1] 孙明忠[2] 刘淑清[3] 王波[1] 

机构地区：[1]大连医科大学病理学教研室,辽宁大连116044 [2]大连医科大学,生物技术专业辽宁大连116044 [3]大连医科大学分子生物学教研室,辽宁大连116044

出　　处：《辽宁师范大学学报（自然科学版）》2009年第4期495-499,共5页Journal of Liaoning Normal University:Natural Science Edition

基　　金：国家自然科学基金项目(30572098)

摘　　要：在前期蛋白质组学和基因组学研究发现凝溶胶蛋白(Gelsolin,GSN)是一个潜在促进小鼠肝癌淋巴道转移基因基础上,本次实验设计合成3条GSN的shRNA短发夹序列(shRNA1、2、3),并成功将它们构建入pSilencer3.1质粒,重组质粒转染鼠肝癌高淋巴道转移力Hca-F细胞,获得了稳定转染的pSilencer-GSN-Hca-F细胞株.结果表明:GSN空白对照组和无关序列对照组细胞中在mRNA及蛋白表达水平均相近;与空白对照组相比较,3个重组质粒在mRNA水平对GSN抑制率分别为46.8%、48.2%、20.2%,在蛋白水平对GSN的抑制率分别为73.9%、45.1%、31.2%;shRNA1重组质粒的抑制效果最明显,统计学意义显著(P<0.01).通过RNA干扰和基因转染技术成功建立了GSN表达稳定下调的鼠肝癌Hca-F细胞株,为进一步研究GSN在恶性肿瘤淋巴道转移中的作用奠定了基础.Previous quantitative proteomic and genomic results from our laboratory indicated the expression of gelsolin（GSN） was correlated positively with the lymphatic metastasis potentials of mouse hepatocarcinoma.In current study,three short hairpins RNA 1,2 and 3（shRNA 1,2,3） of mouse GSN were designed and synthesized and constructed into the pSilencer plasmid,respectively.Then the stably transfected cell line,pSilencer-GSN-Hca-F,was obtained by transfecting the mouse high lymphatic metastasis potential cell line（Hca-F） against the recombined plasmids mentioned above.The results indicated that the GSN expressed closely at both the mRNA level and protein level in both the non-transfected Hca-F and independent sequence transfected Hca-F cell lines.Comparing to the non-transfected group Hca-F cell line,shRNA 1,2 and 3 could down-regulate the GSN expression at mRNA level by 46.8%,48.2% and 20.2%,as well as down-regulate the GSN expression at protein level by 73.9%,45.1% and 31.2%,respectively.Among the three shRNAs,the shRNA1 showed the strongest inhibition with statistics significance.The mouse hepatocarcinoma Hca-F cell lines with stably down-regulated expression of GSN were constructed with the aids of RNAi and gene transfection techniques,which will unravel the role of GSN playing in the lymphatic metastasis mechanism of malignant tumors.

关 键 词：凝溶胶蛋白 淋巴道转移 肝癌 RNA干扰 基因转染 

分 类 号：R735.7[医药卫生—肿瘤]