徐州医学院学报
徐州醫學院學報
서주의학원학보
Acta Academiae Medicinae Xuzhou
2015年
10期
648-653
,共6页
罗文雅%周凯%胡磊%李向阳%孔凡运%尤红娟%汤仁仙
囉文雅%週凱%鬍磊%李嚮暘%孔凡運%尤紅娟%湯仁仙
라문아%주개%호뢰%리향양%공범운%우홍연%탕인선
c-Jun%RNA小干扰%HepG2细胞%增殖%迁移
c-Jun%RNA小榦擾%HepG2細胞%增殖%遷移
c-Jun%RNA소간우%HepG2세포%증식%천이
c-Jun%small interfering RNA%HepG2 cells%proliferation%migration
目的：构建c－Jun特异性的shRNA及研究其抑制c－Jun蛋白表达对人类肝癌细胞HepG2增殖和迁移的影响。方法根据c－Jun编码序列设计并构建shRNA质粒，利用LipofectamineTM 2000转染HepG2细胞，荧光显微镜观察转染效率。 Western blot检测其对c－Jun蛋白表达的影响。 CCK－8增殖实验、平板克隆形成实验观察HepG2细胞的增殖情况，transwell和划痕愈合实验方法观察细胞的迁移情况。结果构建了c－Jun小干扰质粒shRNA－1和shRNA－2。荧光显微镜观察到shRNA质粒在HepG2细胞转染效率约在80％。 Western blot显示shRNA－2小干扰质粒对c－Jun抑制效果要强于shRNA－1小干扰质粒。功能实验结果显示，与阴性对照组相比，shRNA－2能显著抑制HepG2细胞的增殖、克隆形成及其迁移能力（P均＜0．05）。结论成功构建了对c－Jun有效shRNA干扰质粒，下调c－Jun的表达能够明显抑制肝癌细胞的增殖和迁移，为进一步研究c－Jun在肝癌中的作用及相关机制提供了工作基础。
目的：構建c－Jun特異性的shRNA及研究其抑製c－Jun蛋白錶達對人類肝癌細胞HepG2增殖和遷移的影響。方法根據c－Jun編碼序列設計併構建shRNA質粒，利用LipofectamineTM 2000轉染HepG2細胞，熒光顯微鏡觀察轉染效率。 Western blot檢測其對c－Jun蛋白錶達的影響。 CCK－8增殖實驗、平闆剋隆形成實驗觀察HepG2細胞的增殖情況，transwell和劃痕愈閤實驗方法觀察細胞的遷移情況。結果構建瞭c－Jun小榦擾質粒shRNA－1和shRNA－2。熒光顯微鏡觀察到shRNA質粒在HepG2細胞轉染效率約在80％。 Western blot顯示shRNA－2小榦擾質粒對c－Jun抑製效果要彊于shRNA－1小榦擾質粒。功能實驗結果顯示，與陰性對照組相比，shRNA－2能顯著抑製HepG2細胞的增殖、剋隆形成及其遷移能力（P均＜0．05）。結論成功構建瞭對c－Jun有效shRNA榦擾質粒，下調c－Jun的錶達能夠明顯抑製肝癌細胞的增殖和遷移，為進一步研究c－Jun在肝癌中的作用及相關機製提供瞭工作基礎。
목적：구건c－Jun특이성적shRNA급연구기억제c－Jun단백표체대인류간암세포HepG2증식화천이적영향。방법근거c－Jun편마서렬설계병구건shRNA질립，이용LipofectamineTM 2000전염HepG2세포，형광현미경관찰전염효솔。 Western blot검측기대c－Jun단백표체적영향。 CCK－8증식실험、평판극륭형성실험관찰HepG2세포적증식정황，transwell화화흔유합실험방법관찰세포적천이정황。결과구건료c－Jun소간우질립shRNA－1화shRNA－2。형광현미경관찰도shRNA질립재HepG2세포전염효솔약재80％。 Western blot현시shRNA－2소간우질립대c－Jun억제효과요강우shRNA－1소간우질립。공능실험결과현시，여음성대조조상비，shRNA－2능현저억제HepG2세포적증식、극륭형성급기천이능력（P균＜0．05）。결론성공구건료대c－Jun유효shRNA간우질립，하조c－Jun적표체능구명현억제간암세포적증식화천이，위진일보연구c－Jun재간암중적작용급상관궤제제공료공작기출。
Objective To construct specific short hairpin RNA ( shRNA) against c-Jun and explore its effects on the proliferation and migration of HepG2 cells.Methods According to the coding sequence of c-Jun, two shRNA plas-mids were designed and transfected into HepG2 cells by lipofectamineTM 2000.The transfection efficiency was observed by fluorescence microscopy.The effects of shRNA on the level of c-Jun were measured by Western blot.The proliferation of the cells was tested by CCK-8 assay and colony formation assay.The migration of the cells was determined by tran-swell assay and wound healing assay.Results Two shRNA plasmids were constructed ( named shRNA-1 and shRNA-2), and their transfection efficiency was 80%by fluorescence microscopy.According to Western blot analysis, shRNA-2 produced stronger inhibitory effects against c-Jun than shRNA-2 (P<0.05).Compared with the negative control, shRNA-2 could remarkably block the proliferation, colony formation and migration of HepG2 cells (P<0.05).Con-clusion The shRNA plasmid against c-Jun is successfully constructed, which can markedly inhibit the proliferation and migration of HepG2 cells, providing experimental evidence for further research on the function and related mecha-nisms of c-Jun in liver cancer.
