中华肝脏病杂志
中華肝髒病雜誌
중화간장병잡지
CHINESE JOURNAL OF HEPATOLOGY
2011年
11期
853-856
,共4页
公方晓%夏景林%杨毕伟%徐晓晶%吴伟忠
公方曉%夏景林%楊畢偉%徐曉晶%吳偉忠
공방효%하경림%양필위%서효정%오위충
癌,肝细胞%细胞周期蛋白D1%细胞增殖
癌,肝細胞%細胞週期蛋白D1%細胞增殖
암,간세포%세포주기단백D1%세포증식
Carcinoma,hepatocellular%Cyclin D1%Cell proliferation
目的 用瞬时转染的方法 研究let-7c对人肝癌细胞HCCLM3增殖的影响并探讨其机制.方法 用脂质体2000将miRNA转染人人肝癌细胞HCCLM3,设let-7c组转染let-7c,设对照组转染阴性对照miRNA,设空白组不做任何转染.用细胞计数试剂盒CCK-8检测转染后细胞生长增殖的变化,用流式细胞术检测各组细胞周期分布的差异.用Western blot和实时荧光PCR检测转染后细胞周期蛋白(cyclin) D1及其mRNA表达的变化.多组数据的两两比较采用LSD法.结果 let-7c组细胞在转染后48、72h时的吸光度值分别为0.70±0.05、0.77±0.09,低于空白组的0.97±0.10、1.21±0.12和对照组的0.91±0.07、1.12±0.09,各组比较,F值分别为14.431、21.146,P值均＜0.01,差异均有统计学意义.转染后72h,let-7c使癌细胞处在G1期的细胞比例增加.空白组、对照组、let-7c组G1期的细胞比例分别为43.53％±0.86％、44.82％±0.77％、54.52％％±0.13％,各组比较,F=240.739,P＜0.01,差异有统计学意义.let-7c能抑制cyclin D1及其mRNA的表达,空白组、对照组、let-7c组cyclin D1的表达水平分别为0.48±0.09、0.47±0.06、0.23±0.06,各组比较,F=11.316,P＜0.01,差异有统计学意义; mRNA的相对表达量分别为1.03％±0.29％、1.01％±0.11％、0.63％±0.14％,各组比较,F=6.315,P＜0.05,差异有统计学意义.结论 let-7c能抑制HCCLM3细胞的增殖,并使细胞停留在G1期的细胞比例增加.导致这种现象的机制可能为let-7c抑制cyclin Dl及其mRNA的表达.
目的 用瞬時轉染的方法 研究let-7c對人肝癌細胞HCCLM3增殖的影響併探討其機製.方法 用脂質體2000將miRNA轉染人人肝癌細胞HCCLM3,設let-7c組轉染let-7c,設對照組轉染陰性對照miRNA,設空白組不做任何轉染.用細胞計數試劑盒CCK-8檢測轉染後細胞生長增殖的變化,用流式細胞術檢測各組細胞週期分佈的差異.用Western blot和實時熒光PCR檢測轉染後細胞週期蛋白(cyclin) D1及其mRNA錶達的變化.多組數據的兩兩比較採用LSD法.結果 let-7c組細胞在轉染後48、72h時的吸光度值分彆為0.70±0.05、0.77±0.09,低于空白組的0.97±0.10、1.21±0.12和對照組的0.91±0.07、1.12±0.09,各組比較,F值分彆為14.431、21.146,P值均＜0.01,差異均有統計學意義.轉染後72h,let-7c使癌細胞處在G1期的細胞比例增加.空白組、對照組、let-7c組G1期的細胞比例分彆為43.53％±0.86％、44.82％±0.77％、54.52％％±0.13％,各組比較,F=240.739,P＜0.01,差異有統計學意義.let-7c能抑製cyclin D1及其mRNA的錶達,空白組、對照組、let-7c組cyclin D1的錶達水平分彆為0.48±0.09、0.47±0.06、0.23±0.06,各組比較,F=11.316,P＜0.01,差異有統計學意義; mRNA的相對錶達量分彆為1.03％±0.29％、1.01％±0.11％、0.63％±0.14％,各組比較,F=6.315,P＜0.05,差異有統計學意義.結論 let-7c能抑製HCCLM3細胞的增殖,併使細胞停留在G1期的細胞比例增加.導緻這種現象的機製可能為let-7c抑製cyclin Dl及其mRNA的錶達.
목적 용순시전염적방법 연구let-7c대인간암세포HCCLM3증식적영향병탐토기궤제.방법 용지질체2000장miRNA전염인인간암세포HCCLM3,설let-7c조전염let-7c,설대조조전염음성대조miRNA,설공백조불주임하전염.용세포계수시제합CCK-8검측전염후세포생장증식적변화,용류식세포술검측각조세포주기분포적차이.용Western blot화실시형광PCR검측전염후세포주기단백(cyclin) D1급기mRNA표체적변화.다조수거적량량비교채용LSD법.결과 let-7c조세포재전염후48、72h시적흡광도치분별위0.70±0.05、0.77±0.09,저우공백조적0.97±0.10、1.21±0.12화대조조적0.91±0.07、1.12±0.09,각조비교,F치분별위14.431、21.146,P치균＜0.01,차이균유통계학의의.전염후72h,let-7c사암세포처재G1기적세포비례증가.공백조、대조조、let-7c조G1기적세포비례분별위43.53％±0.86％、44.82％±0.77％、54.52％％±0.13％,각조비교,F=240.739,P＜0.01,차이유통계학의의.let-7c능억제cyclin D1급기mRNA적표체,공백조、대조조、let-7c조cyclin D1적표체수평분별위0.48±0.09、0.47±0.06、0.23±0.06,각조비교,F=11.316,P＜0.01,차이유통계학의의; mRNA적상대표체량분별위1.03％±0.29％、1.01％±0.11％、0.63％±0.14％,각조비교,F=6.315,P＜0.05,차이유통계학의의.결론 let-7c능억제HCCLM3세포적증식,병사세포정류재G1기적세포비례증가.도치저충현상적궤제가능위let-7c억제cyclin Dl급기mRNA적표체.
Objective To investigate let-7c's effect on the proliferation of human hepatocellular carcinoma cell HCCLM3 by transient transfection and the mechanism inside.Methods Lipofectamine 2000 was used to transfect miRNAs into HCCLM3 cells.The cells were divided into three groups,let-7c group:let-7c was transfected,negative control group:negative control miRNA was transfected,blank control group:nothing was transfected.The proliferation of HCCLM3 cells was evaluated using Cell Counting Kit8(CCK-8).The cell cycles of each group were assayed by flow cytometry.Western blot and Real time PCR were used to analyze the protein and mRNA expressions of cyclin D1.Statistical analysis was perfomed with SPSS 17.0.Results The absorbances of let-7c group were 0.70 ± 0.05,0.77 ± 0.09 at 48h and 72 h after transfection,lower than that of blank control group (0.97 ± 0.10,1.21 ± 0.12) and negative control group (0.91 ± 0.07,1.12 ± 0.09),48 h:F =14.431,P ＜0.05,72 h:F =21.146,P ＜ 0.05.The flow cytometry at 72 h after traasfection revealed that let-7c increased the percentage of cells in G1 phase.The percentage of blank control group was 43.53％ ± 0.86％,the negative control group was 44.82％±0.77％,and the let-7c group was 54.52％ ± 0.13％,F =240.739,P ＜ 0.05.let-7c suppressed expressions of cyclin D1 at both protein and mRNA levels.The protein levels of cyclin D1 were 0.48 ± 0.09,0.47 ± 0.06 and 0.23 ±:0.06 (F =11.316,P ＜ 0.05) in blank control group,negative control group and let-7c group,respectively.The mRNA levels were 1.03％ ±0.29％,1.01％ ± 0.11％ and 0.63％ ± 0.14％ (F =6.315,P ＜ 0.05) in the above three groups,respectively.Conclusion let-7c can inhibit proliferation of HCCLM3 cells and increase the proportion of cells in Gl phase.The mechanism may be that let-7c represses the expressions of cyclin D 1 at both protein and mRNA levels.
