中华消化杂志
中華消化雜誌
중화소화잡지
Chinese Journal of Digestion
2013年
7期
460-464
,共5页
赵笛%蔡晨雯%刘炯%萧树东%郑青
趙笛%蔡晨雯%劉炯%蕭樹東%鄭青
조적%채신문%류형%소수동%정청
胃肿瘤%细小病毒H-1%病毒非结构蛋白质类%基因,肿瘤抑制%细胞衰老%细胞周期
胃腫瘤%細小病毒H-1%病毒非結構蛋白質類%基因,腫瘤抑製%細胞衰老%細胞週期
위종류%세소병독H-1%병독비결구단백질류%기인,종류억제%세포쇠로%세포주기
Stomach neoplasms%H-1 parvovirus%Viral nonstructural proteins%Genes,tumor suppressor%Cell aging%Cell cycle
目的 探讨细小病毒H-1非结构蛋白1(NS1)基因对人胃癌细胞的抑制作用及其可能的机制.方法 构建表达增强型绿色荧光蛋白(eGFP)标记的NS1重组质粒(peGFP-C1-NS1),分别使用peGFP-C1-NS1(实验组)或空载体质粒(peGFP-C1,阴性对照组)转染人胃癌SGC7901细胞株,空白对照组则予等量磷酸盐缓冲液处理.转染后在荧光显微镜下观察细胞内荧光信号分布,检测NS1基因及蛋白表达情况.绘制细胞生长曲线,检测细胞衰老相关β-半乳糖苷酶(SA-β-Gal)的表达,并使用流式细胞仪分析细胞周期分布变化.两组间比较行t检验.结果 转染peGFP-C1-NS1后SGC7901细胞表达NS1基因及蛋白,与阴性对照组相比,实验组细胞荧光信号集中于细胞核内.实验组SA-β-Gal阳性细胞比例[(30.5±1.4)％]高于阴性对照组[(4.4±1.1)％],差异有统计学意义(t=-12.931,P＜0.01).转染peGFP-C1-NS1后1、2、3、4 d SGC7901细胞的抑制率分别为45％、62％、73％、77％.表达eGFP-NS1融合蛋白的SGC7901细胞细胞周期被阻滞于G0/G1期.结论 细小病毒H-1 NS1在胃癌SGC7901细胞核内发挥作用,可将细胞周期阻滞于G0/G1期,有效抑制SGC7901细胞生长.
目的 探討細小病毒H-1非結構蛋白1(NS1)基因對人胃癌細胞的抑製作用及其可能的機製.方法 構建錶達增彊型綠色熒光蛋白(eGFP)標記的NS1重組質粒(peGFP-C1-NS1),分彆使用peGFP-C1-NS1(實驗組)或空載體質粒(peGFP-C1,陰性對照組)轉染人胃癌SGC7901細胞株,空白對照組則予等量燐痠鹽緩遲液處理.轉染後在熒光顯微鏡下觀察細胞內熒光信號分佈,檢測NS1基因及蛋白錶達情況.繪製細胞生長麯線,檢測細胞衰老相關β-半乳糖苷酶(SA-β-Gal)的錶達,併使用流式細胞儀分析細胞週期分佈變化.兩組間比較行t檢驗.結果 轉染peGFP-C1-NS1後SGC7901細胞錶達NS1基因及蛋白,與陰性對照組相比,實驗組細胞熒光信號集中于細胞覈內.實驗組SA-β-Gal暘性細胞比例[(30.5±1.4)％]高于陰性對照組[(4.4±1.1)％],差異有統計學意義(t=-12.931,P＜0.01).轉染peGFP-C1-NS1後1、2、3、4 d SGC7901細胞的抑製率分彆為45％、62％、73％、77％.錶達eGFP-NS1融閤蛋白的SGC7901細胞細胞週期被阻滯于G0/G1期.結論 細小病毒H-1 NS1在胃癌SGC7901細胞覈內髮揮作用,可將細胞週期阻滯于G0/G1期,有效抑製SGC7901細胞生長.
목적 탐토세소병독H-1비결구단백1(NS1)기인대인위암세포적억제작용급기가능적궤제.방법 구건표체증강형록색형광단백(eGFP)표기적NS1중조질립(peGFP-C1-NS1),분별사용peGFP-C1-NS1(실험조)혹공재체질립(peGFP-C1,음성대조조)전염인위암SGC7901세포주,공백대조조칙여등량린산염완충액처리.전염후재형광현미경하관찰세포내형광신호분포,검측NS1기인급단백표체정황.회제세포생장곡선,검측세포쇠로상관β-반유당감매(SA-β-Gal)적표체,병사용류식세포의분석세포주기분포변화.량조간비교행t검험.결과 전염peGFP-C1-NS1후SGC7901세포표체NS1기인급단백,여음성대조조상비,실험조세포형광신호집중우세포핵내.실험조SA-β-Gal양성세포비례[(30.5±1.4)％]고우음성대조조[(4.4±1.1)％],차이유통계학의의(t=-12.931,P＜0.01).전염peGFP-C1-NS1후1、2、3、4 d SGC7901세포적억제솔분별위45％、62％、73％、77％.표체eGFP-NS1융합단백적SGC7901세포세포주기피조체우G0/G1기.결론 세소병독H-1 NS1재위암SGC7901세포핵내발휘작용,가장세포주기조체우G0/G1기,유효억제SGC7901세포생장.
Objective To investigate the suppression effect of expressing parvovirus H-1 nonstructural protein 1 (NS1) gene on human gastric cancer cells and the possible mechanisms.Methods A recombinant enhanced green fluorescent protein (eGFP) labeled NS1 of parvovirus H-1 plasmid was constructed.Human gastric cancer cell line SGC7901 was transfected with recombinant plasmid (experiment group) or blank vector (negative control group) and blank control group was treated with equal amount of phosphate buffered saline (blank control group).After transfection,the distribution of fluorescent signal was observed under fluorescent microscope.The expression of NS1 at gene and protein level was measured.Cell growth curve of each group was drawn.The expression of cell senescence-associated β-galactosidase (SA-β-Gal) was tested.The changes of cell cycle were investigated by flowcytometry.Two groups' comparision was performed by t-test.Results After transfection,NS1 was expressed in SGC7901 cells at gene and protein level.Compared with negative control group,the fluorescent signal accumulated in cell nucleus in experiment group.The percentage of SA-β-Gal positive cell in experiment group ((30.5 ± 1.4) ％) was higher than that of negative control group ((4.4± 1.1) ％) and the difference was statistically significant (t =-12.931,P ＜ 0.01).The growth inhibition rate of SGC7901 cells from the first day to the fourth day was 45％,62％,73％ and 77％,respectively.The cell cycle of eGFP-NS1 expressed SGC7901 cells was arrested at G0/G1 phase.Conclusion Parvovirus H-1 NS1 play the role in cell nucleus of gastric cancer cell line SGC7901 and could make cell cycle arrested at G0/G1 phase,which effectively inhibited the proliferation SGC7901 cell.
