中华肝胆外科杂志
中華肝膽外科雜誌
중화간담외과잡지
CHINESE JOURNAL OF HEPATOBILIARY SURGERY
2015年
6期
405-409
,共5页
叶健文%周闯%阎冰%杨佳%傅哲%唐文超%翟文龙
葉健文%週闖%閻冰%楊佳%傅哲%唐文超%翟文龍
협건문%주틈%염빙%양가%부철%당문초%적문룡
钙网蛋白%肝癌细胞%小干扰RNA%p-Akt蛋白
鈣網蛋白%肝癌細胞%小榦擾RNA%p-Akt蛋白
개망단백%간암세포%소간우RNA%p-Akt단백
Calreticulin%Hepatocellular carcinoma cells%Small interfering RNA%p-Akt protein
目的 探讨钙网蛋白(CRT)对人肝癌细胞株SMMC-7721、HepG2增殖及侵袭能力的影响.方法 利用siRNA沉默肝癌细胞株SMMC-7721、HepG2 CRT的表达.通过免疫荧光技术及蛋白印迹法检测CRT siRNA转染效果;Cell counting kit-8(CCK-8)检测细胞的增殖;采用流式细胞术检测细胞凋亡率变化;运用Transwell侵袭小室检测细胞侵袭能力;蛋白印迹法检测p-Akt、Akt蛋白表达.结果 siRNA实验组SMMC-7721和HepG2细胞24、36、48 h相对生长抑制率分别为(41.0±2.2)％、(46.5±1.6)％、(59.7±2.2)％和(36.8±2.7)％、(47.3±1.8)％、(61.5±3.2)％.与空白组及阴性对照组比较,siRNA实验组增殖能力明显降低(P＜0.05).沉默CRT 36 h后SMMC-7721、HepG2细胞的凋亡率分别(45.2±9.1)％、(48.9±8.0)％.与空白组及阴性对照组比较,siRNA实验组凋亡率上调(P＜0.05).Transwell实验显示:沉默CRT 36 h后SMMC-7721和HepG2细胞穿膜个数空白组、阴性对照组、siRNA实验组分别为:(96.8±7.3)、(95.6±5.4)、(34.0±4.2)和(124.0±9.9)、(121.6 ±7.0)、(70.4 ±9.5).与空白组及阴性对照组比较,实验组侵袭能力降低(P＜0.05).沉默CRT 36 h后,与空白组及阴性对照组比较,p-Akt蛋白表达水平均下调(P＜0.05).结论 沉默肝癌细胞CRT对肝癌细胞的增殖、侵袭能力有明显的抑制作用,同时可诱导细胞的凋亡,且与PI3K/Akt信号通路相关.
目的 探討鈣網蛋白(CRT)對人肝癌細胞株SMMC-7721、HepG2增殖及侵襲能力的影響.方法 利用siRNA沉默肝癌細胞株SMMC-7721、HepG2 CRT的錶達.通過免疫熒光技術及蛋白印跡法檢測CRT siRNA轉染效果;Cell counting kit-8(CCK-8)檢測細胞的增殖;採用流式細胞術檢測細胞凋亡率變化;運用Transwell侵襲小室檢測細胞侵襲能力;蛋白印跡法檢測p-Akt、Akt蛋白錶達.結果 siRNA實驗組SMMC-7721和HepG2細胞24、36、48 h相對生長抑製率分彆為(41.0±2.2)％、(46.5±1.6)％、(59.7±2.2)％和(36.8±2.7)％、(47.3±1.8)％、(61.5±3.2)％.與空白組及陰性對照組比較,siRNA實驗組增殖能力明顯降低(P＜0.05).沉默CRT 36 h後SMMC-7721、HepG2細胞的凋亡率分彆(45.2±9.1)％、(48.9±8.0)％.與空白組及陰性對照組比較,siRNA實驗組凋亡率上調(P＜0.05).Transwell實驗顯示:沉默CRT 36 h後SMMC-7721和HepG2細胞穿膜箇數空白組、陰性對照組、siRNA實驗組分彆為:(96.8±7.3)、(95.6±5.4)、(34.0±4.2)和(124.0±9.9)、(121.6 ±7.0)、(70.4 ±9.5).與空白組及陰性對照組比較,實驗組侵襲能力降低(P＜0.05).沉默CRT 36 h後,與空白組及陰性對照組比較,p-Akt蛋白錶達水平均下調(P＜0.05).結論 沉默肝癌細胞CRT對肝癌細胞的增殖、侵襲能力有明顯的抑製作用,同時可誘導細胞的凋亡,且與PI3K/Akt信號通路相關.
목적 탐토개망단백(CRT)대인간암세포주SMMC-7721、HepG2증식급침습능력적영향.방법 이용siRNA침묵간암세포주SMMC-7721、HepG2 CRT적표체.통과면역형광기술급단백인적법검측CRT siRNA전염효과;Cell counting kit-8(CCK-8)검측세포적증식;채용류식세포술검측세포조망솔변화;운용Transwell침습소실검측세포침습능력;단백인적법검측p-Akt、Akt단백표체.결과 siRNA실험조SMMC-7721화HepG2세포24、36、48 h상대생장억제솔분별위(41.0±2.2)％、(46.5±1.6)％、(59.7±2.2)％화(36.8±2.7)％、(47.3±1.8)％、(61.5±3.2)％.여공백조급음성대조조비교,siRNA실험조증식능력명현강저(P＜0.05).침묵CRT 36 h후SMMC-7721、HepG2세포적조망솔분별(45.2±9.1)％、(48.9±8.0)％.여공백조급음성대조조비교,siRNA실험조조망솔상조(P＜0.05).Transwell실험현시:침묵CRT 36 h후SMMC-7721화HepG2세포천막개수공백조、음성대조조、siRNA실험조분별위:(96.8±7.3)、(95.6±5.4)、(34.0±4.2)화(124.0±9.9)、(121.6 ±7.0)、(70.4 ±9.5).여공백조급음성대조조비교,실험조침습능력강저(P＜0.05).침묵CRT 36 h후,여공백조급음성대조조비교,p-Akt단백표체수평균하조(P＜0.05).결론 침묵간암세포CRT대간암세포적증식、침습능력유명현적억제작용,동시가유도세포적조망,차여PI3K/Akt신호통로상관.
Objective To explore the effect of calreticulin (CRT) on cell proliferation and invasion of human hepatocellular carcinoma cell line SMMC-7721 and HepG2.Methods SMMC-7721 and HepG2 cells were transfected with small interfering RNA (siRNA).The transfection rate was detected by immunoflurescence and western blot.The cell proliferation,invasion and apoptosis of SMMC-7721 and HepG2 cells were determined by using cell counting kit-8 (CCK-8) assays,transwell assays and flow cytometry,respectively.The p-Akt and Akt levels were detected by western blot.Results The growth inhibition rate in the siRNA experimental group of SMMC-7721 and HepG2 cells for 24,36 and 48 h were (41.0 ±2.2) ％,(46.5 ±1.6)％,(59.7 ±2.2)％ and (36.8 ±2.7)％,(47.3 ± 1.8)％,(61.5 ±3.2)％,respectively.The apoptosis rate after down-regulating the expression of CRT in SMMC-7721 and HepG2 cells for 36h were (45.2 ± 9.1) ％ and (48.9 ± 8.0) ％,respectively.Compared with the blank group and the negative control group,the growth inhibition rate in the siRNA experimental group was lower (P ＜0.05),but the apoptosis rate was significantly higher (P ＜ 0.05).Transwell experiments confirmed that the numbers of invaded SMMC-7721 and HepG2 cells in the blank group and the negative control group and siRNA experimental group were (96.8±7.3),(95.6±5.4),(34.0±4.2) and (124.0 ±9.9),(121.6 ±7.0),(70.4±9.5),respectively,indicating that cell invasion in the siRNA experimental group was significantly suppressed (P ＜ 0.05).The expression of p-Akt was decreased (P ＜ 0.05) after down-regulating the expression of CRT for 36h.Conclusion CRT gene silencing by siRNA can inhibit the SMMC-7721 and HepG2 cell proliferation and invasion,but increase the cell apoptosis by regulating PI3K/Akt signal pathway.
