中华耳鼻咽喉头颈外科杂志
中華耳鼻嚥喉頭頸外科雜誌
중화이비인후두경외과잡지
CHINESE JOURNAL OF OTORHINOLARYNGOLOGY HEAD AND NECK SURGERY
2014年
4期
326-329
,共4页
刘杨%周晓芳%文劲松%邓世山%刘海
劉楊%週曉芳%文勁鬆%鄧世山%劉海
류양%주효방%문경송%산세산%류해
膜联蛋白A5%基因沉默%细胞凋亡%喉肿瘤
膜聯蛋白A5%基因沉默%細胞凋亡%喉腫瘤
막련단백A5%기인침묵%세포조망%후종류
Annexin A5%Gene silencing%Apoptosis%Larygeal neoplasms
目的 探索annexinA5基因及其表达沉默后对人喉癌Hep-2细胞凋亡的影响.方法 培养人喉癌细胞Hep-2,经Lip2000转染特异siRNA片段沉默annexin A5基因;TRlzol法提取细胞总RNA;RT-PCR法检测annexinA5在mRNA水平的表达是否降低;Western blot法检测annexin A5的蛋白表达是否下降;经流式细胞仪AnnexinV-FITC法检测annexin A5基因沉默后的人喉癌Hep-2细胞的凋亡情况.采用SPSS 13.0软件对数据进行单因素方差分析.结果 特异siRNA片段成功转入Hep-2细胞;RT-PCR结果显示实验siRNA组、阴性siRNA组、脂质体组及空白细胞组的相对灰度值为0.70±0.03、1.18±0.05、1.17±0.06及1.23±0.07;经统计学分析表明实验组annexin A5在mRNA水平的灰度明显弱于阴性siRNA组(t=-14.77)、脂质体组(t=-13.23)及空白细胞组(t=-12.99),P值均＜0.05;Western blot结果显示实验siRNA组、阴性siRNA组、脂质体组及空白细胞组的相对灰度值为1.21±0.03、3.88±0.06、3.87±0.02及3.95±0.08;统计学分析表明annexin A5的蛋白的相对灰度值明显弱于阴性siRNA组(t=-70.34)、脂质体组(t=-150.62)及空白细胞组(t=-56.32),P值均＜0.05;AnnexinV-FITC法结果当annexin A5基因沉默后,人喉癌细胞Hep-2凋亡率实验siRNA组、阴性siRNA组、脂质体组及空白细胞组分别为4.43％±0.12％、13.67％±0.22％、13.66％±0.12％及13.35％±0.13％,实验组细胞凋亡率较阴性siRNA组(t=-62.50)、脂质体组(t=-14.16)及空白细胞组(t=-11.47)明显降低,P值均＜0.05.结论 在人喉癌Hep-2细胞中,annexin A5可能促进凋亡,有可能成为喉癌诊断、治疗及预后的新的生物靶标.
目的 探索annexinA5基因及其錶達沉默後對人喉癌Hep-2細胞凋亡的影響.方法 培養人喉癌細胞Hep-2,經Lip2000轉染特異siRNA片段沉默annexin A5基因;TRlzol法提取細胞總RNA;RT-PCR法檢測annexinA5在mRNA水平的錶達是否降低;Western blot法檢測annexin A5的蛋白錶達是否下降;經流式細胞儀AnnexinV-FITC法檢測annexin A5基因沉默後的人喉癌Hep-2細胞的凋亡情況.採用SPSS 13.0軟件對數據進行單因素方差分析.結果 特異siRNA片段成功轉入Hep-2細胞;RT-PCR結果顯示實驗siRNA組、陰性siRNA組、脂質體組及空白細胞組的相對灰度值為0.70±0.03、1.18±0.05、1.17±0.06及1.23±0.07;經統計學分析錶明實驗組annexin A5在mRNA水平的灰度明顯弱于陰性siRNA組(t=-14.77)、脂質體組(t=-13.23)及空白細胞組(t=-12.99),P值均＜0.05;Western blot結果顯示實驗siRNA組、陰性siRNA組、脂質體組及空白細胞組的相對灰度值為1.21±0.03、3.88±0.06、3.87±0.02及3.95±0.08;統計學分析錶明annexin A5的蛋白的相對灰度值明顯弱于陰性siRNA組(t=-70.34)、脂質體組(t=-150.62)及空白細胞組(t=-56.32),P值均＜0.05;AnnexinV-FITC法結果噹annexin A5基因沉默後,人喉癌細胞Hep-2凋亡率實驗siRNA組、陰性siRNA組、脂質體組及空白細胞組分彆為4.43％±0.12％、13.67％±0.22％、13.66％±0.12％及13.35％±0.13％,實驗組細胞凋亡率較陰性siRNA組(t=-62.50)、脂質體組(t=-14.16)及空白細胞組(t=-11.47)明顯降低,P值均＜0.05.結論 在人喉癌Hep-2細胞中,annexin A5可能促進凋亡,有可能成為喉癌診斷、治療及預後的新的生物靶標.
목적 탐색annexinA5기인급기표체침묵후대인후암Hep-2세포조망적영향.방법 배양인후암세포Hep-2,경Lip2000전염특이siRNA편단침묵annexin A5기인;TRlzol법제취세포총RNA;RT-PCR법검측annexinA5재mRNA수평적표체시부강저;Western blot법검측annexin A5적단백표체시부하강;경류식세포의AnnexinV-FITC법검측annexin A5기인침묵후적인후암Hep-2세포적조망정황.채용SPSS 13.0연건대수거진행단인소방차분석.결과 특이siRNA편단성공전입Hep-2세포;RT-PCR결과현시실험siRNA조、음성siRNA조、지질체조급공백세포조적상대회도치위0.70±0.03、1.18±0.05、1.17±0.06급1.23±0.07;경통계학분석표명실험조annexin A5재mRNA수평적회도명현약우음성siRNA조(t=-14.77)、지질체조(t=-13.23)급공백세포조(t=-12.99),P치균＜0.05;Western blot결과현시실험siRNA조、음성siRNA조、지질체조급공백세포조적상대회도치위1.21±0.03、3.88±0.06、3.87±0.02급3.95±0.08;통계학분석표명annexin A5적단백적상대회도치명현약우음성siRNA조(t=-70.34)、지질체조(t=-150.62)급공백세포조(t=-56.32),P치균＜0.05;AnnexinV-FITC법결과당annexin A5기인침묵후,인후암세포Hep-2조망솔실험siRNA조、음성siRNA조、지질체조급공백세포조분별위4.43％±0.12％、13.67％±0.22％、13.66％±0.12％급13.35％±0.13％,실험조세포조망솔교음성siRNA조(t=-62.50)、지질체조(t=-14.16)급공백세포조(t=-11.47)명현강저,P치균＜0.05.결론 재인후암Hep-2세포중,annexin A5가능촉진조망,유가능성위후암진단、치료급예후적신적생물파표.
Objective To study the effect of annexin A5 on the apoptosis of laryngeal cancer cells.Methods Special siRNAs were used to knock annexinA5 down in Hep-2 cell,and RT-PCR and Western blot were applied to identify the efficacy of RNA interference.The flow cytometry assay was performed to detect the Hep-2 cell apoptosis.Results RT-PCR analysis showed that the relative mRNA expression of annexin A5 in siRNA group,negative control group,Lipofectamine2000 group and blank control group were 0.70 ±0.03,1.18 ±0.05,1.17 ±0.06 and 1.23 ±0.07.The relative mRNA expression of annexin A5 in siRNA group was significantly decreased than contrast groups(t =-14.77,t =-13.23,t =-12.99,P ＜0.05).In Western blot assay,the trend of protein expression level was consistent with the mRNA expression levels of annexin A5.The relative levels of proteins in siRNA group,negative control group,Lipofectamine2000 group and blank control group were shown 1.21 ± 0.03,3.88 ± 0.06,3.87 ±0.02 and 3.95 ± 0.08.The relative protein expression of annexin A5 in siRNA group was significantly decreased than contrast groups(t =-70.34,t =-150.62,t =-56.32,P ＜0.05).At the same time in flow cytometry the apoptotic rate of siRNA group,negative control group,Lipofectamine2000 group and blank control group were 4.43％ ±0.12％,13.67％ ±0.22％,13.66％ ±0.12％ and 13.35％ ±0.13％,the difference between the siRNA group and contrast groups was statistically significant (t =-62.50,t =-14.16,t =-11.47,P ＜ 0.05).So after RNA interference,expression of annexin A5 decreased,and the results in the apoptosis inhibition of Hep-2 cell.Conclusion Annexin A5 promotes apoptosis of Hep-2 cells,and it may be a potential therapeutic target for the laryngeal cancer.
