胃癌细胞中长链非编码RNA CCAT2的表达及其作用
| 作者: |
1邓浩,
2刘磊
1 湖北省武汉市红十字会医院 普通外科，湖北 武汉 430015 2 华中科技大学同济医学院附属武汉中心医院 胃肠外科，湖北 武汉 430014 |
| 通讯: |
刘磊
Email: haodeng333@126.com |
| DOI: | 10.3978/.2018.04.008 |
摘要
目的：探讨长链非编码RNA（lncRNA）CCAT2在胃癌细胞中的表达及其作用。
方法：用qRT-PCR检测胃癌细胞系AGS、Hs746T和BSG823及正常胃黏膜上皮细胞GES-1中CCAT2的表达，将胃癌AGS细胞分别转染CCAT2 siRNA（si-CCAT2组）与阴性对照siRNA（阴性对照组）后，以无转染的AGS细胞为空白对照，CCK-8法检测细胞的增殖情况，并分别用流式细胞术分、细胞划痕和Transwell实验、Western blot检测si-CCAT2组与阴性对照组的凋亡情况、迁移和侵袭能力以及凋亡相关蛋白的表达。
结果：各胃癌细胞系中CCAT2相对表达量均明显高于正常胃黏膜上皮细胞GES-1（均P<0.05）；转染后72、96 h，si-CCAT2组的增殖能力明显低于空白对照组与阴性对照组（均P<0.05），而阴性对照组与空白对照组各时间点增殖能力均无明显差异（均P>0.05）；与阴性对照组比较，si-CCAT2组细胞凋亡率明显升高、划痕愈合率明显降低、侵袭细胞数明显减少（均P<0.05）；P53、caspase-8、Bax蛋白表达上调，Bcl-2表达下调（均P<0.05）。
结论：CCAT2在胃癌细胞中高表升高，敲低其表达可抑制胃癌细胞的增殖及迁移与侵袭能力，机制可能与其调控凋亡相关蛋白的表达有关。
关键词：
胃肿瘤；RNA，长链非编码；细胞增殖；细胞凋亡；RNA干扰
方法：用qRT-PCR检测胃癌细胞系AGS、Hs746T和BSG823及正常胃黏膜上皮细胞GES-1中CCAT2的表达，将胃癌AGS细胞分别转染CCAT2 siRNA（si-CCAT2组）与阴性对照siRNA（阴性对照组）后，以无转染的AGS细胞为空白对照，CCK-8法检测细胞的增殖情况，并分别用流式细胞术分、细胞划痕和Transwell实验、Western blot检测si-CCAT2组与阴性对照组的凋亡情况、迁移和侵袭能力以及凋亡相关蛋白的表达。
结果：各胃癌细胞系中CCAT2相对表达量均明显高于正常胃黏膜上皮细胞GES-1（均P<0.05）；转染后72、96 h，si-CCAT2组的增殖能力明显低于空白对照组与阴性对照组（均P<0.05），而阴性对照组与空白对照组各时间点增殖能力均无明显差异（均P>0.05）；与阴性对照组比较，si-CCAT2组细胞凋亡率明显升高、划痕愈合率明显降低、侵袭细胞数明显减少（均P<0.05）；P53、caspase-8、Bax蛋白表达上调，Bcl-2表达下调（均P<0.05）。
结论：CCAT2在胃癌细胞中高表升高，敲低其表达可抑制胃癌细胞的增殖及迁移与侵袭能力，机制可能与其调控凋亡相关蛋白的表达有关。
Expression of long non-coding RNA CCAT2 in gastric cancer cells and its action
CorrespondingAuthor:LIU Lei Email: haodeng333@126.com
Abstract
Objective: To investigate the expression of the long non-coding RNA (lncRNA) CCAT2 in gastric cancer cells and its actions.
Methods: The expressions of CCAT2 in different gastric cancer cell lines (AGS, Hs746T and BSG823) and normal gastric mucosal GES-1 cells were detected by qRT-PCR. The gastric cancer AGS cells were transfected with CCAT2 siRNA (si-CCAT2 group) or scrambled siRNA sequences (negative control group) respectively, and then their proliferative abilities were measured by CCK-8 assay, using untransfected AGS cells as blank control. In si-CCAT2 group and negative control group, the apoptosis, the migration and invasion abilities and the expressions of apoptosis-associated proteins were determined by flow cytometry, scratch assay, Transwell assay and Western blot analysis, respectively.
Results: The relative expression levels of CCAT2 in all studied gastric cancer cell lines were significantly higher than that in the normal gastric mucosal GES-1 cells (all P<0.05). At 72 and 96 h after transfection, the proliferative ability in si-CCAT2 group was significantly lower than that in negative control group or blank control group (all P<0.05), while, no significant difference in proliferative ability was noted between negative control group and blank control group at each predefined time point (all P>0.05). In si-CCAT2 group compared with negative control group, the apoptosis rate was increased, and the wound healing rate and the number of invading cells were decreased significantly (all P<0.05); the protein expressions of P53, caspase-8 and Bax were up-regulated, while the protein expression of Bcl-2 was down-regulated significantly (all P<0.05).
Conclusion: CCAT2 expression is increased in gastric cancer cells. Knockdown of its expression can inhibit the proliferation and abilities of migration and invasion of the gastric cancer cells, and the mechanism may be related to its regulating the expressions of apoptosis-associated proteins.
Keywords：
Stomach Neoplasms; RNA
Long Noncoding; Cell Proliferation; Apoptosis; RNA Interference
Methods: The expressions of CCAT2 in different gastric cancer cell lines (AGS, Hs746T and BSG823) and normal gastric mucosal GES-1 cells were detected by qRT-PCR. The gastric cancer AGS cells were transfected with CCAT2 siRNA (si-CCAT2 group) or scrambled siRNA sequences (negative control group) respectively, and then their proliferative abilities were measured by CCK-8 assay, using untransfected AGS cells as blank control. In si-CCAT2 group and negative control group, the apoptosis, the migration and invasion abilities and the expressions of apoptosis-associated proteins were determined by flow cytometry, scratch assay, Transwell assay and Western blot analysis, respectively.
Results: The relative expression levels of CCAT2 in all studied gastric cancer cell lines were significantly higher than that in the normal gastric mucosal GES-1 cells (all P<0.05). At 72 and 96 h after transfection, the proliferative ability in si-CCAT2 group was significantly lower than that in negative control group or blank control group (all P<0.05), while, no significant difference in proliferative ability was noted between negative control group and blank control group at each predefined time point (all P>0.05). In si-CCAT2 group compared with negative control group, the apoptosis rate was increased, and the wound healing rate and the number of invading cells were decreased significantly (all P<0.05); the protein expressions of P53, caspase-8 and Bax were up-regulated, while the protein expression of Bcl-2 was down-regulated significantly (all P<0.05).
Conclusion: CCAT2 expression is increased in gastric cancer cells. Knockdown of its expression can inhibit the proliferation and abilities of migration and invasion of the gastric cancer cells, and the mechanism may be related to its regulating the expressions of apoptosis-associated proteins.