过表达PTEN基因对甲状腺髓样癌细胞生长及细胞周期的影响
| 作者: |
1翟明慧,
2袁殿宝,
1赵峻峰,
1张敬
1 河北北方学院附属第一医院肿瘤内科，河北 张家口 075000 2 河北北方学院附属第一医院消化内科，河北 张家口 075000 |
| 通讯: |
翟明慧
Email: 1685337381@qq.com |
| DOI: | 10.3978/j.issn.2095-6959.2019.03.002 |
摘要
目的：探讨PTEN基因对甲状腺髓样癌(medullary thyroid carcinoma，MTC)细胞生长、周期的影响以及作用机制。方法：使用酶切酶连的方法构建真核表达载体pcDNA3.1-PTEN，采用脂质体转染法将其转入pA10RP8细胞中。实时荧光定量PCR(real-time quantitative polymerase chain reaction， RT-qPCR)检测细胞内PTEN表达水平的变化，Western印迹法检测PTEN，Ki-67，p21，cyclin D1蛋白表达水平的变化；CCK-8法和平板克隆试验检测PTEN对pA10RP8细胞增殖的影响；细胞流式术检测pA10RP8细胞周期的变化。结果：成功构建了PTEN真核表达载体。转染pcDNA3.1-PTEN质粒的pA10RP8细胞PTEN和p21表达上调，Ki-67和cyclin D1蛋白表达下调；转染pcDNA3.1-PTEN质粒的pA10RP8细胞生长增殖受到抑制；细胞在G0/G1期比例增加，发生周期阻滞。结论：过表达PTEN能够抑制pA10RP8细胞的增殖，诱导细胞周期阻滞，其机制可能与周期蛋白p21上调和cyclin D1下调表达有关。
关键词：
甲状腺髓样癌；PTEN；细胞周期；细胞周期蛋白
Effect of over-expression of PTEN gene on growth and cell cycle of medullary thyroid carcinoma cells
CorrespondingAuthor: ZHAI Minghui Email: 1685337381@qq.com
DOI: 10.3978/j.issn.2095-6959.2019.03.002
Abstract
Objective: To investigate the effect of over-expression of PTEN gene on the growth and cell cycle of medullary thyroid carcinoma cells and to explore its mechanism. Methods: The eukaryotic expression vector pcDNA3.1-PTEN was constructed by restriction enzyme linked method transferred into pA10RP8 cells by liposome transfection. The mRNA expression level of PTEN in the cells were detected by RT-qPCR, the protein expression level of PTEN, Ki-67, p21 and cyclin D1 were detected by Western blot; the effect of PTEN in pA10RP8 cells on the proliferation was detected by CCK-8 and flat clones; and the changes of cell cycle were detected by flow cytometry. Results: The PTEN eukaryotic expression vector was successfully constructed; the expression of PTEN and p21 in cells with pcDNA3.1-PTEN was significantly up-regulated; the expression of Ki-67 and cyclin D1 protein in pA10RP8 cells with pcDNA3.1-PTEN was down-regulated; the proliferation of pA10RP8 cells with pcDNA3.1-PTEN was inhibited; the PTEN gene induced cell cycle arrest of pA10RP8 cells with a significant increase of the proportion of pA10RP8 cells in G0/G1 phase. Conclusion: Over-expression of PTEN could inhibit the proliferation and induce cell cycle arrest of pA10RP8 cells, whose mechanism may be related to p21 and cyclin D1 protein.
Keywords：
medullary thyroid carcinoma; PTEN; cell cycle; cyclin