CAJ | 학술논문

目的：探讨应用四环素调控单纯疱疹病毒胸苷激酶（HSV-tk）基因在 HeLa 细胞内表达及抗肿瘤作用。方法将含有四环素阻遏蛋白基因的质粒载体 pcDNA6/TR 与含有四环素反应元件及 HSV-tk 基因的质粒载体 pcD-NA3．1-TetO2-TKI 先后稳定转染 HeLa 细胞，给予不同浓度的四环素类似物强力霉素，应用 RT-PCR 检测细胞内HSV-tk 基因表达，应用 MTT 法测定转染 HSV-tk 基因的 HeLa 细胞对不同浓度无氧鸟苷（GCV）杀伤的敏感性。结果转染两种质粒载体的 HeLa 细胞经不同浓度的强力霉素作用48 h 后，RT-PCR 结果显示，随着强力霉素浓度的增大，HSV-tk 基因表达量逐渐增加，强力霉素4μg/ml 时，基因表达的水平达到高峰；加入强力霉素后，该细胞对GCV 杀伤细胞的敏感性增加。结论本研究成功地应用四环素基因表达调控系统调控了 HSV-tk 基因在 HeLa 细胞内表达及杀伤肿瘤功能，为进一步探讨自杀基因杀伤肿瘤的可调控性提供理论基础。
목적：탐토응용사배소조공단순포진병독흉감격매（HSV-tk）기인재 HeLa 세포내표체급항종류작용。방법장함유사배소조알단백기인적질립재체 pcDNA6/TR 여함유사배소반응원건급 HSV-tk 기인적질립재체 pcD-NA3．1-TetO2-TKI 선후은정전염 HeLa 세포，급여불동농도적사배소유사물강력매소，응용 RT-PCR 검측세포내HSV-tk 기인표체，응용 MTT 법측정전염 HSV-tk 기인적 HeLa 세포대불동농도무양조감（GCV）살상적민감성。결과전염량충질립재체적 HeLa 세포경불동농도적강력매소작용48 h 후，RT-PCR 결과현시，수착강력매소농도적증대，HSV-tk 기인표체량축점증가，강력매소4μg/ml 시，기인표체적수평체도고봉；가입강력매소후，해세포대GCV 살상세포적민감성증가。결론본연구성공지응용사배소기인표체조공계통조공료 HSV-tk 기인재 HeLa 세포내표체급살상종류공능，위진일보탐토자살기인살상종류적가조공성제공이론기출。
Objective To investigate the effect of tetracycline regulated HSV-tk gene expression and anti-tumor in HeLa cells.Methods The plasmid vector pcDNA6/TR containing the tetracycline repressor protein gene and pcD-NA3.1-TetO2-TKI containing tetracycline response element and HSV-tk gene both were stably transfected into HeLa cells.After treated with different concentrations of tetracycline analogue doxycycline,expression of HSV-tk gene was detected by RT-PCR ,determination of the sensitivity of HSV-tk gene transfected HeLa cells to different concentrations of GCV application the MTT method.Results HeLa cells that transfected with the two kinds of plasmid,was treated with different concentrations of doxycycline,after 48 h,RT-PCR results show that HSV-tk gene expression increased gradually with the increase of concentration of doxycycline,the expression level reached the peak in the doxycycline for 4 μg/ml.when added by doxycycline,the cell sensitivity to GCV was increased.Conclusion HSV-tk gene expression and antitumor function was successfully regulated used tetracycline in HeLa cells then provides experimental and theo-retical basis for further explore expression regulation of suicide gene during antitumor.