人支气管上皮细胞DNA聚合酶β基因RNA干扰后细胞生物学性状分析

An analysis on biological characteristics of DNA polymerase β gene in human bronchial epithelial cells with RNA interference

目的探讨人支气管上皮细胞(HBE)DNA聚合酶β(polβ)基因RNA干扰后的生物学性状变化。方法实验设实验组、空载体组、正常对照组、阳性对照组;运用载体介导的RNA干扰技术抑制polβ基因在HBE中的表达,在光学显微镜下观察转染重组子的实验组细胞及正常对照组细胞的形态学特征;绘制细胞生长曲线;分析细胞倍增时间;运用流式细胞技术分析细胞周期;以Hella细胞作阳性对照,采用软琼脂糖恶性程度鉴定实验分析细胞的恶性程度。结果光镜下可见实验组细胞内颗粒物明显增多,体积稍有增大;3组细胞的生长曲线均呈现相似的S型;实验组、空载体组及正常细胞组的倍增时间相近,分别为0.81、0.84和0.79d;3组细胞的G1期(83.9%、81.0%、84.6%)、G2期(4.9%、5.5%、6.2%)和S期(11.2%、13.5%、9.2%)分布相近;实验组细胞未见在双层软琼脂糖中生长出细胞集落,而阳性对照Hella细胞集落明显。结论 polβ基因缺陷对HBE生物学性状的短期影响不明显。

Objective To explore the biological characteristic of DNA polymerase β （pol β） gene in human bronchial epithelial cells （HBE） with RNA interference （RNAi）.Methods Experimental group,empty vector group,normal control group and positive control group were included.Technology of vector-mediated RNAi was used to inhibit the expression of pol β in HBE.Morphological characteristics of each group were analyzed under light microscope.Growth curve of each group was drawn and doubling generation time was analyzed.Technology of flow cytomertry （FCM） was used to study the cell cycle.Malignancy degree of each group was analyzed by soft agarose experiment.Results In the experimental group,the numbers of cell granulations were increased transparently and cell volume accretion was found,the rest characteristics were common.Growth curve of each group presented similar shape of ＂S＂.The doubling generation time of the experimental group,empty vector group,normal control group were 0.81,0.84 and 0.79 days respectively,while the cell cycle distribution of G1（83.9%,81.0%,84.6%）,G2（4.9%,5.5%,6.2%） and S （11.2%,13.5%,9.2%） in the above mentioned groups appeared similar.In soft agarose experiment,cell colony was not found in the experimental group,but found in the positive control group.Conclusion DNA pol β knock-down might not cause transparent short-term influence on biological characteristic of human bronchial epithelial cells.