视网膜母细胞瘤基因对人骨肉瘤细胞株OS732的影响

目的 观察外源性N端截短的视网膜母细胞瘤（Rb)基因对骨肉瘤细胞株OS732的生长和细胞间缝隙连接信息通讯能力的影响。方法 构建N端截短的长度为1.65kb Rb基因的真核表达质粒,并用DOTAP将其导入骨肉瘤细胞株OS732。应用逆转录－聚合酶链反应（RT－PCR）和Northern blot检测Rb基因的表达;用细胞计数,流式细胞仪分析和软琼脂克隆形成试验观察OS732细胞的生成状况;用半定量RT－PCR法和罗氏黄荧光传输法检测细胞间缝隙连接信息通讯的能力。结果 转染N端截短的Rb基因后,OS732细胞内可检测到内源性和外源性Rb基因的mRNA表达。OS732细胞的形态发生改变,其生长速度减慢,软琼脂形成集落能力降低,细胞周期阻滞于G0－G1期;缝隙连接蛋白基因Gonnexin43的表达和细胞信息通讯能力增强。结论 N端截短的Rb基因可抑制骨肉瘤细胞株OS732的恶性生长表型以及增强细胞间信息通讯能力。

Effect of retinoblastoma gene on human osteosarcoma cell line OS732

Objective To investigate the effect of growth suppression and gap junctional intercellular communication (GJIC) on osteosarcoma cell line OS732 transduced with the exogenous N terminal truncated retinoblastoma (Rb) gene Methods Recombinant eukaryotic expression plasmid harbouring 1 65 kb Rb gene was constructed and transduced into OS732 cells Rb mRNA expression was detected by RT PCR and Northern blot The cell count, cell cycle analysis and soft agar colony formation test were used to observe growth features RT PCR and Lucifer yellow dye transfer were carried out to measure GJIC Results Both exogenous and endogenous Rb gene could be detected in OS732 cell transduced with the N terminal truncated Rb gene Compared with the parental cells, change in morphology of Rb transduced cells were found; growth rate decreased; amount of colony formation reduced; cell cycle arrested at G 0/G 1 checkpoint; Connexin43 expression and GJIC enhanced Conclusions The malignant phenotype of OS732 is inhibited and GJIC is increased by transduction of N terminal truncated Rb gene