Objective Construct a recombinant adenovirus containing the human hydroperoxidase( hT-rnPO )gene, transfer the hTPO genes and human sodium/iodide symporter( hNIS )gene into glioma cells and study rnthe iodide uptake ability of the cells and the killing effect of radioiodine , so as to investigate the feasibility of rarndioiodine therapy on glioma. Methods The recombinant adenovirus AdTPO was constructed by AdEasy system.rnTranfecting hNIS gene into human glioma cell line U251 through recombinant plasmid.the stably expressing hNIS rngene cell line hNIS - U251 was determined as negative control group. Then hTPO gene was transfected into hNIS rn- U251 by recomhinant adenovirus AdTPO, achieving AdTPO - hNIS - U251 , determined as testing group. U251 rncell line without hNIS or hTPO gene was applied as control group. We investigated the iodide uptake assay, perrnchlorate suppressive assay , iodide organification assay to confirm gene expression and function. Killing effect of rarndioiodide on tumor cells was studied by clonogenic assay in vitro. Results Recombinant adenovirus AdTPO was rncorrectly constructed. we were successful in co - transfecting hTPO gene into stably expressing hNIS cell line rn( hNIS - U251 ),obtaining AdTPO - hNIS - U251. In hNIS - U251 group, the iodide uptake ability( 55769. 96 ±rn4353. 26 count · min -1 )was 110 folds higher than that in control group( 507. 67 ±57. 69 count · min -1 ).the effective half - life time was 7 minutes,the organification percentage was 0. 1％,and the rate of clonal forming was rn( 9. 08 ±2. 86 )％ , that was 10 folds lower compared with control group. While in AdTPO - hNIS - U251 group,rnthe iodide uptake ability( 74647. 53 ±3605. 88 count · min -1 )was 147 folds higher than that in control group, the rneffective half - life time prolonged to 13 minutes, the organification percentage reached to 10％ , and the rate of rnclonal forming was( 6. 80 ±2. 09 )％ ,which was 13 folds lower than control group respectively. Conclusion By rnco - transfection of hTPO and hNIS genes into glioma cell line U251 ,the iodide uptake ability increased markedrnly, hTPO increased the retension of iodide in the cells, and the 131Ⅰ could kill tumor cells effectively.%目的 构建含有人甲状腺过氧化物酶基因(hTPO)的重组腺病毒,并与人钠-碘转运体基因(hNIS)共转染至神经胶质瘤细胞中,研究其摄碘能力及对肿瘤细胞的杀伤能力,探讨131I治疗胶质瘤的可能性.方法 应用AdEasy系统构建重组腺病毒AdTPO,利用重组质粒将hNIS基因转染入神经胶质瘤细胞系U251中获得hNIS-U251细胞系作为阴性对照组,再利用重组腺病毒AdTPO将TPO基因转染入hNIS-U251细胞系中获得AdTPO-hNIS-U251作为实验组,未转入hTPO和hNIS的细胞U251作为空白对照组.研究三组细胞的摄碘实验、过氯酸盐抑制实验、有机化测定实验检测其摄碘功能,细胞克隆形成实验评价131I对转染肿瘤的杀伤作用.结果 成功构建出重组腺病毒AdTPO,并在稳定表达hNIS的U251细胞中实现了hTPO的共转染.hNIS-U251组(每分钟放射性计数55769.96±4353.26)比空白对照组(每分钟放射性计数507.67±57.69)摄碘能力增高约110倍,有效半衰期7分钟,有机化程度约为0.1%,细胞克隆形成率(9.08±2.86)%,较对照组减低约10倍.AdTPO-hNIS-U251组(每分钟放射性计数74647.53±3605.88)比空白对照组摄碘能力增高约147倍,有效半衰期延长至13分钟,有机化程度增高至10%,细胞克隆形成率(6.80±2.09)%,较对照组减低约13倍.结论 将hTPO和hNIS共转染至神经胶质瘤细胞后,可有效提高细胞的摄碘能力,hTPO延长了放射性碘在细胞中的停留时间,131I对瘤细胞有较强的杀伤作用.
展开▼
