125I-神经生长因子对人脑胶质瘤细胞株U251的作用

目的 探讨125I-神经生长因子(125I-NGF)对U251细胞的杀伤能力及作用机制.方法 通过噻唑蓝(MTT)试验检测分别以18.5、37.0、74.0、148.0、296.0、592.0、1184.0、1480.0、1850.0、2590.0、3330.0、3700.0 kBq/ml 125I-NGF处理的U251的吸光度值,选择吸光度值趋于最低时最小125I-NGF浓度为其最低有效浓度.采用平板克隆形成试验,比较分别给予完全培养液、592 kBq/mlNa125I、1 mg/L NGF和592 kBq/ml 125I-NGF溶液处理的各组细胞克隆形成率,评价上述因素处理后U251细胞的增殖能力.采用放射自显影技术观察细胞内银颗粒分布,从而反映125I-NGF进入细胞的能力.并通过彗星试验及微核形成试验观察125I-NGF作用后U251细胞DNA的损伤,有明显彗尾及微核形成者DNA损伤严重.通过流式细胞仪检测125I-NGF作用后U251 G0/G1及S期细胞比例变化.结果 125I-NGF的最低有效浓度为592 kBq/ml,在此浓度下,125I-NGF可以有效进入细胞核内并损伤靶细胞DNA,经125I-NGF处理的胶质瘤细胞克隆形成率(0.02±0.01)较对照组(0.33±0.02)明显降低(P<0.01).125I-NGF作用后S期细胞比例(10.69±0.02)较对照组(35.47±0.02)下降,G0/G1期细胞比例(75.10±0.22)较对照组(53.17±0.03)升高(P<0.01).结论 125I-NGF在U251细胞中可以被转运入细胞核内,并可以有效杀伤靶细胞,其杀伤作用主要针对S期胶质瘤细胞.

Effect of 125I labeled nerve growth factor on glioma cell line U251

Objective To explore radiotoxicity of 125I labeled nerve growth factor (125I-NGF) in U251. Methods Absorbance values of U251 treated with 18.5, 37.0, 74.0, 148.0, 296.0, 592.0,1184. 0, 1480. 0, 1850. 0, 2590. 0, 3330. 0, 3700. 0 kBq/ml 125I-NGF were detected by MTT assay. The corresponding minimum concentration of 125I-NGF tending to the minimum absorbance value was selected as minimum effective concentration, and was adopted in follow-up experiments. Plate colony-forming assay was adopted to compare the proliferation ability of cells treated with complete culture medium, 592 kBq/ml Na125I, 1 mg/L NGF and 592 kBq/ml 125I-NGF. Distribution of silver particles within the cells was observed by autoradiography to respond the existence of 125 I-NGF inside the cells. DNA damage was evaluated by comet assay and micronuclei forming assay. The proportion of cells in G0/G1 or S p hase was determined by flow cytometry. Results Minimal effective concentration of 125I-NGF was 592. 0 kBq/ml. Under thisconcentration, 125I-NGF could be transported to the nuclei of U251 and caused DNA damage. Being treated with 125I-NGF, U251 clone forming rate was (0. 02 ± 0. 01 ), compared to control group (0. 33 ± 0. 02),the proliferation of U251 was inhibited remarkably (P <0. 01 ). The proportion of cells in S phase ( 10. 69± 0. 02) was decreased, but that in Go/G1 phase ( 75. 10 ± 0. 22 ) increased remarkably ( P < 0. 01 ).Conclusion After being iodinated with 125I,125I-NGF can be transported to nuclei and causes DNA damage in U251. 125I-NGF has specific lethal effect on U251 cells in S phase.