IFN-γ增强TNF相关凋亡诱导配体对骨巨细胞瘤细胞的抑制

IFN-γ enhances the anti-tumor effect of TRAIL against giant cell tumor of bone in vitro

目的:探讨肿瘤坏死因子相关凋亡诱导配体(tumor necrosis factor-related apoptosis-inducing ligand,TRAIL)对人骨巨细胞瘤(giant cell tumor of bone,GCT)的作用及干扰素-γ(interferon-γ,IFN-γ)对TRAIL抗瘤活性的影响,并探讨其影响机制。方法:选取2003年-2006年于301医院骨科手术切除的骨巨细胞瘤新鲜标本9例,将取自标本的骨巨细胞瘤细胞分为:对照组,TRAIL组,IFN-γ组,IFN-γ诱导后加入TRAIL组,应用CCK-8法检测TRAIL、IFN-γ单独及联合作用对骨巨细胞瘤细胞的增殖抑制作用,流式细胞仪、TUNEL法分析IFN-γ对TRAIL凋亡诱导作用的影响,RT-PCR法检测IFN-γ作用前后肿瘤细胞DR4、DR5、TRAIL的表达。结果:骨巨细胞瘤细胞对照组存活率为(98.64±0.31)%,TRAIL(100、500、1000μg/L)单独作用对肿瘤细胞增殖无明显抑制作用;IFN-γ(500、1000U/ml)单独作用后,细胞的存活率分别为(94.05±1.89)%、(90.47±2.66)%。IFN-γ(500U/ml)诱导24h后与TRAIL(100、200μg/L)联合作用,细胞的存活率分别为(84.65±2.46)%、(77.65±3.14)%。两者联合作用后流式细胞仪测得联合作用组凋亡率为(27.94±2.88)%、(38.65±2.46)%,对照组凋亡率为(1.77±0.49)%;TUNEL法测得联合作用组凋亡率分別为(19.63±3.51)%、(29.28±4.80)%,对照组的凋亡率为(1.1±0.17)%。RT-PCR结果显示,IFN-γ作用24h后肿瘤细胞TRAIL、DR4、DR5 mRNA表达明显上调。结论:骨巨细胞瘤细胞对TRAIL不敏感,IFN-γ可以明显提高TRAIL诱导骨巨细胞瘤细胞凋亡的敏感性,其机制可能与IFN-γ上调TRAIL、DR4、DR5 mRNA的表达有关。

Objective：To explore the anti-tumor effect of tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） against giant cell tumor of bone （GCT）, and to study the influence of IFN-γ on the anti-tumor effects of TRAIL and the possible mechanisms. Methods：Fresh specimens were obtained from 9 patients who were pathologically diagnosed as having GCT in the orthopaedics department of No. 301 Hospital PLA from 2003 to 2006. The GCT cells from specimens were divided into 4 groups： control group, TRAIL group, IFN-γ group and IFN-γ plus TRAIL group. The cell inhibition was examined by CCK-8 assay. Cell apoptosis was detected by flow cytometric analysis and TUNEL assays. RT-PCR was applied to semi-quantitatively assay the mRNA expression of TRAIL, DR4 and DR5 in GCT cells before and after the treatment with IFN-γ （500 U/ml）. Results： The GCT cell survival rate was （98.64±0.31）% in the control group. TRAIL alone （100, 500,1 000 μg/L） had no obvious effect on tumor cell proliferation. After treatment with IF N-γ at 500 and 1 000 U/ml, the tumor cell survival rates were （94.05±1.89） % and （90.47±2.66） %, respectively. After treatement with IFN-γ（500 U /ml） and TRAIL （100,200 μg/L）, the tumor cell survival rates were （84.65±2.46）% and （77.65±3.14）%, respectively; and FCM showed that their apoptosis rates were （27.94±2.88）% and （38.65±2.46）%, respectively; the apoptosis rate was （1.77±0.49）% in the control group. TUNEL revealed that the apoptosis rates were （19.63±3.51）% and （29.28±4.80）% , respectively; and the rate was （1.1±0.17）% in the control group. RT-PCR showed that the expression levels of TRAIL, DR4 and DR5 in tumor cells were up-regulated after treatment with IFN-γ （500 U /ml） for 24 h. Conclusion： GCT cells are not sensitive to TRAIL. IFN-γ can up-regulate the apoptosis rate of GCT cells induced by TRAIL, which might be associated with the up-regulation of TRAIL,DR4 and DR5 mRNA expression by IFN-γ.