摘要
目的:探讨PFT-α(p-fiftythreeinhibitor,PFT-α)对热化疗诱导原代培养肠上皮细胞(intestinalepithelialcells,IECs)凋亡及其凋亡相关基因表达的影响.方法:原代培养IECs分为正常对照组、热化疗组和PFT-α+热化疗组,运用顺铂联合温热(43℃)处理IEEs30min,对比加入不同浓度的PFT-α后,AnnexinV-FITC/PI染色,流式细胞仪检测细胞凋亡.Westernblot检测IECs的p53和Bax蛋白表达,RT-PCR检测BaxmRNA的表达.结果:热化疗导致IECs发生凋亡,Bax蛋白表达明显高于对照组.10,20,30,40μmol/LPFT-α作用于顺铂联合温热处理的IECs后,细胞凋亡率下降且呈剂量依赖陛,p53在IECs胞核/胞质表达比例下降,Bax蛋白和mRNA的表达随PFT-α的剂量升高而逐渐下降.结论:PFT-α热化疗诱导肠上皮细胞凋亡具有抑制作用,其机制可能与改变p53核转位和抑制促凋亡基因Bax的表达相关.
AIM: To investigate the effect of PFT-α (p53 inhibitor, PFT-α) on apoptosis and apoptosis-associated gene expression in intestinal epithelial cells (IECs) induced by hy perthermic chemotherapy. METHODS: Primary cultured IECs were divided into control (CON), hyperthermic chemotherapy (HTC) and PFT-α plus hyperthermic chemotherapy group (PFT-α, HTC). IECs in HTC group were treated with hyperthermic (43 ℃) cisplatin (terminal concentration 10 mg/L) for 30 min. IECs in PFT-α, HTC group were treated with different dosages (10, 20, 30, 40 μmol/L) of PFT-α 4 h before treated the same as those in HTC group. The rate of apoptosis was detected using flow cytometry after double staining of annexin V-FITC and PI. The expression of p53 and Bax protein were detected by Western blotting, and the level of bax mRNA expression was detected using semi-quantitative RT-PCR. RESULTS: PFT-α reduced apoptotic rate of IECs in a dose dependant manner, from (14.83 ± 1.51%) at the concentration of 10 μmol/L to (3.75 ± 0.33%) at 40 μmol/L, which were significantly lower than the rate in HCT group(27.03 ± 2.07%, P <0.01). The ratio of p53 in the nucleus and cytoplasm was 13.22 :1 in HCT group. However, the ratio in PFT-α, HTC group decreased from 0.8 : 1 at the concentration of 10 μmol/L to 0.4 :1 at 40 μmol/L, which were significantly lower than that in HCT group. Hyper thermic chemotherapy upregulated the levels of bax mRNA (128.85 ± 2.38) and protein (14.29 ± 0.43) in lECs, whereas PFT-α significantly reduced (P <0.01) those levels in a dose-dependant manner, from 106.98 ± 3.75 (mRNA) and 5.79 ± 0.24 (protein) at the concentration of 10 μmol/L to 4.79 ± 0.31 (mRNA) and 1.36 ± 0.09 (protein) at 40 μmol/L respectively. CONCLUSION: PFT-α effectively inhibits apoptosis of lECs induced by hyperthermic chemotherapy. The mechanisms may relate to its effect on changing the nuclear transloca tion of p53and inhibiting the expression of bax gene.
出处
《世界华人消化杂志》
CAS
2004年第10期2353-2355,共3页
World Chinese Journal of Digestology
