前列腺癌细胞DU145同步化诱导的DNA损伤反应通路和PI3K/Akt通路对凋亡抑制因子基因API2(BIRC3)mRNA表达的影响

目的探讨在前列腺癌细胞DU145同步化培养后引起的DNA损伤反应通路和PI3K/AKT通路对凋亡抑制因子(IAP)基因API2(BIRC3)mRNA表达的影响,同时分析API2基因所在染色体是否存在特异性的异常。方法采用无血清的饥饿培养法使细胞同步在G0期;分别用含羞草碱(mimosine)、胸腺嘧啶(thymidine)和噻氨酯哒唑(nocodazole)使细胞同步在G1期、S期和G2/M期并引起DNA损伤反应,同时加入PI3K的特异性抑制剂ly294002以阻止PI3K/AKT通路。通过RT-PCR半定量法检测API2 mRNA在各个细胞周期相的表达情况。通过细胞遗传学的常规G式显带法,分析凋亡抑制因子基因所在的染色体是否存在特异性的异常。结果 mimosine同步化的G0/G1期细胞达到了78.04%,thymidine同步化的S期细胞达到62.19%,nocodazole同步化的G2/M 60.5%。API2基因位于染色体11q22-q23,存在易位,如t(11;12)(q;q)。API2mRNA的表达,非同步化的ly294002组分别与同步化的mimosine+ly294002组、nocodazole+ly294002组和thymidine+ly294002组比较,差异均有统计学意义(P<0.05)。结论前列腺癌细胞株DU145存在某些染色体的结构和数目异常,这些异常可能影响某些基因的表达。药物的同步化激活了DNA损伤反应通路和生存信号通路,再通过PI3K/Akt通路,而不是通过P53通路,在细胞周期的某个时相调控API2(BIRC3)mRNA的表达。

Study on the Effect of DNA Damage Response Pathway and PI3K/Akt Pathway Induced by Synchronous Culture of Prostate Cancer Cell DU145 on the mRNA Expression of Apoptosis Inhibitor Gene API2 (BIRC3)

Objective To study on the effect of DNA damage response pathway and PI3K/Akt pathway induced by syn- chronous culture of prostate cancer cell DU145 on the rnRNA expression of apoptosis inhibitor gene API2 （BIRC3）, and to ex- plore the specific abnormality of the chromosome in which API2（BIRC3） is located. Methods The cells were synchronized in CA3 phase by starve culture method. Mimosine, thymidine and nocodazole were used respectively to obtain G1, S and G2/M phase cells, and simultaneously, PI3K/AKT pathway was blocked by a specific inhibitor of PI3K, ly294002. The expression of API2 mRNA at different cell cycle phases was detected by semi - quantitative RT - PCR. The specific abnormality of the chro- mosome in which API2 （BIRC3） was located was analyzed by the routine G- type banding method in cytogenetics. Results The G0/G1 phase cells synchronized by mimosine accounted for 78.04%. The S phase cells synchronized by thymidine were 62.19% ,and the G2/M phase cells synchronized by nocodazole were 60.50%. API2 gene was located in the chromosome 11@2 -q23, which was found to have translocation like t（11;12） （q; q）. Compared respectively with the synchronous cell groups of mimosine ＋ ly294002, nocodazole ＋ ly294002 and thymidine ＋ ly294002, the expression level of API2 mRNA of the non - syn- chronous cell group of ly294002 was significantly different （P 〈 0.05）. Conclusions Some chromosomes in the prostate cancer cell DU145 have abnormal structure or number, which may influence the expression of some genes. Synchronization by the drugs triggers the DNA damage response pathway and survival signal pathway, which is in turn through PI3K/Akt pathway, other than p53 pathway, to control the expression of API2（BIRC3） mRNA at some cell phase.