HIF-1α调控4-羟基他莫昔芬对乳腺癌MCF-7细胞敏感性的研究

Study on HIF-1α regulating the sensitivity of 4-hydroxytamoxifen to breast cancer MCF-7 cells

目的该研究通过建立人乳腺癌4-羟基他莫昔芬(4-OHT)耐药细胞系(MCF-7/TR),探究低氧诱导因子1α(HIF-1α)对乳腺癌MCF-7细胞4-OHT耐药的影响。方法采用他莫昔芬的体内活性形式4-OHT建立乳腺癌耐药细胞MCF-7/TR。MTT实验检测4-OHT对乳腺癌细胞MCF-7、MCF-7/TR细胞活力的影响,并测定耐药指数(RI);Western blot检测MCF-7、MCF-7/TR细胞中HIF-1α蛋白的表达水平;采用MTT法和流式细胞术AV/PI双染法检测,HIF-1α抑制剂(LW6)或siRNA HIF-1α干预作用后4-OHT对MCF-7/TR细胞的影响以及HIF-1α稳定剂(FG-4592)处理后4-OHT对MCF-7细胞的影响。结果结果显示,MCF-7/TR耐药细胞株成功构建,其RI为(5.56±0.80);与MCF-7细胞相比,MCF-7/TR细胞中HIF-1α高表达,且4-OHT作用后MCF-7/TR细胞中HIF-1α的表达水平上调;给予LW6或沉默HIF-1α的表达后,HIF-1α的表达下调增强了4-OHT对MCF-7/TR细胞的抑制作用;给予FG-4592后,HIF-1α的表达上调可降低4-OHT对MCF-7细胞的抑制作用。结论HIF-1α在乳腺癌MCF-7细胞4-OHT耐药中发挥重要作用,靶向HIF-1α可能是增加乳腺癌细胞对他莫昔芬敏感性的有效途径。

Objective To explore the effect of hypoxia inducible factor 1α(HIF-1α)on tamoxifen resistance in breast cancer MCF-7,and to established a human breast cancer cell line(MCF-7/TR)with 4-hydroxy tamoxifen(4-OHT)resistance.Methods 4-OHT was an active form of tamoxifen in vivo,resistant breast cancer cells MCF-7/TR were established in vitro using 4-OHT.MTT assay was used to detect the effect of 4-OHT on the MCF-7 and MCF-7/TR cells,and the drug resistance multiple was detected.Western blot was used to detect the expression level of HIF-1αprotein in MCF-7 and MCF-7/TR cells.The effects of HIF-1αinhibitor(LW6)or siRNA HIF-1αon MCF-7/TR cells and 4-OHT on MCF-7 cells treated with HIF-1αstabilizer(FG-4592)were detected by MTT and flow cytometry AV/PI staining.Results The results showed that the MCF-7/TR was successfully constructed,and the drug resistance ratio was(5.56±0.80).Compared with MCF-7 cells,MCF-7/TR cells had higher expression of HIF-1αprotein and it was up-regulated after tamoxifen treatment.After giving LW6 or silencing the expression of HIF-1α,the down-regulation of HIF-1αexpression enhanced the inhibitory effect of 4-OHT on MCF-7/TR cells.After treatment of FG-4592,the expression level of HIF-1αin breast cancer cells MCF-7 was up-regulated,and hindered the inhibition effect of tamoxifen on MCF-7 cells.Conclusion The above results indicate that HIF-1αplays an important role in 4-OHT resistant breast cancer,and targeting HIF-1αmay be an effective way to increase the sensitivity of MCF7/TR cells to tamoxifen.