AG490对HEL细胞VEGF和HIF-1α表达的影响

 目的: 探讨JAK2抑制剂AG490对人红白血病(HEL)细胞迁移及对VEGF和HIF-1α表达的影响。方法: 用不同浓度的AG490处理HEL细胞,CCK-8法检测细胞活力,Hoechst 33342荧光染色检测细胞凋亡,流式细胞术检测细胞凋亡及周期,Transwell小室检测细胞迁移能力,RT-PCR检测JAK2的mRNA水平,Western blot检测p-JAK2、VEGF和HIF-1α的蛋白水平。结果: AG490能够抑制HEL细胞的活力,不同浓度(20、40、60、80和100 μmol/L)AG490作用HEL细胞48 h后,细胞活力分别为88%、75%、48%、10%和0.12%(P<0.05);Hoechst 33342凋亡细胞染色显示80 μmol/L AG490处理细胞48 h后,亮蓝色凋亡细胞较对照组明显增多(P<0.05);流式结果显示80 μmol/L AG490作用细胞48 h后,凋亡率上升;细胞迁移实验结果显示20 μmol/L AG490处理细胞24 h后漏出细胞明显低于对照组(P<0.05);RT-PCR结果显示不同浓度AG490处理HEL细胞48 h后JAK2 mRNA呈剂量依赖性减低;Western blot结果显示实验组细胞的p-JAK2、VEGF和HIF-1α蛋白水平较对照组明显减低(P<0.05)。结论: AG490可通过抑制JAK2信号通路,抑制HEL细胞血管新生因子VEGF和HIF-1α的表达。

Effect of AG490 on expression of VEGF and HIF-1α in HEL cells

AIM: To investigate the effect of AG490 on the expression of VEGF and HIF-1α, and the capacity of invasion in human erythroleukemia (HEL) cells. METHODS: The HEL cells were treated with AG490 at different concentrations. The cell viability was detected by CCK-8 assay. The apoptosis was detected by Hoechst staining. The apoptosis and the cell cycle were analyzed by flow cytometry. The capacity of migration was evaluated by Transwell assay. The mRNA expression level of JAK2 was measured by RT-PCR. The protein levels of p-JAK2, VEGF and HIF-1α were determined by Western blot. RESULTS: The HEL cell viabilities were 88%, 75%, 48%, 10% and 0.12% after treated with AG490 at 20, 40, 60, 80 and 100 μmol/L for 48 h, respectively. The results of Hoechst staining showed that brilliant blue cells in 80 μmol/L AG490 group was significantly increased compared with control group for 48 h. The apoptosis rate of 80 μmol/L AG490 group was significantly increased compared with control group at 48 h after AG490 treatment. The number of membrane-permeating HEL cells in 20 μmol/L AG490 group at 24 h after AG490 treatment was significantly lower than that in control group (P<0.05). The mRNA level of JAK2 decreased in a concentration-dependent manner after the HEL cells were treated with different concentrations of AG490 for 48 h. The protein levels of p-JAK2, VEGF and HIF-1α were lower in AG490 treatment groups than those in control group (P<0.05). CONCLUSION: AG490 inhibits the expression of VEGF and HIF-1α in HEL cells by inhibiting JAK2 pathway.