硼替佐米增强P16蛋白表达诱导骨髓瘤细胞衰老

Bortezomib Induced Myeloma Cells Senescence by Enhancing the Expression of P16 Protein

目的:研究蛋白酶体抑制剂硼替佐米诱导骨髓瘤RPMI8226、MMH929细胞衰老作用,并进一步探讨其作用机制。方法:硼替佐米0.1-100nmol/L处理骨髓瘤RPMI8226、MMH929细胞48、72h,MTT法检测细胞存活率、药物IC50值。选择药物IC50值1/10剂量处理骨髓瘤RPMI8226、MMH929细胞0、24、48H后检测衰老相关β-半乳糖苷酶染色率。流式细胞术检测细胞周期情况及凋亡率。Western-blot检测相关蛋白表达。结果:硼替佐米处理骨髓瘤细胞RPMI8226、MMH929后48小时IC50值:RPMI8226:19.05 nmol/L,MMH929:18.45nmol/L。以硼替佐米2 nmol/L处理骨髓瘤RPMI8226、MMH929细胞0、24、48H后发现β-半乳糖苷酶染色率、细胞G0/G1期比例明显上升与药物作用时间呈正相关,Western-blot检测细胞周期调控蛋白发现P53、PTEN蛋白无变化,P16蛋白与药物作用时间正相关。结论:硼替佐米通过增强P16蛋白表达诱导骨髓瘤细胞RPMI8226、H929衰老。

Objective： To study the proteasome inhibitor bortezomib induced myeloma RPMI8226,MMH929 cell senescence,and further explore its mechanism.Methods： Myeloma RPMI8226,MMH929 cell were routinely cultured and treated with different concentrations of bortezomib（1-500 nmol/L） for 48-72 h.Viability was determined by MTT assay.The drug of choice IC50 values of 1/10 dose treatment RPMI8226,MMH929 cell 0,24,48 H.Apply PI staining method to test cell cycle by flow cytometry.Apply SA-β galactosidase staining method to test the senescence of RPMI8226,MMH929 cell induced by bortezomib.The protein levels of P53,PTEN,P16 were determined by Western bolt.Results： Bortezomib treatment of myeloma cells RPMI8226,H929 48-hour IC50 values： RPMI8226： 19.05nmol / L,H929： 18.45nmol / L.Treatment with Bortezomib 2 nmol / L after 0,24,48 H,indicating β-galactosidase staining rate,proportion of cells G0/G1 is up-regulated.The protein level of P16 was up-regulated while that of P53,PTEN did not change by Western blot.Conclusion： Bortezomib induced myeloma cells RPMI8226,H929 senescence by enhancing the expression of P16 protein.