The Proliferation Inhibition Effect Of Timolol On Human Multiple Myeloma Cells RPMI 8226 And Its Molecular Mechanism

Objective: To explore the effects of Timolol on proliferation and apoptosis of MM cell line RPMI 8226, and find its potential mechanism, so as to provide evidence for the clinical treatment of multiple myeloma. Methods: The proliferation inhibition effect of Timolol on RPMI 8226 cells was tested by MTT and the concentration for further research were screened out. Flow cytometry was used to detect the apoptosis of RPMI 8226 cells. The expression of caspase-3, caspase-9, Bax and Bcl-2 in transcription and translation level were measured by qRT-PCR and Western Blot respectively. The obtained data was then statistically analyzed. Results:After treatment with Timolol for 24 h, 48 h and 72 h, the viability of RPMI 8226 cells were inhibited, and the inhibition rate increased with the increase of drug concentration under the same time, which with obvious concentration dependence(r=0.548, P=0.019). At the same time, under the same drug concentration, the inhibition rate increased with the extension of the time, which with a significant time dependence(r=0.659, P=0.003). In summary, Timolol could effectively inhibit the proliferation of RPMI 8226 cells in dose- and time- dependent manner. After treatment with Timolol for 72 h, the apoptotic rate of 2.5μmol/L, 20μmol/L, 50μmol/L were(7.17±0.06)%,(10.56±0.65)% and(15.27±0.86)% respectively, which were much higher than that in control group(3.93±0.25)%. The difference was statistically significance(P<0.01). And the apoptotic rate of RPMI 8226 cells was increased with the increasing concentration of Timolol, which was concentration dependent(r=0.953,P=0.047). The results of qRT-PCR showed that, after treated by Timolol for 72 h, the expression of Bax, caspase-3 and casepase-9 mRNA were significantly up-regulatedand that of Bcl-2 was decreased significantly(P<0. 05). Western blot analysis showed that compared with the control group, after treated with 20μmol /L Timolol for 72 h,the expression of Bax protein was increased significantly and that of Bcl-2 was significantly decreased, the difference was statistically significance(P<0.05).However, there were no obvious change at the protein level of caspase-3 and caspase-9 compared with the control group. Conclusion: Timolol has proliferation inhibition and apoptosis induction effect on multiple myeloma cells, the mechanism may be through up regulation of Bax expression and down regulating Bcl-2expression to induce cell apoptosis.