| Objective:This study was aimed at identifying a correlation between the abnormal expression of histone deacetylases(HDACs)and HO-1 in multiple myeloma(MM).Moreover,we assessed the effect of BG45,a novel class I HDAC inhibitor,on the signaling pathway of histone deacetylases(HDACs)inhibition induced apoptosis,as well as its possible mechanism,to determine whether BG45 is a potential candidate for the treatment of MM.Methods:Bone marrow blood samples from MM patients and healthy donors were collected and staged according to the International Staging System(ISS).The expression of HO-1 and HDAC3 mRNA in CD138~+cells were detected by real-time PCR.To determine the level of acetylation and HO-1 on MM patients and cells,the expression of HO-1 and HDACs proteins in MM patients?MM cells and healthy donors were detected by western blot.After being exposed to different concentrations of BG45 for 48h or 72h,cell viability was detected by CCK8.,apoptosis induction was analyzed by flow cytometry,cell migration rate was assessed by transwell assay,and the protein and mRNA expressions of HO-1?HDAC1?HDAC2 and HDAC3 were detected by western blot and real-time PCR.To compare the effects of BG45 with those of LBH589,MM cells were incubated with either BG45 or LBH589,and the expression of HO-1 was detected by western blot.To examine the effect of BG45 on cell cycle and apoptosis,MM cells were incubated with increasing concentrations of BG45 and the changes in cell cycle were determined by flow cytometry,the cycle-associated proteins and the correlation of the mitochondrial apoptosis pathway that were detected by western blot.Moreover,changes in the JAK2/STAT3 pathway was detected by western blot.Finally,the combination of BG45 with lenalidomide was tested in MM cells,the apoptosis rate was detected by flow cytometry,cell viability were detected by the CCK8 assay.Results:The mRNA expression levels of HDAC3 and HO-1 in CD138~+cells from MM patients were significantly higher than those of healthy donors,and followed a descending order of stage III>stage II>stage I(P<0.05),The protein expression levels of HDACs and HO-1 expression in MM patients were significantly higher than those in healthy donors.BG45 significantly inhibited the survival of MM cells(U266,RPMI8226,CD138~+cells)in time-and dose-dependent manners,whereas cells from healthy donors remained unaffected.MM cells were significantly more prone to apoptosis,as measured by flow cytometry,transwell assays showed that BG45 inhibited RPMI8226 cell migration.The transcriptional and protein levels of class I HDACs and HO-1 were strongly down-regulated in the BG45-treated MM cell lines,increased histone H3 acetylation levels.The expression of the antiapoptotic BCL-2 and Bcl-xl protein was strongly decreased and that of Bax protein was increased by BG45 treatment,which also induced cleavage of PARP.Results also revealed less HO-1 was expressed in the MM cells treated with BG45 than in those treated by LBH589,BG45 can potentially be used to treat MM patients with high HO-1 expression.Moreover,BG45 increased the accumulation of U266 cells in the G0/G1 phase,and downregulated the expression of cell cycle-associated proteins.Reduced JAK2/STAT3 activation may enhance MM cell apoptosis and HO-1affected the JAK2/STAT3 pathway.Finally,BG45 synergized with lenalidomide in MM cells were confirmed by flow cytometry and CCK8.Conclusion:BG45downmodulated HO-1 when class I HDACs decreased in MM cells,activated the mitochondrial apoptotic pathway,induced cleavage of PARP,arrested cells in the G0/G1 phase,inhibited the JAK2/STAT3 pathway and strongly induced apoptosis of MM cells,whose effect is related to HO-1.BG45 synergized with lenalidomide in MM cells,test that the correlation between HDACs and HO-1 in MM was found,HO-1may be a potential HDACi therapeutic target,BG45 may be applicable to the treatment of MM and other hematological malignancies... |
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