The Effects Of Proteasome Inhibitor Bortezomib On The Growth And Apoptosis Of K562 Cells In The Presence Of Bone Marrow Mesenchymal Stem Cells

ObjectiveTo investigate the effects of proteasome inhibitor bortezomib on the cell cycle profile and apoptosis of K562 in the presence of bone marrow mesenchymal stem cells. The expression of adhesion molecule VCAM-1 and proangiogenic IL-8 was also detected to investigate whether the anti-leukemia effects of bortezomib were in part through effects on bone marrow microenvironment.MethodsK562 cells were co-cultured in direct contact with MSCs while the control were just cultured alone. Bortezomib was administered at a final concentration of 50nmol/L. Growth curves of K562 cells were drawn and flow cytometry was used to detect the changes of cell cycle distribution. Cell apoptosis was assayed using Annexin-V/PI apoptosis detection kit by flow cytometry. The VCAM-1 and IL-8 gene expression was determined by reverse transcription polymerase chain reaction (RT-PCR).ResultsBortezomib induced cell cycle arrest at G2/M phase of K562 cells cultured alone in a time-dependent manner. A higher percentage of G0/G1 phase was observed in K562 cells co-cultured with MSCs than K562 cells cultured alone in the presence of bortezomib for 8 hours. But the G0/G1 arrest of K562 cells caused by co-cultured with MSCs was abrogated when bortezomib treatment was extended to 24 hours.Bortezomib induced apoptosis of K562 cells as early as 4 hours after treatment. The ratio of apoptotic cells was about 25.8±2.1% when treated for 12 hours. As the bortezomib treatment extended to 24 hours, there was a slightly increase in the ratio of apoptotic cells, but it had no statistic significance compared with that of 12 hours. K562 cells grown on the layer of MSCs demonstrated the similar sensitivity to apoptosis induction of bortezomib.K562 cells which did not express VCAM-1 originally were induced to express VCAM-1 mRNA when co-cultured with MSCs. This effect can be abrogated by bortezomib treatment. Furthermore, bortezomib significantly downregulated the VCAM-1 expression of MSCs .Co-culture of K562 cells in direct contact with MSCs resulted in a significant increase in IL-8 expression of both K562 and MSCs. Bortezomib treatment downregulated the expression of IL-8 in K562 cells , but no changes were observed in MSCs.ConclusionMSCs were able to influence the proliferation and apoptosis of K562 cells by direct cell-to-cell contact. The proteasome inhibitor bortezomib can abrogate the G0/G1 arrest of K562 cells caused by co-cultured with MSCs and induce apoptosis of K562 cells even though with the MSCs layer support. In this study, we demonstrate that bortezomib both acts on leukemia cells and MSCs to induce apoptosis and overcome drug resistance. It directly induces apoptosis of human leukemic cell line K562, and also inhibits the paracrine growth of leukemic cells by decreasing their adherence to MSCs and downregulating the related induction of proangiogenic IL-8 secretion.