The Study Of Upregulation Of Killing Sensitivity Of Multiply Myeloma U266Cells Treated With Bortezomib To γδT Cells

To study the role of Bor in U266cells, cell proliferation was detected by MTT method; Cytotoxicity of γδT cells aganist U266cells was measured by calcein assay before and after treated by Bor; Cell apoptosis were analyzed by AnnexinⅤ-FITC assay; The expressions of NKG2D ligands(ULBP1, ULBP2, ULBP3, MICA/B) and TRAIL (DR4,DR5) on the surface of U266cells before and after treatment with Bor were detected by flow cytometry (Flow cytometry, FCM).The study shows that:we successfully cultured purity median concentration of90%(76%to98%) γδT cells amplified by the peripheral blood of MM patients; The killing rate of γδT cells on multiple myeloma U266cells were significantly increased with effector to target ratio in vitro; The cell growth and proliferation inhibition experimental results show that the U266cells and γδT cells are both able to inhibited by Bor and inhibition rate increased with the increase of drug concentration and treatment times; Bor can induce apoptosis in U266cells, and the apoptosis rate was increased with the prolongation of incubation times; The sensitivity of γδT cells killing U266cells that pre-incubated with selected10.0nmol/L Bor in8hours significantly higher than not pre-incubated group, Compared with the untreated group, selected concentration10.0nmol/L of Bor were incubated U266cells for8hours,16hours,24hours. The expression of NKG2D ligands (ULBP1,ULBP2, ULBP3, MICA/B) and TRAIL (DR4, DR5) on U266cells surface were increased after8hour’s pre-incubated; The expression of ULBP2, MICA/B, and DR4was significantly increased after16hour’s treatment, The expression levels of ULBP1, ULBP3, DR5showed no change. After24hour’s treatment, the U266cell surface NKG2D ligands (ULBP1, ULBP2, ULBP3, MICA/B) and TRAIL (DR4, DR5) expression rates appeared lower.So according to the experimental results we get the following conclusion that: (1) we succeed in culture high purity γδT cells, γδT cell killing effect of U266cells significantly and in dose-dependent manner;(2) Bor were both inhibit U266cells and γδT cells, and the growth inhibition rate were in a time and concentration-dependent manner; Bor induce apoptosis in U266cells, such induction of apoptosis in time-dependent;(3)The amount of expression NKG2D ligands (ULBP1,ULBP2, ULBP3,MICA/B) and TRAIL (DR4, DR5) on U266cells surface showing a trend first and then decreased after Bor incubation,it is great significance for chose the right time points with the combination;(4) The killing sensitivity of U266cells treated with Bor to γδT cells was higher than the untreated group and the monotherapy group, the combination has a synergistic effect; its mechanism may be associated with upregulation expression rate of the cell surface NKG2D ligands (ULBP1,ULBP2,ULBP3, MICA/B) and TRAIL (DR4, DR5).