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Combining norcantharidin with IL-2 and IL-15 enhances the cytotoxicity of PBMC on Raji cells
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Abstract:
Objective:To explore the cytotoxicity and the underlying mechanisms of peripheral blood mononuclear cells (PBMCs) activated by IL-2 and IL-15 on human Burkitt lymphoma Raji cells after being treated by norcantharidin (NCTD).Methods: Trypan blue assay was used to detect the inhibitory effect of NCTD on Raji cells and PBMC. The cytotoxic effects of PBMC activated by IL-2 and IL-15 against K562 cells and Raji cells were analyzed by LDH releasing assay. The expression of NKG2D on the surface of PBMC activated by IL-2 and IL-15 was assayed by flow cytometry (FCM). The expressions of NKG2D ligands (MICA, MICB, ULBP1, ULBP2, ULBP3) on Raji cells were assayed by FCM before and after NCTD treatment. Results: NCTD inhibited the proliferation of Raji cells significantly in a dose- and time-dependent manner (P<0.05), on the contrary, it showed no significant effect on PBMC proliferation (P>0.05). The cytotoxic rates of IL-2 and IL-15 activated PBMC on K562 cells were more than that of inactivated PBMC when the ratio of effect/target (E∶T) were 10∶1 and 20∶1 (\[52.42±3.89\]% vs \[15.82±5.12\]%, \[79.55±922\]% vs \[2767±366\]%, P<0.05), respectively. The cytotoxic rates of inactivated PBMC against Raji cells after being treated by NCTD were increased compared with untreated Raji cells (\[23.63±6.20\]% vs \[5.04±1.25\]%, \[41.80±409\]% vs \[8.59±2.19\]%, P<0.05) with E∶T of 10∶1 and 20∶1 respectively. Meanwhile, the cytotoxic rates of PBMC after being activated by IL-2 and IL-15 on NCTD-treated Raji cells were also increased compared with that of untreated PBMC (\[38.97±2.76\]% vs \[13.19±3.67\]%, \[63.09±7.30\]% vs \[19.89±4.15\]%, P<0.05). The expression of NKG2D on PBMC activated by IL-2 and IL-15 was increased compared with untreated PBMC (\[4491±585\]% vs \[25.28±769\]%, P<0.05). The expressions of NKG2D ligands on Raji cells, especially ULBP2 were up-regulated (\[12.69±3.99\]% vs \[1.03±0.42\]%, P<0.05) after being treated by NCTD, while no significant changes were found in other ligands (P>0.05). Conclusion: Combining NCTD with IL-2 and IL-15 can increase the cytotoxicity of PBMC on Raji cells, which may be related with NCTD-increased expression of ULBP2 on Raji cell surface and cytokine-increased expression of NKG2D on PBMC surface.
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Project Supported:
Project supported by the National Natural Science Foundation of China (No.30973454)
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