Effect of siRNA targeting inhibition of 4-1BBL gene expression in HL-60 tumor cells on lymphocytic proliferation and apoptosis

Objective:To observe the effects of 4-1BBL siRNA on the cytokine production of HL-60 cells,compare the effects of culture supernatant of HL-60 cells on lymphocyte proliferation and inducing apoptosis between before and after interfering 4-1BBL,and explore the immune escape mechanism of tumor cells.Methods:The siRNA targeting 4-1BBL gene was designed and synthesized in vitro,then transfected into HL-60 cells using Lipofectamine.The expression levels of 4-1BBL mRNA and protein in HL-60 cells were detected using RT-PCR and flow cytometry,respectively.The production levels of TGF-β and VEGF in culture supernatant of HL-60 cells were determined by enzyme linked immunosorbent assay(ELISA).Peripheral blood mononuclear cells(PBMC) were collected by density gradient centrifugation,and co-cultured with HL-60 cells culture supernatant in vitro.The proliferation and apoptosis rates of lymphocytes were determined by MTT assay and flow cytometry,respectively.Results:After the siRNA targeting 4-1BBL gene was transfected into HL-60 cells,the expression levels of 4-1BBL gene and protein significantly decreased(P<0.01).The results of ELISA showed that the secretion levels of TGF-β and VEGF in culture supernatant of HL-60 cells significantly decreased after the 4-1BBL gene expression was interfered(P<0.01 and P<0.05).Compared with before interfering,the effects of the culture supernatant of HL-60 cells on inhibiting the proliferation and inducing apoptosis of lymphocyte decreased after interfering(P<0.05).Conclusions:The high expression of 4-1BBL in HL-60 cells has a reversely regulatory function.The production of TGF-β and VEGF in HL-60 cells can strengthen the inhibiting effect of tumor cell culture supernatant on lymphocyte activity,and promote the lymphocytes apoptosis,which can be an important mechanism of tumor cells escaping from the immune surveillance of tumor-bearing host.