Biological Effects Of Soluble CD40 Ligand On Leukemia HL-60 Cells And Its Mechanism

Objective:To investigate the biological effects of sCD40L on inleukemia HL-60 cells and its possible molecular mechanism in regulating the proliferation, differentiation, and apoptosis. And provide certain theoretical basis for rhsCD40L to be used in clinical practice.Methods:1) HL-60 cells were co-incubated with sCD40L of different concentrations. MTT assay were used to detect the cell proliferation and chose the best concentration of sCD40L;2) HL-60 cells were co-incubated with the best concentration of sCD40L for different treating time.And used MTT assay to measure the cell growth inhibitory effect.3) Flow cytometry (FCM) was used to evalue their apoptosis.4) The expression changes of CD95 and Bcl-2 were obversed by FCM.5) HL-60 cells were co-incubated with the different concentrations of sCD40L for different treating time. The levels of IL-6 were determined by ELISA.Results:1) Compared with AS2O3 positive and HL-60 cells negative control groups, sCD40L was able to inhibit HL-60 cells proliferation in a dose-dependent manner for 48 hours (P<0.05),The increase of growth inhibitory rate was most significant at its cconcentration of 2.00μg/ml and 4.00μg/ml.2) The positive OD of sCD40L 2.00μg/ml group, sCD40L 4.00μg/ml group, AS2O3 positive control group and HL-60 cells negative control group respectively were 1.290±0.006,0.568±0.012,0.799±0.013,2.154±0.004 (P<0.05) and the OD of sCD40L 4.00μg/ml group was the lowest one.3) In this four groups the hightest of cells apoptosis rate was the sCD40L 4.00 ug/ml group for 48 hours (the hightest of cells apoptosis rate was over>20%, P< 0.01).4) In HL-60 cells cultured with sCD40L 4.00μg/ml for 48 hours the expression of CD95 was up-regulated most obviously (P<0.05) and Bcl-2 expression was down-regulated most significantly (P <0.05).5)The levels of IL-6 in sCD40L 4.00μg/ml group for 48 hours was the lowest one (P<0.05) and the next one was AS2O3 positive control group for 48 hours.However,there was no significant dose-and time- dependent manner of the IL-6 level (P>0.05)Conclusion:1) sCD40L can induce growth inhibition and apoptosis in HL-60 cells.2) In the process of HL-60 cells apoptosis, the the mitochondria pathway and death receptor pathway both play an important role.In another words,the molecular mechanism of HL-60 cells apoptosis induced by sCD40L is related to the up-regulation of CD95 and down-regulation of Bcl-2.3) There may be a relationship between the growth inhibition of HL-60 cells induced by sCD40L with the accommodation of IL-6.