Effects of curcumin on in vitro proliferation and apoptosis of mouse hepatocellular carcinoma cells
Objective To explore the effects of curcumin on the in vitro proliferation and apoptosis of mouse liver can-cer Hepa1-6 cells.Methods Hepa1-6 cells in the logarithmic growth phase were divided into the control group,10 μmol/L group,20 μmol/L group and 40 μmol/L group,respectively,which were treated with 0,10,20,and 40 μmol/L of cur-cumin.Cell proliferation was assessed by the MTT assay,and apoptosis level was detected by FACS.Real-time quantitative PCR(RT-qPCR)was employed to detect the levels of apoptosis-related molecules Bcl-2,Caspase-3,Toll-like receptor 2(TLR2),IFN-α,and IFN-β in Hepa1-6 cells.The secretion levels of IFN-α and IFN-β in the cell culture supernatant were determined using ELISA.Hepa1-6 cells were incubated with CD4+T cells,CD8+T cells and NK cells in the control group and 40 μmol/L curcumin group,respectively.FACS was used to detect the expression levels of CD69 in CD4+T cells,CD8+T cells and NK cells.Results The proliferation inhibition rates of Hepa1-6 cells in the 10,20 and 40 μmol/L groups increased in a dose-dependent manner,and the inhibition rates were(36.45±2.11)%,(46.40±2.08)%and(82.16±3.32)%,respectively.There was statistically significant difference in comparison with that of the control group(P<0.05).The apoptotic proportions of Hepa1-6 cells in the 10 μmol/L,20 μmol/L and 40 μmol/L groups were 10.47%,26.66%and 49.81%,respectively.And the difference was statistically significant in comparison with that of the control group(P<0.01).RT-qPCR results showed that the transcription levels of anti-apoptotic gene Bcl-2 in the 10 μmol/L,20 μmol/L and 40 μmol/L groups were(0.77±0.06)times,(0.44±0.06)times and(0.25±0.08)times that of the control group,respectively,with statistically significant difference(all P<0.01).At the same time,the expression of pro-apoptotic gene Caspase-3 significantly increased,and the transcription levels of pro-apoptotic gene Caspase-3 in the 10 μmol/L,20 μmol/L and 40 μmol/L groups was(1.73±0.31)times,(2.52±0.45)times and(3.25±0.20)times that of the control group,respectively,with statistically significant difference(all P<0.01).With the increase of curcumin concentrations,TLR2 transcription levels increased gradually.TLR2 transcription levels in the 10 μmol/L,20 μmol/L and 40 μmol/L groups were(3.56±0.59)times,(4.93±0.04)times and(6.60±0.47)times that of the control group,respectively,with statistically significant difference(all P<0.05).In addition,the transcription levels of IFN-α in the 10 μmol/L,20 μmol/L and 40 μmol/L groups significantly increased,which were(4.21±0.22),(5.54±0.39)and(8.05±0.93)times that of the control group,respectively,and there were statistically significant differences among these groups(all P<0.05).The transcription levels of IFN-β were(3.08±0.51),(5.31±0.55)and(8.35±0.59)times that of the control group,and the difference was statistically significant(all P<0.01).For IFN-α,compared with the control group[(36.26±6.12)pg/mL],the levels of IFN-α secreted by cells in the 10 μmol/L,20 μmol/L and 40 μmol/L groups were(57.00±8.8),(82.47±4.42),and(100.37±8.99)pg/mL,respectively;the levels of IFN-α secreted by cells in the 10,20,and 40 μmol/L groups were higher that that of the control group(all P<0.01).The levels of IFN-β secreted by the cells in the 10,20,and 40 μmol/L groups were(60.67±6.81),(91.33±5.51),and(144.00±10.15)pg/mL,respec-tively,while that in the control group was(44.67±1.15)pg/mL.The levels of IFN-β secreted by the cells in the 10,20,and 40 μmol/L groups were higher than that in the control group(all P<0.01).In the control group,CD69 expression lev-els of CD4+T cells,CD8+T cells and NK cells were(24.53±1.39)%,(23.40±2.32)%,(32.33±3.08)%,respectively.The CD69 expression levels of CD4+T cells,CD8+T cells and NK cells in the 40 μmol/L group were(39.90±3.51)%,(31.47±3.88)%,(50.83±4.74)%,respectively.There were significant differences in CD69 levels of CD4+T cells,CD8+T cells and NK cells between control group and 40 μmol/L group(all P<0.05).Conclusion Curcumin promotes the secretion of type Ⅰ interferon by regulating the expression of TLR2,thus inhibits the proliferation of hepatocellular car-cinoma cells,induces the apoptosis of hepatocellular carcinoma cells,as well as promotes the activation of immune cells.
hepatocellular carcinomacurcuminToll-like receptor 2cell proliferationapoptosisimmune function
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