ABSTRACT
Objective:To explore the mechanism of the cytotoxicity of human NK cells induced by atorvastatin to colon cancer cell lines. Methods:After colon cancer cells (HCT-116,SW-480,Caco-2) were cultured with different concentrations of atorvastatin, CCK-8 assay was used to assess the effect of atorvastatin on growth of colon cancer cells. The amplification of human NK cells was induced by SCGM medium in vitro. Automatic biochemical analyzer was applied to test the cytotoxicity of NK cells to colon cancer cells which cultured with different concentration of atorvastatin. FCM was used to detect the expression rate of MICA/B on the cells. Results:(1) The cultivation of NK cells:The proportion of NK cells attained to 93. 1% from 4. 5% after cultured for 10 days. (2) The effects of atorvastatin on the growth of the colon cancer cells:After cultured with atorvastatin,the inhibition rate of HCT-116 cells was higher than that in control when the density of atorvastatin increased from 5 μmol/L to 40 μmol/L after 48 h and from 1. 25 μmol/L to 40 μmol/L after 96 h ( P<0. 05 ) . Correlation analysis showed that the concentration of atorvastatin and the growth inhibition rate of HCT-116 cells were positively correlated(r[48 h]=0. 13,r[96 h]=0. 22,P<0. 05). (3) The cytotoxicity of NK cells to colon cancer cells effected after atorvastatin: In different atorvastatin concentrations groups,the cytotoxicity of NK cells to three colon cancer cell lines was all higher than that in control ( P<0. 05 ) . The atorvastatin concentration was from 2. 5 μmol/L to 10 μmol/L for HCT-116 cells,from 5 μmol/L to 20μmol/L for SW-480 cells,and from 2. 5μmol/L to 20μmol/L for Caco-2 cells. Among the three cell lines, the cytotoxicity of NK cells to HCT116 was the highest in the same concentration. (4)NK cells by atorvastatin cutting statins 96 h,the concentration of 20 mmol/L and 40 mmol/L inhibition rate was higher than that of control group,more than other groups on NK cell growth without significant effect. ( 5 ) The impact of atorvastatin on MICA/B expression of colon cancer cells: After cultured with different concentrations of atorvastatin,the expression of MICA/B on colon cancer cells was higher than that in control(P<0. 05). The concentration was 2. 5μmol/L and 5μmol/L for HCT-116 cells,10μmol/L and 20μmol/L for SW-480 cells,and from 2. 5μmol/L to 40 μmol/L for Caco-2 cells. Conclusion:Atorvastatin could inhibit the growth of colon cancer cells (HCT-116,SW-480 and Caco-2) in a dose-dependent manner;and it could enhance the cytotoxicity of NK cells to colon cancer cells;it also could promote the expression of MICA/B of colon cancer cells,and improve the immunogenicity of colon cancer cells.
ABSTRACT
Objective:To investigate the effect and mechanism of Wogonin on CD3AK cell proliferation and cytotoxicity to SMMC-7721.Methods:CD3AK cells were cultured from peripheral blood mononuclear cells ( PBMC) in vitro by a variety of cytokines for 7 d,and treated with different concentrations of Wogonin for 48 h.CD3AK cells proliferation was measured by CCK-8 assay.SMMC-7721 cell growth was detected by MTT.The expression of perforin (PFP),granzyme B (GrB) and CD107a on CD3AK cells were measured by flow cytometry ( FCM).The cytotoxicity to SMMC-7721 cells was detected by LDH release assay.The expression of ERK1/2 on CD3AK cells was detected by Western blot.The mobility of SMMC-7721 cells was detected with transwell chambers.The merge of SMMC-7721 cells were measured with Wound healing assay.Results:Wogonin could significantly promote CD3AK cells proliferation, especially at 3.2 mg/L (23%higher than that of control group,P<0.05).The highest cytotoxicity to SMMC-7721 was also at the con-centration 3.2 mg/L (60.4%).The expression of PFP,GrB,CD107a were significantly higher than that of control group( P<0.05).The expression of ERK1/2 was obviously improved,especially at 12.5-0.8mg/L.After treated with Wogonin 50,12.5,3.2,0.8,0.2 mg/L for 48 h,the lowest transwell cell was at 12.5 mg/L and lowest merge rate was at 3.2 mg/L.Conclusion:Wogonin could promote CD3AK cell proliferation and enhance the cytotoxicity to SMMC-7721.Wogonin could also inhibit SMMC-7721 cell growth,migration and cell merge.The mechanism may be related to activated ERK1/2 and increase the expression of PFP,GrB,CD107a.