阿托伐他汀对人NK 细胞杀伤结肠癌细胞的影响及其机制研究

目的:探讨阿托伐他汀对人NK 细胞杀伤结肠癌细胞的影响及其机制。方法:不同浓度的阿托伐他汀作用于3 株结肠癌细胞(HCT-116、SW-480、Caco-2),CCK-8 法测定阿托伐他汀对结肠癌细胞生长抑制率的影响。SCGM 培养基体外扩增人NK 细胞,自动生化分析仪测定NK 细胞对3 株结肠癌细胞的杀伤活性;流式细胞仪检测结肠癌细胞MICA/ B 的表达率。结果:(1)NK 细胞的培养:培养前CD3- CD56+的NK 细胞占外周血单个核细胞的比例为4.5%,培养10 d 时NK 细胞的比例增至93.1%。(2)阿托伐他汀对3 株结肠癌细胞生长抑制率的影响:阿托伐他汀作用48 h 后,在浓度5 ~40 μmol/ L 的4 个实验组中,HCT-116 细胞的生长抑制率较对照组升高(P<0.05)。作用96 h 后,在1.25 ~ 40 μmol/ L 的所有浓度组(6 个)中,HCT-116 细胞的生长抑制率均显著高于对照组(P<0.05)。另外, HCT-116 细胞的生长抑制率随着阿托伐他汀浓度的升高而逐渐上升,相关分析显示,阿托伐他汀的浓度与HCT-116 细胞的生长抑制率呈正相关(r48 h] = 0.13,r96 h] = 0.22,P<0.05)。(3)NK 细胞经阿托伐他汀作用96 h 后,除浓度为20 μmol/ L 和40 μmol/ L 时抑制率高于对照组,其他各组对NK 细胞生长无明显影响。(4)阿托伐他汀对NK 细胞杀伤结肠癌细胞活性的影响:NK 细胞对HCT-116 细胞的杀伤活性在阿托伐他汀浓度2.5 ~10 μmol/ L 组均显著高于对照组,对SW-480 的杀伤活性在5 ~20 μmol/ L 组较对照组明显升高,对Caco-2 的杀伤活性在2.5 ~20 μmol/ L 组显著高于对照组(P<0.05),其中同一浓度时对HCT-116 细胞杀伤作用最强。(5)阿托伐他汀对结肠癌细胞MICA/ B 表达的影响:在阿托伐他汀浓度2.5 μmol/ L 及5 μmol/ L 组,HCT-116 细胞MICA/ B 的表达率与对照组比较显著升高(P <0.05),在10 μmol/ L 及20 μmol/ L 组,SW-480 细胞MICA/ B 表达率显著高于对照组(P <0.05),在2.5 ~40 μmol/ L 组,Caco-2 细胞MICA/ B 的表达率均较对照组显著升高(P<0.05)。结论:(1)阿托伐他汀能够呈剂量依赖性抑制结肠癌细胞HCT-116、SW-480 及Caco-2 的生长;(2)阿托伐他汀能够增强NK 细胞对结肠癌细胞的杀伤活性,可能与阿托伐他汀上调结肠癌细胞MICA/ B 的表达有关;(3)阿托伐他汀可以上调3 株结肠癌细胞MICA/ B 的表达率,提高结肠癌细胞的免疫原性。

Effect and mechanism of atorvastatin on cytotoxicity of human NK cells to colon cancer cells

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(r48 h] =0.13,r96 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 20 μmol/ L for SW-480 cells,and from 2.5 μmol/ L to 20 μmol/ 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 2.5 μmol/ L and 5 μmol/ L for HCT-116 cells,10 μmol/ L and 20 μmol/ L for SW-480 cells,and from 2.5 μmol/ 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.