姜黄素对肿瘤相关巨噬细胞分泌细胞因子基因表达的影响

Effect of curcumin on gene expressions of cytokines secreted by tumor-associated macrophages

目的:研究姜黄素对口腔鳞状细胞癌(OSCC)细胞(Cal27细胞)上清液诱导的肿瘤相关巨噬细胞(TAMs)分泌细胞因子基因表达的影响,阐明姜黄素抗OSCC的作用机制。方法:将Raw264.7细胞随机分为对照组及不同浓度(1.25、2.50、5.00、10.00、20.00和40.00μmol·L^-1)姜黄素组,各组细胞加入Cal27细胞上清液共同孵育48 h,CCK-8法检测细胞活性。将Raw264.7细胞随机分为对照组及不同浓度(5、10和20μmol·L^-1)姜黄素组,各组细胞加入Cal27细胞上清液共同孵育36和48 h后,采用Real-time PCR法检测各组细胞中白细胞介素12(IL-12)、诱导型一氧化氮合酶(iNOS)、肿瘤坏死因子α(TNF-α)和白细胞介素10(IL-10)mRNA表达水平;Cal27细胞上清液孵育Raw264.7细胞48 h后,姜黄素组加入不同浓度(5、10和20μmol·L^-1)姜黄素干预48 h,采用Real-time PCR法检测各组细胞中IL-12、iNOS、TNF-α、IL-10和精氨酸酶1(Arg-1)mRNA表达水平。结果:与对照组比较,40μmol·L^-1姜黄素组Raw264.7细胞活性明显降低(P<0.01)。不同浓度姜黄素与Cal27细胞上清液共同孵育Raw264.7细胞36 h,与对照组比较,20μmol·L^-1姜黄素组Raw264.7细胞中IL-12 mRNA表达水平明显升高(P<0.01),10和20μmol·L^-1姜黄素组Raw264.7细胞中iNOS和TNF-αmRNA表达水平明显降低(P<0.05或P<0.01),20μmol·L^-1组Raw264.7细胞中IL-10 mRNA表达水平降低(P<0.01);不同浓度姜黄素与Cal27细胞上清液共同孵育Raw264.7细胞48 h,与对照组比较,5和20μmol·L^-1姜黄素组Raw264.7细胞中IL-12 mRNA表达水平明显升高(P<0.01),5和20μmol·L^-1姜黄素组Raw264.7细胞中iNOS、TNF-α和IL-10 mRNA表达水平均明显降低(P<0.01)。Cal27细胞上清液孵育Raw264.7细胞48 h后,采用不同浓度姜黄素进行干预,与对照组比较,10和20μmol·L^-1姜黄素组Raw264.7细胞中IL-12 mRNA表达水平升高(P<0.01),不同浓度姜黄素组Raw264.7细胞中iNOS、TNF-α和Arg-1 mRNA表达水平降低(P<0.05或P<0.01),20μmol·L^-1姜黄素组Raw264.7细胞中IL-10 mRNA表达水平均明显降低(P<0.01)。结论:姜黄素可以在诱导TAMs的不同阶段调节TAMs分泌细胞因子IL-12、iNOS、TNF-α、IL-10和Arg-1 mRNA表达水平。

Objective:To investigate the effect of curcumin on the gene expressions of cytokines secreted by tumorassociated macrophages(TAMs)induced by the supernatants of the oral squamous cell carcinoma(OSCC)cells(Cal27 cells),and to clarify the mechanism of curcumin’s anti-OSCC effect.Methods:The Raw264.7 cells were randomly divided into control group and different concentrations(1.25,2.50,5.00,10.00,20.00 and 40.00μmol·L^-1)of curcumin groups.The cells from each group were added with Cal27 cell supernatants and incubated for 48 h,and the cell activities were detected by CCK-8 assay.The Raw264.7 cells were randomly divided into control group and different concentrations(5,10 and 20μmol·L^-1)of curcumin groups.The cells from each group were added with Cal27 cell supernatants and incubated for 36 and 48 h,and Real-time PCR method was used to detect the mRNA expression levels of interleukin-12(IL-12),inducible nitric oxide synthase(iNOS),tumor necrosis factor-α(TNF-α)and interleukin-10(IL-10)in the cells in various groups.After the Raw264.7 cells were incubated with Cal27 cell supernatants for 48 h,the cells in curcumin groups were treated with different concentrations(5,10 and 20μmol·L^-1)of curcumin for 48 h,and Real-time PCR curcumin method was used to detect the mRNA expression levels of IL-12,iNOS,TNF-α,IL-10 and Arginase 1(Arg-1)in the cells in various groups.Results:Compared with control group,the cell activity of Raw264.7 cells in 40μmol·L^-1 curcumin group was significantly reduced(P<0.01).After the Raw264.7 cells were incubated with different concentrations of curcumin and Cal27 cell supernatants for 36 h,compared with control group,the IL-12 mRNA expression level in 20μmol·L^-1 curcumin group was increased(P<0.01);the iNOS and TNF-αmRNA expression levels in the Raw264.7 cells in 10 and 20μmol·L^-1 groups were significantly decreased(P<0.05 or P<0.01);the IL-10 mRNA expression level in 20μmol·L^-1 group was decreased(P<0.01).After the Raw264.7 cells were incubated with different concentrations of curcumin and Cal27 cell supernatants for 48 h,compared with control group,the IL-12 mRNA expression levels in the Raw264.7 cells in 5 and 20μmol·L^-1 groups were increased(P<0.01);the iNOS TNF-αand IL-10 mRNA expression levels in 5 and 20μmol·L^-1 groups were decreased(P<0.01).After the Raw264.7 cells were incubated with Cal27 cell supernatants for 48 h,and intervented with different concentrations of curcumin,compared with control group,the IL-12 mRNA expression levels in 10 and 20μmol·L^-1 curcumin groups were increased(P<0.01);the iNOS,TNF-αand Arg-1 mRNA expression levels in the Raw264.7 cells in different concentrations of curcumin groups were decreased(P<0.05 or P<0.01);the IL-10 mRNA expression level in the Raw264.7 cells in 20μmol·L^-1 curcumin group was significantly reduced(P<0.01).Conclusion:Curcumin can regulate the mRNA expression levels of cytokines IL-12,iNOS,TNF-α,IL-10 and Arg-1 secreted by TAMs at different stages of TAMs induction.