塞来昔布抑制COX-2和PD-1发挥抗肝癌作用  

Antihepatocarcinoma effect of Celecoxib by inhibiting COX-2 and PD-1

作　　者：詹磊[1,2] 程良斌[1] 蔡岳[3] 程良斌 ZHAN Lei;CHENG Liang-Bin;CAI Yue;CHENG Liang-Bin(Hepatology Department,Hubei Provincial Traditional Chinese Medical Hospital,Wuhan 430060,China)

机构地区：[1]湖北省中医院肝病科,武汉430060 [2]湖北省中医药研究院,武汉430074 [3]湖北中医药大学,武汉430065

出　　处：《中国免疫学杂志》2018年第4期595-599,共5页Chinese Journal of Immunology

摘　　要：目的:研究塞来昔布的抗肝癌作用及其潜在的分子机制。方法:选取2012年2月~2016年6月在我院进行手术治疗的原发性肝细胞癌(HCC)患者65例作为研究对象,另选取肝胆外科提供的正常肝脏组织标本15例作为对照。采用免疫组织化学法(IHC)检测肿瘤及正常组织样本中COX-2与PD-1的表达。采用Pearson相关性分析HCC患者COX-2与PD-1的相关性。建立H22肝癌细胞BALB/c小鼠模型,随机分为对照组、塞来昔布组和PD-1抗体组(每组15只)。给药4周后处死全部存活小鼠,取出肿瘤。绘制肿瘤生长曲线及小鼠生存曲线。采用IHC分析各组肿瘤组织中COX-2、PD-1的表达水平及CD8^+T与Foxp3阳性Treg细胞数。流式细胞术检测外周血中CD8^+T与CD4^+CD25^+Treg细胞数量。分离正常BALB/c小鼠的外周血单核细胞(PBMC),分别转染PD-1及对照siRNA,使用塞来昔布处理后,Western blot检测COX-2、PD-1、CD8及Foxp3水平。结果:IHC结果显示HCC患者肿瘤组织中COX-2与PD-1的表达水平均显著高于对照组(P<0.001)。Pearson相关性分析显示HCC患者肿瘤组织中COX-2的表达水平与PD-1呈显著正相关(R2=0.673,P<0.001)。塞来昔布、PD-1抗体治疗小鼠的肿瘤生长曲线和生存曲线均显著优于对照组(P<0.05),塞来昔布组、PD-1抗体组小鼠的肿瘤生长曲线和生存曲线相比差异均无统计学意义(P>0.05)。IHC与流式分析显示,塞来昔布能显著降低肿瘤组织中PD-1的表达(P<0.05);塞来昔布与PD-1抗体均能使肿瘤组织及外周血CD8^+T细胞显著增多(P<0.05),使Treg细胞显著减少(P<0.05)。Western blot结果显示,塞来昔布能显著降低小鼠PBMC的COX-2、PD-1、CD8及Foxp3水平,而不影响转染PD-1 siRNA后PBMC的CD8及Foxp3水平。结论:HCC患者肿瘤组织中COX-2与PD-1均显著增高,塞来昔布可能通过抑制COX-2调节PD-1影响肿瘤免疫从而发挥抗肝癌的作用。Objective:To investigate the role of celecoxib in inhibiting liver cancer and to explore the potential molecular mechanisms.Methods:A total of 65 cases of patients with primary hepatocellular carcinoma(HCC)underwent surgical treatment in our hospital from February 2012 to June 2016 were recruited as study object.Another 15 cases of normal liver tissue offered by the department of hepatobiliary surgery were selected as control.Immunohistochemical staining(IHC)was used to detect the expression level of COX-2 and PD-1 in tumor and normal tissue samples.Pearson correlation analysis were used to evaluate the correlation between COX-2 and PD-1 in HCC patients.Construction of H22 hepatoma cells bearing BALB/c mice model,randomly divided them into control group,celecoxib group and PD-1 antibody group(15 mice for each group).Sacrificed all mice alive at the end of the 4th weeks after the treatment and removed the tumors then.The tumor growth curves and survival curves were drawn to observe the anti-tumor effect.IHC were used to evaluate the expression of COX-2,PD-1 as well as the number of CD8+T and foxp3 positive Treg cells in tumor tissue.Flow cytometry were used to determine the number of CD8+T and CD4+CD25+Treg cells in peripheral blood.The peripheral blood mononuclear cell(PBMC)of BALB/c mice were separated and scramble or PD-1 siRNA were transfected then,Western blot analysis were used to detected the level of COX-2,PD-1,CD8 and Foxp3 after the treatment of celecoxib.Results:IHC results showed that the expression level of COX-2 and PD-1 in tumor tissue of HCC patients were significant higher than control(P<0.001).Pearson correlation analysis showed COX-2 were both positive correlated with PD-1 in the tumor tissue of HCC patients(R 2=0.673,P<0.001).The tumor growth curves and survival curves in celecoxib or PD-1 antibody groups were significant better than in control group(P<0.05).There were no significant difference of tumor growth curves and survival curves between celecoxib group and PD-1 antibody group(P>0.05).IH

关 键 词：塞来昔布 COX-2 PD-1 肝细胞癌 

分 类 号：R392[医药卫生—免疫学]