肝癌细胞微环境对不同分化阶段树突状细胞功能状态的影响  

The effect of hepatocellular carcinoma cell microenvironment on the functional status of dendritic cells at different differentiation stages

作　　者：曾柱[1] 龙金华[2] 陈琳[3] 

机构地区：[1]贵州省贵阳医学院生物技术教研室,贵阳550004 [2]贵州省肿瘤医院头颈肿瘤科,贵阳550004 [3]贵州省贵阳医学院物理教研室,贵阳550004

出　　处：《重庆医科大学学报》2010年第12期1773-1778,共6页Journal of Chongqing Medical University

基　　金：教育部科学技术重点项目(编号:210196);贵州省省长基金(编号:黔省专字2009-79);贵州省自然科学基金(编号:黔省合J字2008-2274);贵州省贵阳市科学技术项目(编号:2010筑科农合同字第1-社-12)

摘　　要：目的:肿瘤的发生发展伴随有其对免疫系统各效应单元的显著抑制,树突状细胞(Dendritic cells,DCs)是目前所知功能最为强大的抗原呈递细胞(Antigen presenting cells,APC)并在天然和适应性免疫应答的启动方面发挥着至关重要的作用。本文探讨肝癌细胞(Hepatocellular carcinoma cells,HCC)分泌的可溶性细胞因子营造的微环境对树突状细胞的功能状态的影响,以探索傅立叶变换红外光谱(Fourier transformed infrared spectrum,FT-IR)技术在基于DCs的抗肿瘤细胞免疫治疗中的临床应用。方法:用免疫磁珠(Miltenyibiotec)从人外周血分离CD14+单核细胞,加入粒-巨噬细胞集落刺激因子(Recombinant human granulocyte-macrophage CSFr,hGM-CSF)、白介素4(Recombinant human interleukins 4r,hIL-4)将单核细胞诱导分化为未成熟DCs(ImmatureDCsi,mDCs),利用肿瘤坏死因子α(Tumor necrosis factor-α,TNF-α)将imDCs诱导为成熟DCs(Mature DCs,mDCs),分别将imDCs和mDCs与HCC在Transwell中共培养48 h,正常培养和撤生长因子培养的DCs作为对照,采用FT-IR技术和分子生物学技术研究HCC对DCs功能状态的影响。结果:与正常培养和撤生长因子培养的DCs相比,与HCC共培养DCs的基因转录活性和能量状态受到了显著的抑制,免疫印迹的结果进一步证实DCs的转录因子NF-κB的确受到了由HCC分泌的细胞因子的抑制。结论:HCC能够抑制DCs的基因转录活性和能量状态,FT-IR的变化与转录因子NF-κB的表达具有高度的相关性,这为FT-IR技术应用于检测DCs的功能状态奠定了基础,对基于DCs的抗肿瘤免疫治疗具有重要意义。Objective：The generation and progression of tumor is accompanied by a marked suppression of the effect units of immune system.DCs are the most powerful antigen presenting cells as now known and play significant roles in the initiation of innate and acquired immune response.This research aims to study the influence of microenvironment created by soluble cytokines derived from hepatocellular carcinoma cells（HCC） on the functional status of DCs,and to explore the clinical application of Fourier-Transformed Infrared（FT-IR） technology to the DCs-based cellular immune therapy against tumor.Methods：The monocytes were cultured in RPMI1640/10% FBS supplemented with 100 ng/ml rhGM-CSF and 100 ng/ml rhIL-4 for 7 days to develop into immature DCs（imDCs）.Maturation was induced by addition of 20 ng/ml rhTNF-α to imDCs for another 3 days of culture.Acquired imDCs and mDCs were respectively co-cultured with human HCC for 48 hours in Transwell.The functional status of treated cells was investigated by FT-IR scanning and western blotting with untreated cells as control.Results：Compared with untreated DCs,the DCs co-cultured with HCC appeared to be suppressed in gene transcription activity and energy status.The results of western blotting further demonstrated the NF-κB transcription factors of DCs treated with HCC were inhibited by tumor-derived factors.Conclusion：HCC can inhibit the gene transcription activity and energy status.The ratio of absorption intensity of FT-IR at given wave numbers is markedly correlative with the expression of NF-κB.Clinically,these results may lay the foundation for the application of FT-IR to the analyses of DCs functional status in cellular immune therapy against cancer.

关 键 词：树突状细胞 傅立叶变换红外光谱 基因转录活性 能量状态 

分 类 号：Q6-3[生物学—生物物理学]