表达CD40L基因的卵巢癌细胞诱导树突状细胞成熟及分泌Th1型细胞因子的作用机制  

Enhanced expression of CD40L cDNA on ovarian cancer cell line OVHM induces the secretion of Thl cytokines from dendritic cells

作　　者：张正茂[1] 张风华[3] 王喜梅[4] 张超[2] 刘洁[1] 顾来梅 李全海 单保恩[2] 田川雅敏 

机构地区：[1]河北医科大学第四医院妇产科,石家庄050011 [2]河北医科大学第四医院科研中心,石家庄050011 [3]河北省人民医院普外科 [4]河北省南皮县人民医院妇产科 [5]日本千葉かんセンタ-研究局病理研究部

出　　处：《中华肿瘤杂志》2008年第3期174-178,共5页Chinese Journal of Oncology

基　　金：河北省自然科学基金资助项目（C2007001058）

摘　　要：目的探讨转染CD40L cDNA的卵巢癌细胞株OVHM对树突状细胞（DC）成熟、分化的影响及诱导其分泌细胞因子的机制。方法采用脂质体转染法将鼠全长CD40L基因转染人小鼠卵巢癌细胞株OVHM中，用C418筛选出表达CD40L基因的抗药性克隆细胞（CD40L-OVHM）。应用免疫磁珠分选并纯化小鼠骨髓DC。将DC与转染和未转染CD40L cDNA的OVHM细胞混合培养，流式细胞术检测DC细胞表面人主要组织相容性复合物Ⅰ类分子（MHC-Ⅰ）、Ⅱ类分子（MHC-Ⅱ）、CD80、CD86和CCR7的表达，逆转录聚合酶链反应（RT—PCR）检测共培养细胞中自细胞介素（IL）-12、IL-23、IL-27、IL-18、γ干扰素（IFN-γ）、Mig基因的表达。结果DC与CD40L—OVHM细胞混合培养后可形成集落，且上调了DC细胞MHC-Ⅰ、MHC-Ⅱ、CD80、CD86、CCR7的共刺激分子和黏附分子的表达。在DC＋CD40L-OVHM组可检测到细胞因子IL-12、IL-23、IL-27、IL-18、IFN-γ、Mig基因的表达，而在DC组、DC＋0VHM组、CD40L-OVHM组、OVHM组未检测到这些基因的表达。结论表达CD40L的卵巢癌细胞促进了DC的成熟，诱导了Th1型细胞因子的分泌，这可能是转染CD40L基因的卵巢癌细胞产生抗肿瘤效应的机制之一。Objective To examine whether the enhanced expression of CD40L eDNA on murine ovarian cancer （OVHM） cells could induce the secretion of Thl cytokines from dendritic cells （DC）. Methods OVHM cells were transfected with the full-length mouse CD40L eDNA by lipofectamine^TM 2000 and then G418 resistant cells as positive cells were selected. They were examined for their expression of CD40L with flow cytometry. Bone marrow cells were firstly depleted of erythrocytes, maerophages, T and B cells with PE-conjugated magnetic beads, and then cultured in 10% FCS RPMI 1640 medium supplemented with recombinant mouse GM-CSF and IL-4 for 10 days. PKH67-1abeled tumor cells were cultured with DC, and then the stained cells were analyzed for the expression of MHC-Ⅰ, MHC-Ⅱ, CD80, CD86, CCR7 in DC with flow cytometry. The expression of P40, p19, p35, p28, EBi3 subunits, IL-18, IFN-γ, Mig gene in cocultured DC-tumor cells were detected by RT-PCR. Results The CD40L eDNA was successfully transfected into OVHM cells. Bone marrow-derived DCs, when cultured with CD40L/OVHM, formed clusters with the tumors and showed an upregulated expression of MHC- Ⅰ , MHC-Ⅱ , CD80, CD86, CCR7. Expression of the IL-12, IL-23, IL-27, IL-18, interferon-γ （IFN-γ） and Mig （monokine induced by IFN- γ） genes was induced in the DCs that were cultured with CD40L/OVHM but not with OVHM cells. Conclusion These data directly showed that the expression of CD40L on ovarian cancer cells facilitates the interaction between DCs and tumors, enhances the maturation of DCs, induces secretion of Thl cytokines, esoeeially for IL-12, IL-23 and IL-27, which maybe one of the possible antitumor mechanism for CD40L-transfected ovarian cancer cell line.

关 键 词：CD40L 树突状细胞 细胞因子 卵巢肿瘤 

分 类 号：R686[医药卫生—骨科学]