Nakano Yoshiteru, Kuroda Etsushi, Kito Tomohiro, Uematsu Satoshi, Akira Shizuo, Yokota Akira, Nishizawa Shigeru, Yamashita Uki
Department of Neurosurgery, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
J Neurosurg. 2008 Feb;108(2):311-9. doi: 10.3171/JNS/2008/108/2/0311.
Microglia are one of the members of monocyte/macrophage lineage in the central nervous system (CNS) and exist as ramified microglia in a normal resting state, but they are activated by various stimuli, such as tumors. Activated microglia induce immune responses in the CNS, but the precise functions of microglia in glioma microenvironments are not clear. It has been reported that glioma cells produce prostaglandin (PG)E2, which promotes the growth of tumor cells and possesses immunosuppressive activity. The authors previously reported that PGE2 production by peritoneal macrophages was enhanced by glioma-derived soluble factors, which induce an immunosuppressive state. In this study, they investigated PGE2 production by microglia treated with glioma cells and assessed the role of microglia in glioma microenvironments in the mouse.
Microglia and peritoneal macrophages were cultured in vitro with or without lipopolysaccharide, and tumor necrosis factor (TNF) and PGE2 in the culture supernatant were measured using L929 bioassay and enzyme immunoassay. The expression of mRNA was measured using reverse transcriptase polymerase chain reaction, and the protein expression was assayed with Western blotting. In some experiments glioma cells and conditioned glioma medium were added to the microglia cultures.
Glioma cells studied in this report did not produce a significant amount of PGE2. However, the coculture of microglia with glioma cells or conditioned glioma medium led to the production of a large amount of PGE2. The enhancement of PGE2 production by microglia was more significant than that by peritoneal macrophages. The expression of cyclooxygenase (COX)-2 and particularly the expression of microsomal PGE synthase (mPGES)-1 (a terminal enzyme of the arachidonate cascade) in microglia were enhanced by conditioned glioma medium. The enhancement of mPGES-1 expression in microglia was more significant than that in peritoneal macrophages. The production of TNF was suppressed when culturing microglia with conditioned glioma medium, but this suppression was abrogated by the addition of a COX inhibitor (NS-398) and a PGE2 receptor (EP4) antagonist. Furthermore, TNF production was not suppressed in microglia from mPGES-1-deficient mice.
These results indicate that PGE2 production by microglia is enhanced by conditioned glioma medium, which induces an immunosuppressive state in the CNS. Therefore, the manipulation of microglia, from the standpoint of PGE2, provides investigators with an important strategy to induce an effective antiglioma immune response.
小胶质细胞是中枢神经系统（CNS）中单核细胞/巨噬细胞谱系的成员之一，在正常静息状态下以分支状小胶质细胞的形式存在，但它们会被各种刺激（如肿瘤）激活。活化的小胶质细胞可在中枢神经系统中诱导免疫反应，但小胶质细胞在胶质瘤微环境中的具体功能尚不清楚。据报道，胶质瘤细胞可产生前列腺素（PG）E2，其可促进肿瘤细胞生长并具有免疫抑制活性。作者之前报道，胶质瘤衍生的可溶性因子可增强腹膜巨噬细胞产生PGE2，这些因子可诱导免疫抑制状态。在本研究中，他们研究了用胶质瘤细胞处理的小胶质细胞产生PGE2的情况，并评估了小胶质细胞在小鼠胶质瘤微环境中的作用。
小胶质细胞和腹膜巨噬细胞在体外培养，添加或不添加脂多糖，使用L929生物测定法和酶免疫测定法测量培养上清液中的肿瘤坏死因子（TNF）和PGE2。使用逆转录聚合酶链反应测量mRNA的表达，并用蛋白质印迹法测定蛋白质表达。在一些实验中，将胶质瘤细胞和条件性胶质瘤培养基添加到小胶质细胞培养物中。
本报告中研究的胶质瘤细胞未产生大量PGE2。然而，小胶质细胞与胶质瘤细胞或条件性胶质瘤培养基共培养导致产生大量PGE2。小胶质细胞产生PGE2的增强比腹膜巨噬细胞更显著。条件性胶质瘤培养基可增强小胶质细胞中环氧化酶（COX）-2的表达，尤其是微粒体PGE合酶（mPGES）-1（花生四烯酸级联反应的末端酶）的表达。小胶质细胞中mPGES-1表达的增强比腹膜巨噬细胞更显著。用条件性胶质瘤培养基培养小胶质细胞时，TNF的产生受到抑制，但添加COX抑制剂（NS-398）和PGE2受体（EP4）拮抗剂可消除这种抑制作用。此外，mPGES-1缺陷小鼠的小胶质细胞中TNF的产生未受到抑制。
这些结果表明，条件性胶质瘤培养基可增强小胶质细胞产生PGE2，从而在中枢神经系统中诱导免疫抑制状态。因此，从小胶质细胞产生PGE2的角度进行调控，为研究人员提供了一种诱导有效的抗胶质瘤免疫反应的重要策略。
