Fujitsu Youichiro, Fukuda Ken, Kimura Kazuhiro, Seki Keisuke, Kumagai Naoki, Nishida Teruo
Department of Biomolecular Recognition, Yamaguchi University School of Medicine, Ube City, Yamaguchi 755-8505, Japan.
Invest Ophthalmol Vis Sci. 2005 Mar;46(3):797-802. doi: 10.1167/iovs.04-1016.
To examine the possible roles of T helper type 2 (Th2) cell-derived cytokines in formation of the giant papillae characteristic in individuals with vernal keratoconjunctivitis, the effects of these cytokines on the proliferation and apoptosis of cultured human conjunctival fibroblasts were investigated.
Apoptosis was induced by the NO donor sodium nitroprusside. Cell viability was determined by measurement of mitochondrial metabolic activity, and apoptotic cells were identified on the basis either of nuclear morphology after staining with 4',6-diamidino-2-phenylindole or of TUNEL staining. The activation of antiapoptotic signaling mediated by the protein kinase Akt was assessed by immunoblot analysis and by an in vitro kinase assay. Expression of interleukin (IL)-4 and IL-13 receptor subunits was examined by reverse transcription and polymerase chain reaction analysis and by flow cytometry.
IL-4 and IL-13, but not IL-5, IL-9, or IL-10, induced the proliferation of conjunctival fibroblasts as well as protecting these cells from NO-induced apoptosis. Both IL-4 and IL-13 induced the phosphorylation of Akt and increased the kinase activity of this enzyme in a manner that was sensitive to the phosphatidylinositol 3-kinase inhibitors LY294002 or wortmannin. These inhibitors also blocked the antiapoptotic effects of IL-4 and IL-13. Transcripts encoding IL-4 and IL-13 receptor components were detected in conjunctival fibroblasts, and the proteins were expressed at the cell surface.
Among the various Th2 cytokines tested, only IL-4 and IL-13 induced the proliferation of human conjunctival fibroblasts and protected these cells from apoptosis. These effects may contribute to the formation of giant papillae in individuals with vernal keratoconjunctivitis.
为研究2型辅助性T细胞（Th2）衍生的细胞因子在春季角结膜炎患者特征性巨大乳头形成中的可能作用，本研究调查了这些细胞因子对培养的人结膜成纤维细胞增殖和凋亡的影响。
通过一氧化氮供体硝普钠诱导细胞凋亡。通过测量线粒体代谢活性来确定细胞活力，并用4',6-二脒基-2-苯基吲哚染色后的核形态或TUNEL染色来鉴定凋亡细胞。通过免疫印迹分析和体外激酶测定评估蛋白激酶Akt介导的抗凋亡信号传导的激活。通过逆转录、聚合酶链反应分析和流式细胞术检测白细胞介素（IL）-4和IL-13受体亚基的表达。
IL-4和IL-13而非IL-5、IL-9或IL-10可诱导结膜成纤维细胞增殖，并保护这些细胞免受一氧化氮诱导的凋亡。IL-4和IL-13均诱导Akt磷酸化，并以对磷脂酰肌醇3-激酶抑制剂LY294002或渥曼青霉素敏感的方式增加该酶的激酶活性。这些抑制剂也阻断了IL-4和IL-13的抗凋亡作用。在结膜成纤维细胞中检测到编码IL-4和IL-13受体成分的转录本，并且这些蛋白在细胞表面表达。
在所测试的各种Th2细胞因子中，只有IL-4和IL-13可诱导人结膜成纤维细胞增殖并保护这些细胞免受凋亡。这些作用可能有助于春季角结膜炎患者巨大乳头的形成。
