Abstract：
Chemokines and adhesion molecules are major inflammatory mediators of chronic and recurrent vernal keratoconjunctivitis (VKC). Sulforaphane (SFN) is a natural plant extract that is known to have anti-inflammatory and antioxidant properties. SFN is demonstrated to be effective against a variety of human diseases. The current investigation examines the effects and the molecular mechanisms of SFN on cytokine-induced human corneal fibroblasts (HCFs) expression of adhesion molecules and chemokines. HCFs were exposed to both interleukin (IL)-4 and tumor necrosis factor (TNF)-α in the absence or presence of SFN treatment. The levels of thymus- and activation-regulated chemokine (TARC) and eotaxin-1 in culture supernatants were evaluated using enzyme-linked immunosorbent assay (ELISA). Reverse transcription-polymerase chain reaction analysis (RT-PCR) enabled quantification of mRNA levels of vascular cell adhesion molecule (VCAM)-1, eotaxin-1, and TARC along with cytokine receptors. An immunoblotting assay was used to evaluate the activities of VCAM-1, nuclear factor-kappa B (NF-κB), mitogen-activated protein kinases (MAPKs), signal transducer and activator of transcription factor (STAT)6 pathways, along with the expression of the cytokine receptors including IL-4 receptor (R)α, IL-13Rα1, TNFRI, as well as TNFRII. SFN inhibited TARC and eotaxin-1 release in HCFs stimulated by TNF-α and IL-4 in a manner dependent on dose and time. SFN suppressed transcriptions of TARC, eotaxin-1, and VCAM-1. Furthermore, the mRNA and protein expression levels of IL-4Rα, TNFRI, and TNFRII were also attenuated by SFN exposure, however, those of IL-13Rα1 remained unaffected. In addition, SFN downregulated the expression of VCAM-1 and the phosphorylation of MAPKs, IκBα, and STAT6. These results suggest that SFN inhibited cytokine-stimulated TARC, eotaxin-1 secretion as well as VCAM-1 expression in HCFs, with these effects likely occurring as a result of cytokine receptor inhibition and attenuation of MAPK, NF-κB, and STAT6 signaling. SFN may therefore have therapeutic potential in VKC treatment.
摘要：
趋化因子和粘附分子是慢性和复发性春季角膜结膜炎（VKC）的主要炎症介质。萝卜硫烷（SFN）是一种天然植物提取物，已知具有抗炎和抗氧化特性。SFN被证明对多种人类疾病有效。当前的研究检查了SFN对细胞因子诱导的人角膜成纤维细胞（HCFs）粘附分子和趋化因子表达的影响和分子机制。在不存在或存在SFN治疗的情况下，将HCF暴露于白介素（IL）-4和肿瘤坏死因子（TNF）-α。使用酶联免疫吸附测定（ELISA）评估培养上清液中胸腺嘧啶和活化调节趋化因子（TARC）和eotaxin-1的水平。逆转录聚合酶链反应分析（RT-PCR）可以定量血管细胞粘附分子（VCAM）-1，eotaxin-1和TARC以及细胞因子受体的mRNA水平。免疫印迹法用于评估VCAM-1，核因子-κB（NF-κB）的活性，丝裂原活化蛋白激酶（MAPK），信号转导和转录因子激活因子（STAT）6个途径，随着细胞因子受体包括IL-4受体（R）α的表达，IL-13Rα1，TNFRI，以及TNFRII。SFN抑制TNF-α和IL-4刺激的HCFs中TARC和eotaxin-1的释放，其方式取决于剂量和时间。SFN抑制了TARC的转录，eotaxin-1和VCAM-1。此外，IL-4Rα的mRNA和蛋白表达水平,TNFRI,和TNFRII也被SFN暴露衰减，然而,IL-13Rα1未受影响。此外,SFN下调VCAM-1的表达和MAPK的磷酸化，IκBα,STAT6这些结果表明，SFN抑制细胞因子刺激的TARC，eotaxin-1分泌以及VCAM-1在HCFs中的表达，这些效应可能是细胞因子受体抑制和MAPK减弱的结果，NF-κB,和STAT6信令。因此,SFN在VKC治疗中可能具有治疗潜力。
