IOBA-University of Valladolid, Campus Miguel Delibes, Paseo de Belén 17, Valladolid E-47011, Spain.
Exp Eye Res. 2013 Feb;107:88-100. doi: 10.1016/j.exer.2012.11.017. Epub 2012 Dec 7.
A role for transforming growth factor (TGF)-β in the pathogenesis of some ocular surface diseases has been proposed. We determined if secretion of TGF-β and expression of TGF-β receptors RI, RII, and RIII by human ocular surface epithelial cells were modified under inflammatory conditions. We also determined how these cells responded to TGF-β. A human corneal epithelial (HCE) cell line and a conjunctival epithelial cell line (IOBA-NHC) were exposed to TGF-β1 and -β2 and to proinflammatory cytokines. TGF-β receptor mRNAs were analyzed by real time reverse transcription polymerase chain reaction (RT-PCR) in both cell lines, and in conjunctival, limbal, and corneal epithelial cells from post-mortem human specimens. Expression of TGF-β receptors and pSMAD2/SMAD2 were determined by Western blot and immunofluorescence assays. Secretion of TGF-β isoforms, cytokine/chemokine, and metalloproteinases (MMPs) were analyzed in cell supernatants by immunobead-based assays. Secretory leukocyte proteinase inhibitor (SLPI) secretion was analyzed by enzyme-linked immunosorbent assay. TGF-β isoform and receptor gene expression was determined by RT-PCR in conjunctival epithelium of dry eye (DE) patients and healthy subjects. Our results showed that TGF-β RI expression was down-regulated with IL-4 exposure, whereas TGF-β RII and TGF-β2 were upregulated by TNF-α in HCE cells. TGF-β RIII receptor expression was upregulated in IOBA-NHC cells by TNF-α and IFN-γ. SMAD2 phosphorylation occurred in HCE and IOBA-NHC cells after TGF-β treatment. TGF-β significantly up- and down-regulated secretion of several cytokines/chemokines by both cell lines and MMP by HCE cells. TGF-β2 and TGF-β3 were upregulated and TGF-β RIII mRNA was down-regulated in DE conjunctival epithelium. These results show that TGF-β plays an important role in directing local inflammatory responses in ocular surface epithelial cells.
转化生长因子 (TGF)-β 在某些眼表疾病发病机制中的作用已被提出。我们确定了人眼表面上皮细胞在炎症条件下分泌 TGF-β 以及表达 TGF-β 受体 RI、RII 和 RIII 的情况是否发生了改变。我们还确定了这些细胞对 TGF-β 的反应方式。我们用 TGF-β1 和 -β2 以及促炎细胞因子处理人角膜上皮 (HCE) 细胞系和结膜上皮细胞系 (IOBA-NHC)。我们通过实时逆转录聚合酶链反应 (RT-PCR) 分析了这两种细胞系以及来自死后人类标本的结膜、角膜缘和角膜上皮细胞中的 TGF-β 受体 mRNA。通过 Western blot 和免疫荧光测定法分析了 TGF-β 受体和 pSMAD2/SMAD2 的表达。通过免疫珠基检测法分析了细胞上清液中 TGF-β 同工型、细胞因子/趋化因子和金属蛋白酶 (MMP) 的分泌情况。通过酶联免疫吸附试验分析了分泌白细胞蛋白酶抑制剂 (SLPI) 的分泌情况。我们通过 RT-PCR 确定了干燥性角结膜炎 (DE) 患者和健康受试者结膜上皮中的 TGF-β 同工型和受体基因表达情况。我们的结果表明，HCE 细胞中 IL-4 暴露可下调 TGF-β RI 表达，而 TNF-α 可上调 TGF-β RII 和 TGF-β2。TNF-α 和 IFN-γ 可使 IOBA-NHC 细胞中 TGF-β RIII 受体表达上调。TGF-β 处理后 HCE 和 IOBA-NHC 细胞中的 SMAD2 发生磷酸化。TGF-β 显著上调和下调了这两种细胞系的几种细胞因子/趋化因子和 HCE 细胞的 MMP 的分泌。DE 结膜上皮中 TGF-β2 和 TGF-β3 上调，TGF-β RIII mRNA 下调。这些结果表明，TGF-β 在指导眼表面上皮细胞中的局部炎症反应方面发挥着重要作用。
