Human Corneal Fibrosis: An In Vitro Model
Human Corneal Fibrosis: An In Vitro Model
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DOI：
10.1167/iovs.09-3860
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发表时间：
2010-03-01
影响因子：
4.4
通讯作者：
Zieske, James D.
中科院分区：
文献类型：
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作者：
Karamichos, Dimitris;Guo, Xiaoqing Q.;Zieske, James D.
目的。角膜损伤最终可能通过角膜纤维化形成疤痕，其特征是肌成纤维细胞的存在和细胞外基质（ECM）成分的不当沉积。已知tgf - β 1可刺激ECM成分的过度产生和沉积。此前，利用稳定的维生素C （VitC）刺激原代人角膜成纤维细胞（HCFs），建立了角膜基质的体外三维（3d）模型。这个模型模拟了角膜的发育。作者推测，在tgf - β 1的加入下，可以生成三维角膜瘢痕模型。hcf在四种培养基条件下生长4或8周：仅维生素c；维生素c + tgf - β 1；维生素c + tgf - β 1治疗1周，然后维生素c仅治疗3或7周；维生素c 4周，然后维生素c + tgf - β 1 4周。用透射电镜和间接免疫荧光分析培养物。与对照组相比，tgf - β 1的添加显著增加了构建体的厚度，在第4周和第8周，tgf - β 1全程存在的构建体的最大增幅分别为2.1至3.2倍。在所有tgf - β处理的培养中，细胞变得长而扁平，可见大量丝状细胞，胶原蛋白水平增加，可见长胶原原纤维。平滑肌肌动蛋白、细胞纤维连接蛋白和III型胶原蛋白表达均增加。第4周和第8周的培养显示出极小的差异。受维生素c和tgf - β 1刺激的人角膜成纤维细胞似乎产生了一个类似于在人角膜纤维化中观察到的过程的模型。该模型可用于检查伤口愈合情况下基质的沉积和组装。（中国眼科杂志，2010;51:1382-1388）DOI:10.1167/iovs.09-3860
PURPOSE. Corneal injury may ultimately lead to a scar by way of corneal fibrosis, which is characterized by the presence of myofibroblasts and improper deposition of extracellular matrix (ECM) components. TGF-beta 1 is known to stimulate overproduction and deposition of ECM components. Previously, an in vitro three-dimensional (3-D) model of a corneal stroma was developed by using primary human corneal fibroblasts (HCFs) stimulated with stable vitamin C (VitC). This model mimics corneal development. The authors postulate that with the addition of TGF-beta 1, a 3-D corneal scar model can be generated.METHODS. HCFs were grown in four media conditions for 4 or 8 weeks: VitC only; VitC+TGF-beta 1 for the entire time; VitC+TGF-beta 1 for 1 week, then VitC only for 3 or 7 weeks; and VitC for 4 weeks, then VitC+TGF-beta 1 for 4 weeks. Cultures were analyzed with TEM and indirect immunofluorescence.RESULTS. Compared with the control, addition of TGF-beta 1 increased construct thickness significantly, with maximum increase in constructs with TGF-beta 1 present for the entire time-2.1- to 3.2-fold at 4 and 8 weeks, respectively. In all TGF-beta-treated cultures, cells became long and flat, numerous filamentous cells were seen, collagen levels increased, and long collagen fibrils were visible. Smooth muscle actin, cellular fibronectin, and type III collagen expression all appeared to increase. Cultures between weeks 4 and 8 showed minimal differences.CONCLUSIONS. Human corneal fibroblasts stimulated by VitC and TGF-beta 1 appear to generate a model that resembles processes observed in human corneal fibrosis. This model should be useful in examining matrix deposition and assembly in a wound-healing situation. (Invest Ophthalmol Vis Sci. 2010;51:1382-1388) DOI:10.1167/iovs.09-3860