Corneal epithelial basement membrane assembly is mediated by epithelial cells in coordination with corneal fibroblasts during wound healing.

IF 1.8 3区医学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY

Molecular Vision Pub Date : 2023-05-20 eCollection Date: 2023-01-01

Thomas Michael Shiju, Lycia Pedral Sampaio, Guilherme S L Hilgert, Steven E Wilson

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引用次数: 0

Abstract

Purpose: To understand which cell types, either alone or in combination, contribute to the assembly of the epithelial basement membrane (BM) during corneal wound healing.

Methods: A 3D corneal organotypic model and an in situ rabbit photorefractive keratectomy (PRK) model were used in this study. The 3D corneal organotypic model was established by culturing the rabbit corneal epithelial cells with either corneal fibroblasts or myofibroblasts embedded in collagen type I for 18 days. Corneal fibroblasts were isolated from fresh rabbit corneas, and the myofibroblasts were derived either directly from bone marrow or differentiated from corneal fibroblasts. Immunocytochemistry for alpha-smooth muscle actin (SMA), vimentin, desmin, and vinculin markers confirmed well-differentiated myofibroblasts. Immunohistochemistry was performed in cryofixed sections for BM markers, including laminin alpha-5, laminin beta-3, perlecan, nidogen-1, and collagen type IV. Specimens were also examined with transmission electron microscopy (TEM). Corneas were collected from rabbits after -3 diopter (D) PRK at different time points after surgery, with four corneas at each time point in each group. Cryofixed corneal sections were stained for vimentin, alpha-SMA, and nidogen-1.

Results: The formation of an epithelial BM with expression of laminin alpha-5, laminin beta-3, perlecan, nidogen-1, and collagen IV was observed at the interface between the corneal epithelial cells and corneal fibroblasts. TEM images further confirmed the presence of epithelial BM in organotypic cultures of epithelial cells and corneal fibroblasts. No epithelial BM was observed in cultures of corneal epithelial cells and myofibroblasts (cornea or bone marrow derived), corneal epithelial cells alone, or corneal fibroblasts alone. In rabbit corneas after -3D PRK, a strong association was observed between the regenerating epithelial BM and the presence of corneal fibroblasts at the site of epithelial BM generation.

Conclusions: The corneal epithelial BM assembly is mediated by epithelial cells in coordination with corneal fibroblasts during wound healing.

Abstract Image

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在伤口愈合过程中，角膜上皮基底膜的组装是由上皮细胞与角膜成纤维细胞协调进行的。

目的：了解在角膜伤口愈合过程中，哪些细胞类型（单独或联合）有助于上皮基底膜（BM）的组装：方法：本研究使用了三维角膜器官型模型和原位兔光屈光性角膜切除术（PRK）模型。三维角膜器官型模型是通过将兔角膜上皮细胞与角膜成纤维细胞或肌成纤维细胞包埋在 I 型胶原蛋白中培养 18 天而建立的。角膜成纤维细胞是从新鲜兔角膜中分离出来的，而肌成纤维细胞则直接来源于骨髓或从角膜成纤维细胞分化而来。α-平滑肌肌动蛋白（SMA）、波形蛋白、desmin 和 vinculin 标记的免疫细胞化学证实了肌成纤维细胞分化良好。对冷冻固定切片进行了免疫组化，以检测生物膜标记物，包括层粘蛋白α-5、层粘蛋白β-3、perlecan、nidogen-1和IV型胶原。标本还通过透射电子显微镜（TEM）进行了检查。在手术后的不同时间点采集-3屈光度（D）角膜屈光手术后兔子的角膜，每组在每个时间点采集四个角膜。对冷冻固定的角膜切片进行波形蛋白、α-SMA 和 nidogen-1 染色：结果：在角膜上皮细胞和角膜成纤维细胞的交界处观察到上皮BM的形成，其中表达了层粘连蛋白α-5、层粘连蛋白β-3、perlecan、nidogen-1和胶原蛋白IV。TEM 图像进一步证实了上皮细胞和角膜成纤维细胞的器官型培养物中存在上皮基质。在角膜上皮细胞和肌成纤维细胞（角膜或骨髓来源）、单独角膜上皮细胞或单独角膜成纤维细胞的培养液中均未观察到上皮基质。在-3D PRK术后的兔角膜中，观察到再生的上皮基质与上皮基质生成部位的角膜成纤维细胞之间存在密切联系：结论：在伤口愈合过程中，角膜上皮细胞与角膜成纤维细胞共同参与角膜上皮基质的形成。

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来源期刊

Molecular Vision 生物-生化与分子生物学

CiteScore

4.40

自引率

0.00%

发文量

审稿时长

1 months

期刊介绍： Molecular Vision is a peer-reviewed journal dedicated to the dissemination of research results in molecular biology, cell biology, and the genetics of the visual system (ocular and cortical). Molecular Vision publishes articles presenting original research that has not previously been published and comprehensive articles reviewing the current status of a particular field or topic. Submissions to Molecular Vision are subjected to rigorous peer review. Molecular Vision does NOT publish preprints. For authors, Molecular Vision provides a rapid means of communicating important results. Access to Molecular Vision is free and unrestricted, allowing the widest possible audience for your article. Digital publishing allows you to use color images freely (and without fees). Additionally, you may publish animations, sounds, or other supplementary information that clarifies or supports your article. Each of the authors of an article may also list an electronic mail address (which will be updated upon request) to give interested readers easy access to authors.