Purpose

Hydrogels derived from decellularized tissues provide superior biocompatibility, tenability and tissue-specific extracellular matrix (ECM) components. Based on the preparation of decellularized porcine cornea (DPC), here we developed an injectable and transparent hydrogel for the regeneration of epithelium and stroma in focal corneal defects.

Methods

The DPC-derived hydrogel was prepared with N-cyclohexyl-N′-(2-morpholinethyl) carbodiimide metho-p-toluenesulfonate/N-hydroxysuccinimide (CMC/NHS) as cross-linkers. The characteristics of the hydrogel were analyzed and its cytocompatibility was assessed by Live/Dead and Cell Counting Kit (CCK)-8 assays. Immunofluorescence staining, quantitative PCR and Western blot analyses were performed to assess the relative protein and gene expression in corneal fibroblasts on hydrogel. The safety and efficiency of the hydrogel for repairing focal corneal defects in rabbit were measured by slit-lamp, anterior segment optical coherence tomography (AS-OCT), confocal microscopy and histological analyses.

Results

The DPC-derived hydrogel cross-linked with CMC/NHS assumed favorable transparency, exhibited distinct mechanical properties and preserved the ECM components of native porcine cornea (NPC). In vitro experiments showed that the hydrogel maintained the phenotype, supported the proliferation and promoted the ECM synthesis of corneal fibroblasts. When injected onto rabbit corneas, the hydrogel rapidly covered, solidified and formed a smooth surface on the focal defect. Corneal epithelium was fully regenerated within 3 days. The thickness of the corneal epithelium and stroma was restored at 12 weeks after surgery without significant inflammation or scar formation. Notably, the hydrogel showed no harmful effects on the resident stroma and endothelium.

Conclusions

The DPC-derived hydrogel may represent a promising biomaterial for corneal epithelial and stromal regeneration.

中文翻译：

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去细胞猪角膜衍生水凝胶，用于局灶性角膜缺损中上皮和基质的再生。

目的

衍生自脱细胞组织的水凝胶具有出色的生物相容性，韧性和组织特异性细胞外基质（ECM）成分。在制备脱细胞猪角膜（DPC）的基础上，我们在这里开发了一种可注射的透明水凝胶，用于再生局灶性角膜缺损中的上皮和基质。

方法

DPC衍生的水凝胶是用N-环己基-N' -（2-吗啉基乙基）碳二亚胺，甲基-对甲苯磺酸盐/ N-羟基琥珀酰亚胺（CMC / NHS）作为交联剂制备的。分析了水凝胶的特性，并通过活/死和细胞计数试剂盒（CCK）-8分析评估了其细胞相容性。进行了免疫荧光染色，定量PCR和蛋白质印迹分析，以评估水凝胶上角膜成纤维细胞中的相对蛋白质和基因表达。通过裂隙灯，前节光学相干断层扫描（AS-OCT），共聚焦显微镜和组织学分析来测量水凝胶修复兔局灶性角膜缺损的安全性和效率。

结果

与CMC / NHS交联的DPC衍生的水凝胶具有良好的透明性，表现出独特的机械性能，并保留了天然猪角膜（NPC）的ECM成分。体外实验表明，水凝胶保持表型，支持增殖并促进角膜成纤维细胞的ECM合成。当注射到兔角膜上时，水凝胶迅速覆盖，固化并在局灶性缺损处形成光滑表面。角膜上皮在3天内完全再生。术后12周角膜上皮和基质的厚度恢复，没有明显的炎症或疤痕形成。值得注意的是，水凝胶对驻留的基质和内皮没有有害作用。

结论

DPC衍生的水凝胶可能代表了有希望的角膜上皮和基质再生的生物材料。