School of Pharmacy, University of Reading, Reading, UK.
Tissue Eng Part A. 2010 Jun;16(6):2091-100. doi: 10.1089/ten.TEA.2009.0748.
We have investigated the use of a laminin-coated compressed collagen gel containing corneal fibroblasts (keratocytes) as a novel scaffold to support the growth of corneal limbal epithelial stem cells. The growth of limbal epithelial cells was compared between compressed collagen gel and a clinically proven conventional substrate, denuded amniotic membrane (AM). Following compression of the collagen gel, encapsulated keratocytes remained viable and scanning electron microscopy showed that fibers within the compressed gel were dense, homogeneous, and similar in structure to those within denuded AM. Limbal epithelial cells were successfully expanded upon the compressed collagen, resulting in stratified layers of cells containing desmosome and hemidesmosome structures. The resulting corneal constructs of both the groups shared a high degree of transparency, cell morphology, and cell stratification. Similar protein expression profiles for cytokeratin 3 (CK3) and CK14 and no significant difference in CK12 mRNA expression levels by real-time polymerase chain reaction were also observed. This study provides the first line of evidence that a laminin-coated compressed collagen gel containing keratocytes can adequately support limbal epithelial cell expansion, stratification, and differentiation to a degree that is comparable to the leading conventional scaffold, denuded AM.
我们研究了使用含有角膜成纤维细胞（角膜基质细胞）的层粘连蛋白包被的压缩胶原凝胶作为一种新型支架，以支持角膜缘上皮干细胞的生长。比较了压缩胶原凝胶和临床上已证实的传统基质，即脱细胞羊膜（AM），对角膜缘上皮细胞生长的影响。胶原凝胶压缩后，包裹的角膜基质细胞仍然存活，扫描电子显微镜显示压缩凝胶内的纤维密集、均匀，结构与脱细胞 AM 内的纤维相似。角膜缘上皮细胞在压缩胶原上成功扩增，形成具有桥粒和半桥粒结构的分层细胞。两组的角膜构建体都具有高度的透明度、细胞形态和细胞分层。实时聚合酶链反应也观察到细胞角蛋白 3（CK3）和 CK14 的相似蛋白表达谱，以及 CK12mRNA 表达水平无显著差异。这项研究提供了第一个证据，表明含有角膜基质细胞的层粘连蛋白包被的压缩胶原凝胶可以充分支持角膜缘上皮细胞的扩增、分层和分化，达到与主要的传统支架脱细胞 AM 相当的程度。
