Chirco Kathleen R, Worthington Kristan S, Flamme-Wiese Miles J, Riker Megan J, Andrade Joshua D, Ueberheide Beatrix M, Stone Edwin M, Tucker Budd A, Mullins Robert F
The Stephen A. Wynn Institute for Vision Research, The University of Iowa, Iowa City, IA 52246, USA; Department of Ophthalmology and Visual Sciences, The University of Iowa, Iowa City, IA 52246, USA.
Proteomics Laboratory, New York University School of Medicine, New York, NY 10016, USA.
Acta Biomater. 2017 Jul 15;57:293-303. doi: 10.1016/j.actbio.2017.05.011. Epub 2017 May 5.
Endothelial cells (ECs) of the choriocapillaris are one of the first cell types lost during age-related macular degeneration (AMD), and cell replacement therapy is currently a very promising option for patients with advanced AMD. We sought to develop a reliable method for the production of human choroidal extracellular matrix (ECM) scaffolds, which will allow for the study of choroidal EC (CEC) replacement strategies in an environment that closely resembles the native tissue. Human RPE/choroid tissue was treated sequentially with Triton X-100, SDS, and DNase to remove all native cells. While all cells were successfully removed from the tissue, collagen IV, elastin, and laminin remained, with preserved architecture of the acellular vascular tubes. The ECM scaffolds were then co-cultured with exogenous ECs to determine if the tissue can support cell growth and allow EC reintegration into the decellularized choroidal vasculature. Both monkey and human ECs took up residence in the choriocapillary tubes of the decellularized tissue. Together, these data suggest that our decellularization methods are sufficient to remove all cellular material yet gentle enough to preserve tissue structure and allow for the optimization of cell replacement strategies.
Age-related macular degeneration (AMD) is a devastating disease affecting more than 600 million people worldwide. Endothelial cells of the choriocapillaris (CECs) are among the first cell types lost in early AMD, and cell replacement therapy is currently the most promising option for restoring vision in patients with advanced AMD. In order to study CEC replacement strategies we have generated a 3D choroid scaffold using a novel decellularization method in human RPE/choroid tissue. To our knowledge, this is the first report describing decellularization of human RPE/choroid, as well as recellularization of a choroid scaffold with CECs. This work will aid in our development and optimization of cell replacement strategies using a tissue scaffold that is similar to the in vivo environment.
脉络膜毛细血管的内皮细胞（ECs）是年龄相关性黄斑变性（AMD）过程中最早丢失的细胞类型之一，细胞替代疗法目前是晚期AMD患者非常有前景的选择。我们试图开发一种可靠的方法来生产人脉络膜细胞外基质（ECM）支架，这将有助于在与天然组织非常相似的环境中研究脉络膜内皮细胞（CEC）替代策略。用人RPE/脉络膜组织依次用 Triton X-100、SDS 和 DNase 处理以去除所有天然细胞。虽然所有细胞都成功从组织中去除，但IV型胶原、弹性蛋白和层粘连蛋白保留下来，无细胞血管管的结构得以保留。然后将ECM支架与外源性ECs共培养，以确定该组织是否能支持细胞生长并使EC重新整合到去细胞化的脉络膜血管系统中。猴和人的ECs都在去细胞化组织的脉络膜毛细血管管中定居。总之，这些数据表明我们的去细胞化方法足以去除所有细胞物质，但又足够温和以保留组织结构，并允许优化细胞替代策略。
年龄相关性黄斑变性（AMD）是一种毁灭性疾病，全球有超过6亿人受其影响。脉络膜毛细血管的内皮细胞（CECs）是早期AMD中最早丢失的细胞类型之一，细胞替代疗法目前是晚期AMD患者恢复视力最有前景的选择。为了研究CEC替代策略我们使用一种新的去细胞化方法在人RPE/脉络膜组织中生成了3D脉络膜支架。据我们所知，这是第一份描述人RPE/脉络膜去细胞化以及用CECs对脉络膜支架进行再细胞化的报告。这项工作将有助于我们使用与体内环境相似的组织支架来开发和优化细胞替代策略。
