Gilpin Sarah E, Li Qiyao, Evangelista-Leite Daniele, Ren Xi, Reinhardt Dieter P, Frey Brian L, Ott Harald C
Center for Regenerative Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, United States.
Department of Chemistry, University of Wisconsin-Madison, Madison, WI, United States.
Biomaterials. 2017 Sep;140:212-219. doi: 10.1016/j.biomaterials.2017.06.027. Epub 2017 Jun 22.
Organ engineering based on native matrix scaffolds involves combining regenerative cell populations with corresponding biological matrices to form functional grafts on-demand. The extracellular matrix (ECM) that is retained following lung decellularization provides essential structure and biophysical cues for whole organ regeneration after recellularization. The unique ECM composition in the early post-natal lung, during active alveologenesis, may possess distinct signals that aid in driving cell adhesion, survival, and proliferation. We evaluated the behavior of basal epithelial stem cells (BESCs) isolated from adult human lung tissue, when cultured on acellular ECM derived from neonatal (aged < 1 week) or adult lung donors (n = 3 donors per group). A significant difference in cell proliferation and survival was found. We next performed in-depth proteomic analysis of the lung scaffolds to quantify proteins significantly enriched in the neonatal ECM, and identified the glycoproteins Fibrillin-2 (FBN-2) and Tenascin-C (TN-C) as potential mediators of the observed effect. BESCs cultured on Collagen Type IV coated plates, supplemented with FBN-2 and TN-C demonstrated significantly increased proliferation and decreased cellular senescence. No significant increase in epithelial-to-mesenchymal transition was observed. In vitro migration was also increased by FBN-2 and TN-C treatment. Decellularized lung scaffolds treated with FBN-2 and TN-C prior to re-epithelialization supported greater epithelial proliferation and tissue remodeling. BESC distribution, matrix alignment, and overall tissue morphology was improved on treated lung scaffolds, after 3 and 7 days of ex vivo lung culture. These results demonstrate that scaffold re-epithelialization is enhanced on neonatal lung ECM, and that supplementation of FBN-2 and TN-C to the native scaffold may be a valuable tool in lung tissue regeneration.
基于天然基质支架的器官工程涉及将再生细胞群体与相应的生物基质相结合，以按需形成功能性移植物。肺去细胞化后保留的细胞外基质（ECM）为再细胞化后的全器官再生提供了基本结构和生物物理线索。出生后早期肺在活跃的肺泡形成过程中独特的ECM组成可能具有有助于驱动细胞黏附、存活和增殖的独特信号。我们评估了从成人肺组织分离的基底上皮干细胞（BESC）在源自新生儿（年龄<1周）或成人肺供体（每组n = 3个供体）的脱细胞ECM上培养时的行为。发现细胞增殖和存活存在显著差异。接下来，我们对肺支架进行了深入的蛋白质组学分析，以量化在新生儿ECM中显著富集的蛋白质，并确定糖蛋白原纤维蛋白-2（FBN-2）和腱生蛋白-C（TN-C）是观察到的效应的潜在介质。在涂有IV型胶原蛋白的平板上培养并补充FBN-2和TN-C的BESC表现出显著增加的增殖和细胞衰老减少。未观察到上皮-间质转化有显著增加。FBN-2和TN-C处理也增加了体外迁移。在重新上皮化之前用FBN-2和TN-C处理的脱细胞肺支架支持更大程度的上皮增殖和组织重塑。在体外肺培养3天和7天后，处理过的肺支架上的BESC分布、基质排列和整体组织形态得到改善。这些结果表明，新生儿肺ECM上的支架再上皮化得到增强，并且向天然支架中补充FBN-2和TN-C可能是肺组织再生中的一种有价值的工具。
