Table3_A Pulmonary Vascular Model From Endothelialized Whole Organ Scaffolds.XLSX

The development of an in vitro system for the study of lung vascular disease is critical to understanding human pathologies. Conventional culture systems fail to fully recapitulate native microenvironmental conditions and are typically limited in their ability to represent human pathophysiology for the study of disease and drug mechanisms. Whole organ decellularization provides a means to developing a construct that recapitulates structural, mechanical, and biological features of a complete vascular structure. Here, we developed a culture protocol to improve endothelial cell coverage in whole lung scaffolds and used single-cell RNA-sequencing analysis to explore the impact of decellularized whole lung scaffolds on endothelial phenotypes and functions in a biomimetic bioreactor system. Intriguingly, we found that the phenotype and functional signals of primary pulmonary microvascular revert back—at least partially—toward native lung endothelium. Additionally, human induced pluripotent stem cell-derived endothelium cultured in decellularized lung systems start to gain various native human endothelial phenotypes. Vascular barrier function was partially restored, while small capillaries remained patent in endothelial cell-repopulated lungs. To evaluate the ability of the engineered endothelium to modulate permeability in response to exogenous stimuli, lipopolysaccharide (LPS) was introduced into repopulated lungs to simulate acute lung injury. After LPS treatment, proinflammatory signals were significantly increased and the vascular barrier was impaired. Taken together, these results demonstrate a novel platform that recapitulates some pulmonary microvascular functions and phenotypes at a whole organ level. This development may help pave the way for using the whole organ engineering approach to model vascular diseases.

研究肺部血管疾病体外模型的开发对于理解人类病理学至关重要。传统的培养系统无法完全重现原生的微环境条件，且通常在代表人类病理生理学以研究疾病和药物机制方面存在局限性。整体器官去细胞化提供了一种方法，用以构建能够重现完整血管结构结构、机械和生物学特征的构建体。在此，我们开发了一种培养方案，以改善整个肺支架中的内皮细胞覆盖度，并利用单细胞RNA测序分析探索去细胞化整体肺支架对生物模拟生物反应器系统中内皮表型和功能的影响。引人入胜的是，我们发现原发性肺微血管的表型和功能信号至少部分地回归到原生的肺内皮。此外，源自人诱导多能干细胞培养的去细胞化肺系统中的内皮开始获得各种原代人内皮表型。血管屏障功能部分恢复，而在内皮细胞再植的肺中，小血管保持通畅。为了评估工程化内皮调节通透性的能力，对再植肺引入了脂多糖（LPS），以模拟急性肺损伤。在LPS处理后，促炎信号显著增加，血管屏障受损。综上所述，这些结果展示了一个新的平台，能够在整体器官水平上重现某些肺微血管的功能和表型。这一发展可能有助于为使用整体器官工程方法来模拟血管疾病铺平道路。