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Recently, the complexity of lung organoid models derived from adult mouse and human stem cells has increased substantially in terms of cellular composition and structural complexity. However, a murine lung organoid system with a clear integrated endothelial compartment is still missing. Here, we describe a novel method that adds another level of intricacy to our published bronchioalveolar lung organoid (BALO) model by microinjection of FACS-sorted lung endothelial cells (ECs) into differentiated organoid cultures. Before microinjection, ECs obtained from the lung homogenate of young mice expressed typical EC markers such as CD31 and vascular endothelial cadherin and showed tube formation capacity. Following microinjection, ECs surrounded the BALO\u2019s alveolar-like compartment, aligning with type I and type II alveolar epithelial cells, as demonstrated by confocal and electron microscopy. Notably, expression of Car4 and Aplnr was as well detected, suggesting the presence of EC microvascular phenotypes in the cultured ECs. Moreover, upon epithelial cell injury by LPS and influenza A virus, endothelialized BALOs released proinflammatory cytokines, leading to the upregulation ICAM-1 (intercellular adhesion molecule 1) in ECs. In summary, we characterized for the first time an organoid model that incorporates ECs into the alveolar structures of lung organoids, not only increasing our previous model\u2019s cellular and structural complexity but also providing a suitable niche to model lung endothelium responses to injury ex vivo.<\/jats:p>","DOI":"10.1165\/rcmb.2023-0373ma","type":"journal-article","created":{"date-parts":[[2024,9,3]],"date-time":"2024-09-03T12:28:45Z","timestamp":1725366525000},"page":"124-132","update-policy":"https:\/\/doi.org\/10.1164\/crossmarkpolicies","source":"Crossref","is-referenced-by-count":19,"title":["Endothelialized Bronchioalveolar Lung Organoids Model Endothelial Cell Responses to Injury"],"prefix":"10.1093","volume":"72","author":[{"given":"Anna-Lena","family":"Ament","sequence":"first","affiliation":[{"name":"Department of Medicine V (Internal Medicine, Infectious Diseases and Infection Control), Universities of Giessen and Marburg Lung Center (UGMLC), Giessen, 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