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Our objective is to optimize a method for the isolation and culture of primary bronchial epithelial cells (PBECs) and to provide an ex vivo model to study mechanisms of epithelial airway inflammation. PBECs were isolated and cultured from the airways of calves in a submerged cell culture and liquid\u2013liquid interface system. A higher yield and cell viability were obtained after stripping the epithelium from the bronchial section compared to cutting the bronchial section in smaller pieces prior to digestion. <jats:italic>Mannheimia haemolytica<\/jats:italic> and lipopolysaccharide (LPS) as stimulants increased inflammatory responses (IL-8, IL-6 and TNF-\u03b1 release), possibly, by the activation of \"TLR-mediated MAPKs and NF-\u03baB\" signaling. Furthermore, <jats:italic>M. haemolytica<\/jats:italic> and LPS disrupted the bronchial epithelial layer as observed by a decreased transepithelial electrical resistance and zonula occludens-1 and E-cadherin expression. An optimized isolation and culture method for calf PBECs was developed, which cooperated with animal use Replacement, Reduction and Refinement (3R's) principle, and can also contribute to the increased knowledge and development of effective therapies for other animal and humans (childhood) respiratory diseases.<\/jats:p>","DOI":"10.1038\/s41598-020-69982-0","type":"journal-article","created":{"date-parts":[[2020,8,3]],"date-time":"2020-08-03T10:04:58Z","timestamp":1596449098000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["Mannheimia haemolytica and lipopolysaccharide induce airway epithelial inflammatory responses in an extensively developed ex vivo calf model"],"prefix":"10.1038","volume":"10","author":[{"given":"Yang","family":"Cai","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Soheil","family":"Varasteh","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jos P. 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