Background: Neonatal respiratory system disease is closely associated with embryonic lung development. Our group found that Integrin β4(ITGB4) is downregulated in the airway epithelium of asthma patients. Asthma is the most common chronic respiratory illness in childhood. Therefore, we suspect whether the deletion of ITGB4 would affect fetal lung development? In this study, we characterized the role of ITGB4 deficiency in Bronchopulmonary dysplasia (BPD).
Methods: ITGB4 was conditionally knocked out in CCSP-rtTA; Tet-O-Cre; ITGB4f/f triple transgenic mice administrating doxycycline from E7.5 to P42. Lung tissues at different developmental stages were collected for experimental detection and transcriptome sequencing. The expression of lung-development-relevant factors were evaluated by transcriptome sequencing, WB and Immunohistochemical. Bronchoalveolar lavage fluid were collected to perform Total cell and differential inflammatory cell counts. The effects of ITGB4 deficiency on lung branching morphogenesis were observed by fetal mouse lung explants culture.
Results: We found that β4CCSP.Cre mice exhibit significant lighter body weight comparing to age-matched littermate control β4f/f, and the difference became more distinct as their age increase. ITGB4 deficiency shows no significant impact on the ratio of lung weight to body weight and the overall survival rates of mice. Deleting ITGB4 from the airway epithelial cells result in enlarged alveolar airspaces, inhibition of branching, the overlapping of epithelium cells and the impairment of cilial growth during lung development. Scanning electron microscopy showed that the airway epithelial cilia of β4CCSP.Cre group appear to be sparse, shortened, and lodging. Lung-development-relevant factors such as SPTPC、SOX2 significantly decreased in both mRNA and protein level. KEGG pathway analysis indicated that multiple ontogenesis-regulating-relevant pathway converge to FAK, accordingly, ITGB4 deletion decreased phospho-FAK, phospho-GSK3β, SOX2 level, and correspondingly contrary consequence was detected after treatment with GSK3β agonist（wortmannin）. Airway branching defect of β4CCSP.Cre mice lung explants was also partly recovered after wortmannin treatment.
Conclusions: Airway epithelial-specific deletion of ITGB4 contributes to lung developmental defect, which could be achieved through the FAK/GSK3β/SOX2 signal pathway.