IRF4 affects the protective effect of regulatory T cells on the pulmonary vasculature of a bronchopulmonary dysplasia mouse model by regulating FOXP3.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is a common chronic lung disease in preterm infants, characterised by compromised alveolar development and pulmonary vascular abnormalities. Emerging evidence suggests that regulatory T cells (Tregs) may confer protective effects on the vasculature. Knockdown of their transcription factor, interferon regulatory factor 4 (IRF4), has been shown to promote vascular endothelial hyperplasia. However, the involvement of Tregs and IRF4 in the BPD pathogenesis remains unclear. This study aimed to investigate the regulation of Tregs by IRF4 and elucidate its potential role in pulmonary vasculature development in a BPD mouse model. METHODS: The BPD model was established using 85% hyperoxia exposure, with air exposure as the normal control. Lung tissues were collected after 7 or 14 days of air or hyperoxia exposure, respectively. Haematoxylin-eosin staining was performed to assess lung tissue pathology. Immunohistochemistry was used to measure platelet endothelial cell adhesion molecule-1 (PECAM-1) level, flow cytometry to quantify Treg numbers, and Western blot to assess vascular endothelial growth factor (VEGFA), angiopoietin-1 (Ang-1), forkhead box protein P3 (FOXP3), and IRF4 protein levels. We also examined the co-expression of IRF4 and FOXP3 proteins using immunoprecipitation and immunofluorescence double staining. Furthermore, we employed CRISPR/Cas9 technology to knock down the IRF4 gene and observed changes in the aforementioned indicators to validate its effect on pulmonary vasculature development in mice. RESULTS: Elevated IRF4 levels in BPD model mice led to FOXP3 downregulation, reduced Treg numbers, and impaired pulmonary vascular development. Knockdown of IRF4 resulted in improved pulmonary vascular development and upregulated FOXP3 level. CONCLUSION: IRF4 may affect the protective role of Tregs in the proliferation of pulmonary vascular endothelial cells and pulmonary vascular development in BPD model mice by inhibiting the FOXP3 level.

IL-33-ST2 pathway regulates AECII transdifferentiation by targeting alveolar macrophage in a bronchopulmonary dysplasia mouse model.

Evidence points to the indispensable function of alveolar macrophages (AMs) in normal lung development and tissue homeostasis. However, the importance of AMs in bronchopulmonary dysplasia (BPD) has not been elucidated. Here, we identified a significant role of abnormal AM proliferation and polarization in alveolar dysplasia during BPD, which is closely related to the activation of the IL-33-ST2 pathway. Compared with the control BPD group, AMs depletion partially abolished the epithelialmesenchymal transition process of AECII and alleviated pulmonary differentiation arrest. In addition, IL-33 or ST2 knockdown has protective effects against lung injury after hyperoxia, which is associated with reduced AM polarization and proliferation. The protective effect disappeared following reconstitution of AMs in injured IL-33 knockdown mice, and the differentiation of lung epithelium was blocked again. In conclusion, the IL-33-ST2 pathway regulates AECII transdifferentiation by targeting AMs proliferation and polarization in BPD, which shows a novel strategy for manipulating the IL-33-ST2-AMs axis for the diagnosis and intervention of BPD.

Type 2 innate lymphoid cell-derived amphiregulin regulates type II alveolar epithelial cell transdifferentiation in a mouse model of bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD) is a common complication in preterm infants characterized by alveolar growth arrest. Interleukin (IL)-33 and type 2 innate lymphoid cell (ILC2) affect type II alveolar epithelial cell (AECII) differentiation in BPD mice and may cause increased lung epithelial-mesenchymal transition (EMT). Amphiregulin (AREG) can be produced by ILC2 and is associated with tissue repair. However, the action mechanism of AREG produced by ILC2 to alveolar development in BPD is unclear. In this study, we aimed to demonstrate the role and mechanism of AREG in influencing AECII transdifferentiation in the lung tissue of BPD mice. The effects of ILC2-derived AREG on AECII transdifferentiation were verified in vivo and in vitro, and the role of IL-33 on ILC2-derived AREG in AECII transdifferentiation in BPD mice and a preliminary investigation of the role of AREG's receptor-epidermal growth factor receptor (EGFR) on AECII transdifferentiation. The results showed that neonatal mice developed severe lung injury after hyperoxia, and IL-33 induced AREG production via ILC2 affected normal AECII differentiation and promoted EMT. In addition, the blockade of EGFR was found to alleviate the impaired AECII differentiation under hyperoxia in an in vitro study. In summary, our study demonstrates that AREG secreted by ILC2 affects AECII transdifferentiation in BPD mice, which provides a new idea for the clinical treatment of BPD.