330 - Pathophysiology features of evolving bronchopulmonary dysplasia: insights from a clinical and translational study

Publication Number: 1314.330

Carlotta Boggi, University of Parma, Parma, Emilia-Romagna, Italy; Chiara Catozzi, Chiesi Farmaceutici Spa, Parma, Emilia-Romagna, Italy; Matteo Storti, Chiesi Farmaceutici, Parma, Emilia-Romagna, Italy; Erica Ferrini, Chiesi Farmaceutici S.p.A., Parma, Emilia-Romagna, Italy; Franco Fabio Stellari, Chiesi Farmaceutici S.p.A., Parma, Emilia-Romagna, Italy; Enrica Scalera, CHIESI FARMACEUTICI S.P.A., Parma, Emilia-Romagna, Italy; Aurora Radicati, Chiesi S.p.A, Parma, Emilia-Romagna, Italy; Giorgio Aquila, Chiesi Farmaceutici Spa, Parma, Emilia-Romagna, Italy; Simone De Meo, University of Siena, Formia, Lazio, Italy; Leonardo Novelli, University of Parma, Santo Stefano di Magra, Liguria, Italy; Angelo Modena, Università del Piemonte Orientale, Parma, Italy, Emilia-Romagna, Italy; Xabier Murguia Esteve, Consultant in Biomedical Science (Freelance), Bilbao, Pais Vasco, Spain; Francesca Ricci, Chiesi Farmaceuitci, PARMA, Emilia-Romagna, Italy; Daniele De Luca, Paris Saclay University-APHP, Paris, Ile-de-France, France

Background: Bronchopulmonary dysplasia (BPD) is a chronic respiratory condition of preterm infants. Its pathophysiology has shifted from ventilator-induced injury to disrupted alveolarization, but the early evolving phase remains poorly characterized, limiting early identification and personalized management.

Objective: We combined clinical and translational approaches to investigate the BPD development in neonates and in a hyperoxia-exposed preterm rabbit model.

Design/Methods: The clinical arm was a secondary analysis of the PATH-BPD cohort (ISRCTN12233815) including preterm infants ≤36 weeks GA. Functional lung imaging with quantitative lung ultrasound (LUS) and weight-indexed thoracic fluid content (TFC) was performed at 10, 21, and 28 days of life, and at 34-36 weeks postconceptional age. Infants received mainly non-invasive respiratory support under a shared care policy. The experimental arm used a validated preterm rabbit model exposed to hyperoxia (70% O₂) from birth to postnatal day (PND) 14. Preterm rabbits were delivered at day 28 of gestation (term 31 days); age-matched term-born pups reared by dams served as controls. Functional micro-CT imaging was performed at two developmental timepoints: T1=PND7 and T2=PND14 in preterm rabbits; T1=PND4 and T2=PND11 in term controls. Lung function (inspiratory capacity [IC], static compliance [Cst], tissue damping [G], and tissue elastance [H]) was assessed with the FlexiVent system at PND14 and PND11, in hyperoxia-exposed preterm and control rabbits, respectively. Morphometric and histopathological analyses, including radial alveolar count (RAC), and acute lung injury (ALI) score were performed on lung sections. In addition, differential gene expression and pathway enrichment analyses were performed on rabbit lung samples.

Results: In infants, extended LUS aeration scores showed significant differences between those who developed BPD and those who did not, with progressive divergence over time (Fig.1), while TFC showed no significant difference. In rabbits, hyperoxia-exposed preterm animals exhibited reduced lung aeration (Fig.2), lower IC and Cst, higher G and H, decreased RAC, higher ALI, and enrichment of inflammatory (upregulated) and morphogenesis-related (downregulated) pathways compared to term controls (Fig.3).

Conclusion(s): This integrative study provides novel insights into the early pathophysiological features of evolving BPD, highlighting measurable functional and structural alterations before clinical diagnosis. These findings support the use of preterm rabbits as a translational model to mimic BPD-like phenotypes and to test new pharmacological therapies.

Figure 1. Functional imaging in infants: ultrasound-assessed lung aeration
Grey and black circles indicate patients with and without BPD, respectively; T-bars indicate two standard errors. Post hoc, between-patients, head-to head significant comparisons were performed by Student test and indicated by the following symbols: *p < 0.001. Lung aeration was evaluated with the dimensionless extended lung ultrasound aeration score, calculated at 10, 21, 28 days of life and 34- and 36-weeks postconceptional age.

Figure 2. Functional imaging in the experimental rabbit model: parameters measured with µCT.
Minute Ventilation (MV), expiratory volume (Vexp), tidal volume (Vt) and the gas volume at end-expiration (Gasexp which represents functional residual capacity) are shown in Panel A, B, C and D, respectively. Grey and black circles indicate hyperoxia-exposed preterm rabbits and term control rabbits, respectively; T-bars indicate standard deviation. Statistical comparisons between the two experimental groups were performed using a two-way ANOVA, followed by Sidak's post-hoc test for multiple comparisons. Significant differences are indicated by vertical bars as follows: * p<0.05, ** p<0.01, *** p<0.001. T1-measurements are performed at 4 and 7 days of life for term and preterm rabbits, respectively; T2-measurements are performed at 11 and 14 days of life for term and preterm rabbits, respectively

Figure 3. Histological, lung function, and molecular analyses of preterm rabbits
(A) Lung mechanics in the experimental rabbit model: parameters were measured with the FlexiVent. Inspiratory capacity, respiratory system compliance (Crs), tissue damping (G) and tissue elastance (H) are shown. Columns and T-bars represent mean and standard deviation, respectively. Black and grey columns represent term control rabbits and hyperoxia-exposed preterm rabbits, respectively; *p < 0.01, **p < 0.001 (Student test). (B) Radial alveolar count (RAC), and acute lung injury (ALI) score in preterm rabbits. Columns and T-bars represent mean and standard deviation, respectively; black and grey represent term control rabbits and hyperoxia-exposed preterm rabbits, respectively; *p < 0.0001 (Student test). (C) Pathway enrichment analysis performed with Gene Set Enrichment Analysis results comparing different conditions. The bar plots display the top 10 upregulated and downregulated pathways when comparing preterm-hyperoxic and normoxic rabbits. The normalized enrichment score (NES) is shown on the x-axis, indicating pathway activation or suppression; the scale gradient color indicates the adjusted p-value. Analyses were performed in R using GSEA.