Bronchopulmonary dysplasia (BPD) remains one of the main diseases in extremely premature infants. To prevent ventilator-induced lung injury and respiratory distress syndrome (RDS), high-frequency oscillatory ventilation (HFOV) with volume guarantee (VG) especially with high frequencies, has been previously proposed as an alternative method to invasive conventional mechanical ventilation (CMV). Upon HFOV-VG, improvement in several histopathological markers has been observed, however, no ex vivo studies have been performed to assess the impact of HFOV-VG on the function and composition of lung surfactant (LS). We have characterized the surface-active properties of LS from bronchoalveolar lavages (BAL) of piglets subjected to either HFOV-VG (12 animals) or CMV with low tidal volume (Vt) (6 animals) after lung-induced injury. Adsorption of LS at the air-liquid interface was assessed by Surfactant Adsorption Test (SAT), whereas the level of surfactant protein A (SP-A) was measured by Western Blot. A significantly higher LS adsorption/accumulation at the interface was observed in the HFOV-VG compared to the CMV group after 30 minutes of adsorption [16548,72(9701,10) vs 4690,94(3081,52)RFU, p=0.010]. Within the HFOV-VG group, no significant differences were found among animals ventilated at low (10 Hz) or high (20 Hz) frequencies. Moreover, there were no significant differences between the two groups in the amount of SP-A [32,18(20,90) vs 79,31(84,80)%ADU p=0,229]. This indicates that HFOV-VG could be a promising strategy for improving respiratory outcomes and highlights the importance of surfactant monitoring in the treatment of premature infants.