Azithromycin and its non-antibacterial derivative EP317 attenuate select aspects of ventilator-induced lung injury in rats

Ventilator-induced lung injury (VILI) is a significant complication of mechanical ventilation, contributing to morbidity and mortality in critically ill patients. Excessive mechanical stress triggers an inflammatory cascade, leading to lung damage, pulmonary oedema, and systemic complications. While macrolide antibiotics such as azithromycin (AZM) have demonstrated anti-inflammatory and epithelial barrier strengthening effects, their use is limited by concerns regarding antibiotic resistance. EP317, an azithromycin-derived compound with negligible antibacterial activity, was designed to retain the immunomodulatory and epithelial barrier enhancing effects of AZM while mitigating the risk of bacterial resistance. This study investigated the effects of AZM and EP317 in a rat model of VILI. Forty-eight rats were divided into six groups (N = 8 per group), receiving either placebo, AZM, or EP317 (5 mg/kg daily for seven days), with half of the animals subjected to mechanical ventilation with peak airway pressure at 35 cmH 2 O. Both compounds attenuated ventilation-induced pulmonary oedema, reflected in a smaller increase in lung wet/dry ratio, whereas ventilation-induced increases in total protein in bronchoalveolar lavage fluid (BALF) were not significantly reduced by either treatment. Cytokine profiling in plasma (L-Selectin, PAI-1, RANTES, MCP-3, IL-6, SCF, IL-17a, RAGE, MIP-3α, IL-13, G-CSF, IL-10, M-CSF, IFN-γ, GRO-KC and MMP9) revealed that both treatments attenuated inflammatory responses, while cytokine levels in BALF in ventilated animals were either unchanged or increased. Survival rates were improved in the treatment groups. Histological analysis indicated increased oedema in ventilated animals, likely due to the high-pressure ventilation, while drug-treated groups exhibited reduced oedema. Global gene expression profiling by RNA sequencing revealed robust upregulation of inflammatory and stress-response pathways in ventilated animals, with partial attenuation following AZM or EP317 pre-treatment. These findings suggest that EP317 and AZM offer protective effects against VILI. EP317 presents a promising alternative to conventional macrolide therapy by circumventing antibiotic resistance concerns.