Acute lung injury model in human Precision Cut Lung Slices.

Background: Repetitive alveolar injury contributes to fibrotic interstitial lung disease (ILD) development such as idiopathic pulmonary fibrosis. Alveolar injury can have various causes. Evidence in the literature shows strong evidence of cytotoxicity, pro-inflammatory, and pro-oxidative properties of heme (Soares MP,Curr Opin Immunol.2016)and its association with acute respiratory distress syndrome (ARDS),the clinical presentation of major acute lung injury. Reliable pre-clinical models to study acute lung injury (ALI) are sparse. Therefore, we have developed an advanced exvivo model of heme-induced acute lung injury using precision cut lung slices (PCLS). We used human PCLS to investigate heme-induced acute injury. Methods: PCLS were obtained from 12 human donors undergoing lung resection surgery. PCLS were cultured and stimulated with 100µM heme for 24h. Viability was assessed by XTT assay at different concentrations of Heme. Inflammation was determined by cytokine secretion (ELISA). The PCLS were collected and bulk RNA-sequencing was performed. Results: Heme-treated PCLS had reduced viability at 100µM and above compared to untreated controls. Pro-inflammatory and injury signals were induced after heme stimulation. The transcriptomic analysis showed upregulation of a total of 732 genes, including mostly inflammatory genes like IL-6,CXCL8,and IL-1β after heme stimulation. Inflammatory cytokines such as IL-6,TNF-α,and IL-8 were significantly elevated in supernatants from PCLS treated with heme compared to untreated PCLS. Conclusion: Heme induces inflammation in human PCLS and is cytotoxic at higher levels as observed during ALI. Future projects will investigate the role of heme-induced lung injury in fibrosis development in our new model.