Exploring the role of circular RNAs in alveolar epithelial damage in pulmonary fibrosis

Introduction Pulmonary fibrosis is a rare disease characterised by scarring of the lung where lung transplant remains the only effective cure. Type II alveolar epithelial (AT2) cells play a key role in the fibrotic process. Several cell systems have been proposed as human models of AT2 cells, including AT2 cells generated from patient-derived induced pluripotent stem cells (iAT2) and the SALI model. Non-coding RNAs (ncRNAs), namely microRNAs (miRNAs) and circular RNAs (circRNAs) are thought to be involved in disease pathogenesis but little is known about their precise role in AT2 injury. Aim Here we aim to assess the expression and function of ncRNAs in AT2 injury using the iAT2 and the SALI model with the ultimate goal to explore their therapeutic potential in pulmonary fibrosis. Methods MiRNA and circRNA expression was measured in iAT2s carrying the SFPTC mutation vs CRISPR-Cas9 corrected clone using Arraystar Human Small RNA and CircRNA Microarray, respectively. Differentially expressed ncRNAs were identified using Volcano Plot filtering. Bioinformatic analysis was carried out to identify a list of candidate circRNAs. Results A total of 2560 miRNAs and 13548 circRNAs were detectable in the iAT2 model. 131 miRNAs and 905 circRNAs were significantly upregulated in SFPTC-mutant compared to corrected iAT2s, while 97 and 684 were downregulated, respectively. KEGG pathway enrichment highlighted two important pathways for pulmonary fibrosis among the top 10 enriched pathways. Seventeen circRNAs were selected for validation in qRT-PCR and further functional analysis. Conclusions NcRNAs are heavily dysregulated in SFPTC-mutant vs corrected iAT2s. Expression of the 17 candidate circRNAs will be validated in iAT2 and SALI models in presence or absence of pro-fibrotic cocktail