118 - The R213G Polymorphism in SOD3 Protects Early Bleomycin-Induced Pulmonary Inflammation and Attenuates Induction of Genes Involved in Agranulocyte/granulocyte Adhesion and Diapedesis

Abstract

Extracellular superoxide dismutase (SOD3), a key lung antioxidant enzyme, protects against lung and vascular inflammation. A single nucleotide polymorphism (SNP) substituting arginine to glycine at the 213 amino acid position (R213G) in SOD3 decreases tissue binding, thereby increasing SOD3 in the extracellular fluids such as epithelial lining fluid and plasma. We tested the hypothesis that the R213G SNP modulates key inflammatory signaling pathways in bleomycin induced lung injury to protect against progression to lung fibrosis and pulmonary hypertension (PH). In wild type (WT) mice, bleomycin increased multiple proinflammatory cytokines, measured with V-plex (Meso Scale Discovery) over 21 days. In R213G mice, inflammation increased 3 days post-bleomycin, however resolved by day 21 (n=6-8). Unlike the WT mice, R213G mice did not progress to lung fibrosis or PH, shown by physiologic, morphometric and biochemical measures. We performed high throughput RNAseq and bioinformatics analyses in lung RNA to evaluate gene pathways 7 days post-bleomycin in WT and R213G mice. The top enriched pathways, determined by ingenuity pathway analysis, were agranulocyte and granulocyte adhesion and diapedesis (2.51E-11 and 5.75E-06; p-value, respectively). These innate immune responses have important roles in the recruitment, adhesion and extravasation of leukocytes. Genes involved in adhesion (ITGAM and ITGB2), superoxide generation via NADPH oxidase (CYBB and NCF), and hydrolyzation of extracellular matrix (MMP12 and TIMP1) were preferentially upregulated in the WT mice compared to R213G mice. These findings were validated by qPCR. Bioinformatic analysis identifies potential pathways responsible for the protective effects of the R213G SNP and provides a foundation to better understand the role of redox regulation in inflammatory lung and pulmonary vascular diseases.