Effects of ω‐3 Polyunsaturated Fatty Acids and Aspirin on Expression of Arginase 2, a Protein Implicated in Airway Remodeling in Human Lung Fibroblasts

The hallmarks of airway remodeling (AR) in asthma are increased airway smooth muscle mass, subepithelial fibrosis, and increased amounts of fibroblasts and myofibroblasts. Once established, AR becomes resistant to current asthma therapies, possibly leading to irreversible airflow limitation and increasing asthma severity. Dysregulation of the arginine metabolism pathway has been implicated in AR. We hypothesized that activity of arginase 2 (ARG2), a key regulatory enzyme in the arginine metabolism pathway, is altered in AR, contributing to fibrosis. We investigated ARG2 levels in human lung fibroblasts (HLF) after exposure to various compounds, including ω‐3 polyunsaturated fatty acids and acetylsalicylic acid (ASA), that are thought to influence asthma development. Fibroblasts from normal (NHLF) and diseased (DHLF) asthmatic donors were treated with 10 μM, 50 μM, or 100 μM eicosapentaenoic acid (EPA) or 10 μM docosahexaenoic acid (DHA) for 24 h. ARG2 mRNA levels were detected by reverse‐transcriptase quantitative polymerase chain reaction (RT‐qPCR). ARG2 mRNA levels were measured after 24 h treatment with 1 μM RvD1 or a combination of 1 μM RvD1 and 10 mM ASA or 25 ng/mL T H 1 cytokine cocktail. HLF were treated with 1 nM, 10 nM, 50 nM, or 100 nM RvE1 for 24 h. HLF were also treated with 100 μM 8‐bromo‐cAMP (8‐Br‐cAMP), 2 μg/mL dexamethasone (DEX) or a combination for 24 h. HLF were treated with 10 mM ASA for 24 h, then ARG2 protein levels were identified via enzyme‐linked immunosorbent assay (ELISA) and cell immunochemistry. ELISA was performed with nuclear, mitochondrial, and cytosolic protein fractions. HLF were treated with 10 mM ASA for 24 h, and then stained for ARG2. In NHLF, changes in ARG2 mRNA levels showed no real pattern, while mRNA levels decreased in DHLF after treatment with 50 μM or 100 μM EPA. Exposure to 1 μM RvD1 did not affect ARG2 mRNA levels in HLF, but 25 ng/mL T H 1 cytokine cocktail or a combination of 1 μM RvD1 and 25 ng/mL T H 1 cytokine cocktail produced a significant (P<0.01) decrease in ARG2 mRNA in both NHLF and DHLF. Treatment with 1–100 nM RvE1 did not significantly affect ARG2 mRNA levels in HLF. 10 mM ASA or a combination of 10 mM ASA and 1 μM RvD1 produced significant (P<0.001) increases in ARG2 mRNA levels in both NHLF and DHLF. cAMP treatment decreased ARG2 mRNA levels in DHLF, while DEX did not produce a significant change. Increased levels of ARG2 protein were observed in both NHLF and DHLF mitochondrial fractions after 24 h of ASA treatment compared to untreated cells, with the DHLF mitochondrial fractions showing a significant (P<0.01) increase in ARG2. Cell immunochemistry experiments indicated an increase in ARG2 in DHLF treated with 10 mM ASA. A decrease in ARG2 mRNA was observed in DHLF with higher concentrations (50 μM and 100 μM) of EPA, the T H 1 cytokine cocktail, and with addition of 8‐Br‐cAMP, which may make these compounds useful as potential therapies in ARG2‐influenced AR. ASA produced an increase in ARG2 in HLF which may make it useful for studying the development of AR. Further study is needed to see if any of these compounds have long‐term effects on HLF ARG2 expression and activity, and on fibrosis in AR. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .