Abstract 2073 KNOCKDOWN OF CES1 IN THP-1 MACROPHAGES PROMOTES A PROINFLAMMATORY PHENOTYPE

Macrophages are innate immune cells with the capacity to differentiate into classically activated (M1) or alternatively activated (M2) cells that promote or attenuate inflammation, respectively. Inflammation caused in response to pathogen- or damage-associated molecular patterns is normally resolved during healthy physiology. However, dysregulated resolution can lead to chronic inflammation, contributing to disease development. Carboxylesterase 1 (CES1) is a serine hydrolase that can metabolize neutral lipids. Our previous results indicated that THP-1 macrophages with deficient CES1 expression (CES1KD cells) have a distinctly foamy phenotype compared to CES1-expressing THP-1 macrophages (control cells), which is due to greater quantities of TAG-containing lipid droplets. Further, CES1KD cells produce more IL-1b and PGE2 than control cells following lipopolysaccharide (LPS) stimulation. It is known that classically-activated macrophages exhibit a broken TCA cycle, causing a buildup of proinflammatory citrate and succinate metabolites. Here, we report that CES1KD cells have higher levels of several polar organic acid metabolites than control cells under both baseline and LPS-stimulated conditions. These included increased lactate (glycolytic waste product) and citrate, cis-aconitate, and a-ketoglutarate (TCA metabolites), although succinate was depressed. There was also a striking reduction in UDP-GlcNAc levels in CES1KD cells, which is a metabolite that affords O-GlcNAc PTMs on proteins during M2 macrophage polarization. In line with this, M2 gene markers ALOX15, FABP4, and CD206 in CES1KD cells were downregulated compared to those in control cells. Based on our RNA-seq data, we observed a high degree of similarity in the differentially expressed genes in baseline CES1KD cells with those of control cells treated with LPS/IFNg (r=0.728), which is concordant with the altered metabolite profiles of CES1KD cells and their more proinflammatory nature. Finally, Gene Ontology, KEGG, and Reactome database analyses of unstimulated CES1KD cells revealed an enrichment of upregulated pathways involved in antibacterial and antiviral defense that is comparable to that seen in control cells stimulated with LPS and IFN-γ. Thus, we conclude that CES1KD macrophages under baseline conditions (M0) appear to exhibit a immunophenotype that is similar to that of M1 control cells. These findings further suggest that CES1 may regulate the macrophage inflammatory immune response and enhancing CES1 activity might reverse, or attenuate diseases caused by chronic inflammation.