Investigation of the contribution of non-canonical inflammasome activation to LPS-induced foam cell formation in macrophages

<p dir="ltr">Bacterial lipopolysaccharide (LPS) has been shown to promote the accumulation of cytosolic lipid droplets in macrophages, converting them into so-called ‘foam cells’. This process may contribute to atherosclerosis, but it is not clear how LPS signalling connects to lipid metabolism in macrophages. In this project, the hypothesis that detection of LPS via the non-classical caspase-11 inflammasome may contribute to macrophage foam cell formation was investigated using small molecule inhibitors, gene knockout and overexpression experiments.</p><p dir="ltr">Supportive of the hypothesis, foam cell formation was increased by transfection of LPS into the cytosol of macrophages, in comparison to exposure without transfection. LPS was also found to both prime and activate caspase-11 at later timepoints (>48 h) without requirement for transfection, suggesting that it can enter the cell by natural means. Two different inhibitors of caspase-11, wedelactone and scutellarin, significantly blunted lipid accumulation in response to LPS treatment. LPS-induced lipid accumulation was lower, but not completely absent, in a Tlr4-/- macrophage cell-line in comparison to wild-type control cells. Further investigation using Asc-/- macrophages showed that there is no requirement for the classical inflammasome for this process to occur.</p><p dir="ltr">Bioinformatics analysis revealed potential caspase-4 cleavage sites in the lipid regulatory proteins sterol regulatory element binding protein (SREBP)1a, SREBP1c, and SREBP2. Overexpression of human caspase-4 in HEK-293 cells by transfection successfully replicated the gasdermin-D cleavage and pyroptosis responses of non-classical inflammasomes in response to LPS. However, as a FLAG tagged SREBP construct did not express well in this system, it remains unclear if SREBPs may be cleaved by caspase-4 activation.</p><p dir="ltr">The present findings suggest that caspase-11 signalling contributes to LPS-induced lipid accumulation in macrophages cultured in vitro. Further experiments using in vivo studies of caspase-11 knockout mice will be necessary to test this hypothesis further.</p>