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INVESTIGATING THE ROLE OF THE LIPID-IMMUNE SIGNALING AXIS IN SALMONELLA ENTERICA SEROVAR TYPHIMURIUM INFECTIONS
Taddeo, Jessica
Taddeo, Jessica
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2025-05
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Biomedical Sciences
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https://doi.org/10.34944/brpf-bh02
Abstract
Salmonella enterica serovar Typhimurium (STm) is a gram-negative facultative intracellular pathogen that poses a significant health risk due to its route of infection through fecal-oral contamination. This pathogen invades the intestinal epithelial cells causing gastroenteritis and diarrhea and can disseminate into other tissues through survival in phagocytes. Eicosanoids-derived bioactive lipids play critical roles in both the induction and resolution of inflammation resulting from an infection, helping to precisely regulate the inflammatory response to both clear the pathogen while also limiting sustained tissue damage. A subset of these lipids are endogenous ligands for nuclear receptors/transcription factor Peroxisome proliferator-activated receptor (PPAR), composed of PPARα, PPARβ/δ, and PPARγ subtypes. PPAR plays major roles in fatty acid metabolism, modulation of immune functions, and formation of subcellular lipid bodies (sites of eicosanoid metabolism). PPAR also modulates macrophage polarization in which microenvironment, cytokines, and bacterial products influence the functional status. Previous studies have shown that STm infects and persists in M2 (alternatively activated) macrophages and induces M2 polarization upon infection through non-canonical STAT3 activation and IL-4 production. In the present study, we aimed to elucidate the role of how fatty acids, eicosanoids, and PPARα modulate the inflammatory responses during STm infections. We infected streptomycin-pretreated C57BL/6 (wildtype) and Ppara–/– mice with STm (colitis model), determining that Ppara–/– mice have lower cecal inflammation, cecal inflammatory gene expression, and bacterial dissemination in liver and spleen tissue, as well as lower numbers of STm-infected M2 macrophages from the mesenteric lymph nodes, compared to wildtype mice at 48-hours post infection. We also determined that Ppara–/– mice have a more pronounced increase in eicosanoid metabolism and a more significant shift in fatty acid profile following infection compared to wildtype, suggesting a localized effect of STm infection on fatty acid and eicosanoid metabolism in the cecum, which may contribute to increased infection resistance.
To interrogate the mechanisms by which the lipid-PPARα axis affects macrophage polarization during STm infections, we polarized wildtype and Ppara–/– macrophages to naive or M2b polarization subtypes and then infected them with STm. We found that that STm infection increased eicosanoid production and lipid body production in wildtype macrophages, and these macrophages had higher bacterial internalization compared to Ppara–/– macrophages. We also measured levels of ceramide, a long-chain fatty acid that has been shown to promote cell death; we found that STm-infected wildtype M2b-polarized macrophage have lower ceramide levels compared to uninfected, suggesting that STm intracellular infection interferes with the ceramide-induced cell death pathway to promote intracellular survival. Lastly, we determined that inhibition of fatty acid oxidation via CPT-1 inhibitor (etomoxir) during STm infection increases cell death, decreases bacterial load, and increases inflammatory response (increased Cxcl1 expression), suggesting that inhibiting fatty acid oxidation (downstream of PPARα) may help wildtype macrophages fight STm infection more effectively. We also determined that STm infection in wildtype mice treated with Etomoxir increased ceramide and phosphatidylcholine production and inflammatory gene expression in the cecum, as well as increased infected M1 macrophages from mesenteric lymph nodes (MLNs), suggesting that inhibition of fatty acid oxidation in mice can provide a localized effect on antibacterial activities in the cecum, including more pro-inflammatory macrophage polarization and bacterial clearance.
From our studies on ceramide in STm-induced cell death, we also took an interest in fatty acid metabolism in pyroptosis, caspase-1 dependent cell death that is most often triggered by infection with intracellular pathogens. We first measured fatty acid profiles in wildtype and Ppara–/– macrophages stimulated with PAM3CSK4, a TLR2/1 agonist, or WY14643, an agonist of PPARα, finding that activation of PPARα or TLR2/1 increases fatty acid production in wildtype macrophages, while combination treatment did not, suggesting that TLR stimulation and PPARα activation modulate fatty acid metabolism in potentially antagonistic mechanisms that may serve to regulate each other. We then measured pyroptotic cell death, finding that Ppara–/– macrophages have higher pyroptotic cell death compared to wildtype macrophages. In addition, accumulations of ceramide, phosphatidylethanolamines, and lysophosphatidylethanolamines in pyroptotic cells lacking GasderminD highlight a role for upregulation of LCFA in induction of cell death and suggest a role dysregulated lipid metabolism and lipotoxicity in Ppara–/– macrophages that contributes to higher cell death.
Overall, this study indicates that PPARα exacerbates STm infection through the modulation of lipid-mediated immunometabolism and macrophage polarization. As well, this work highlights the importance of understanding how lipid metabolism contributes to orchestrating inflammatory activities, such as cell death, and provides new insights into discovering innovative targets for modulating host inflammation & immune responses to combat bacterial infections.
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