Identification and characterization of a novel anti-inflammatory lipid isolated from Mycobacterium vaccae, a soil-derived bacterium with immunoregulatory and stress resilience properties

Abstract

<div class="section"> <a class="named-anchor" id="S1"> <!-- named anchor --> </a> <h5 class="section-title" id="d10833758e298">Rationale</h5> <p id="P2"> <i>Mycobacterium vaccae</i> (NCTC 11659) is an environmental saprophytic bacterium with anti-inflammatory, immunoregulatory, and stress resilience properties. Previous studies have shown that whole, heat-killed preparations of <i>M. vaccae</i> prevent allergic airway inflammation in a murine model of allergic asthma. Recent studies also demonstrate that immunization with <i>M. vaccae</i> prevents stress-induced exaggeration of proinflammatory cytokine secretion from mesenteric lymph node cells stimulated ex vivo, prevents stress-induced exaggeration of chemically induced colitis in a model of inflammatory bowel disease, and prevents stress-induced anxiety-like defensive behavioral responses. Furthermore, immunization with <i>M. vaccae</i> induces anti-inflammatory responses in the brain and prevents stress-induced exaggeration of microglial priming. However, the molecular mechanisms underlying anti-inflammatory effects of <i>M. vaccae</i> are not known. </p> </div><div class="section"> <a class="named-anchor" id="S2"> <!-- named anchor --> </a> <h5 class="section-title" id="d10833758e318">Objectives</h5> <p id="P3">Our objective was to identify and characterize novel anti-inflammatory molecules from M. vaccae NCTC 11659. </p> </div><div class="section"> <a class="named-anchor" id="S3"> <!-- named anchor --> </a> <h5 class="section-title" id="d10833758e323">Methods</h5> <p id="P4">We have purified and identified a unique anti-inflammatory triglyceride, 1,2,3-tri [ <i>Z</i>-10-hexadecenoyl] glycerol, from <i>M. vaccae</i> and evaluated its effects in freshly isolated murine peritoneal macrophages. </p> </div><div class="section"> <a class="named-anchor" id="S4"> <!-- named anchor --> </a> <h5 class="section-title" id="d10833758e334">Results</h5> <p id="P5">The free fatty acid form of 1,2,3-tri [ <i>Z</i>-10-hexadecenoyl] glycerol, 10( <i>Z</i>)-hexadecenoic acid, decreased lipopolysaccharide-stimulated secretion of the proinflammatory cytokine IL-6 ex vivo. Meanwhile, next-generation RNA sequencing revealed that pretreatment with 10( <i>Z</i>)-hexadecenoic acid upregulated genes associated with peroxisome proliferator-activated receptor alpha (PPARα) signaling in lipopolysaccharide-stimulated macrophages, in association with a broad transcriptional repression of inflammatory markers. We confirmed using luciferase-based transfection assays that 10( <i>Z</i>)-hexadecenoic acid activated PPARα signaling, but not PPARγ, PPARδ, or retinoic acid receptor (RAR) α signaling. The effects of 10( <i>Z</i>)-hexadecenoic acid on lipopolysaccharide-stimulated secretion of IL-6 were prevented by PPARα antagonists and absent in PPARα-deficient mice. </p> </div><div class="section"> <a class="named-anchor" id="S5"> <!-- named anchor --> </a> <h5 class="section-title" id="d10833758e354">Conclusion</h5> <p id="P6">Future studies should evaluate the effects of 10( <i>Z</i>)-hexadecenoic acid on stress-induced exaggeration of peripheral inflammatory signaling, central neuroinflammatory signaling, and anxiety- and fear-related defensive behavioral responses. </p> </div>