Clostridium butyricum MIYAIRI 588-Induced Protectin D1 Has an Anti-inflammatory Effect on Antibiotic-Induced Intestinal Disorder.
ABSTRACT: Metabolites are thought as the end products in cellular regulatory processes and their levels show the strongest relationships with the phenotype. Previously, we showed that the administration of Clostridium butyricum MIYAIRI 588 (CBM 588) upregulated protectin D1, an anti-inflammatory lipid metabolite, in colon tissue under antibiotic therapy. However, how CBM 588 induces protectin D1 expression and whether the metabolite has anti-inflammatory effects on antibiotic-induced inflammation are unclear. Therefore, here, we evaluated the effect of CBM 588 on lipid metabolism and protectin D1 in gut protection from antibiotic-induced intestinal disorders. In the CBM 588 treatment group, expression levels of genes encoding lipid receptors related to the conversion of DHA to protectin D1, such as polyunsaturated fatty acid (PUFA) receptors, G-protein coupled receptor 120 (GPR120), and 15-lipoxygenase (LOX), were increased in colon tissue. CD4+ cells producing interleukin (IL)-4, the main component of T helper type 2 (Th2) cells that can activate 15-LOX, also increased in CBM 588-treated groups even after clindamycin co-administration. In addition, similar to CBM 588, exogenously administered protectin D1 reduced inflammatory cytokines, while IL-10 and TGF-?1, works as anti-inflammatory cytokines, were increased. Our data revealed that CBM 588 activated 15-LOX to enhance protectin D1 production by increasing IL-4-producing CD4+ cell population in the intestinal tract. Additionally, CBM 588-induced protectin D1 clearly upregulated IL-10-producing CD4+ cells to control antibiotic-induced gut inflammation. We provide new insights into CBM 588-mediated lipid metabolism induction for the treatment of gut inflammatory diseases.
Project description:Metabolites are thought as the end products in cellular regulatory processes and their levels show the strongest relationships with the phenotype. Previously, we showed that the administration of Clostridium butyricum MIYAIRI 588 (CBM 588) upregulated protectin D1, an anti-inflammatory lipid metabolite, in colon tissue under antibiotic therapy. However, how CBM 588 induces protectin D1 expression and whether the metabolite has anti-inflammatory effects on antibiotic-induced inflammation are unclear. Therefore, here, we evaluated the effect of CBM 588 on lipid metabolism and protectin D1 in gut protection from antibiotic-induced intestinal disorders. In the CBM 588 treatment group, expression levels of genes encoding lipid receptors related to the conversion of DHA to protectin D1, such as polyunsaturated fatty acid (PUFA) receptors, G-protein coupled receptor 120 (GPR120), and 15-lipoxygenase (LOX), were increased in colon tissue. CD4+ cells producing interleukin (IL)-4, the main component of T helper type 2 (Th2) cells that can activate 15-LOX, also increased in CBM 588-treated groups even after clindamycin co-administration. In addition, similar to CBM 588, exogenously administered protectin D1 reduced inflammatory cytokines, while IL-10 and TGF-β1, works as anti-inflammatory cytokines, were increased. Our data revealed that CBM 588 activated 15-LOX to enhance protectin D1 production by increasing IL-4-producing CD4+ cell population in the intestinal tract. Additionally, CBM 588-induced protectin D1 clearly upregulated IL-10-producing CD4+ cells to control antibiotic-induced gut inflammation. We provide new insights into CBM 588-mediated lipid metabolism induction for the treatment of gut inflammatory diseases.
Project description:Clostridium butyricum MIYAIRI 588 (CBM 588) is a probiotic bacterium that has previously been used to prevent antibiotic-associated diarrhea. However, the underlying mechanism by which CBM 588 protects the gut epithelial barrier remains unclear. Here, we show that CBM 588 increased the abundance of Bifidobacterium, Lactobacillus, and Lactococcus species in the gut microbiome and also enhanced the intestinal barrier function of mice with antibiotic-induced dysbiosis. Additionally, CBM 588 significantly promoted the expansion of IL-17A-producing ??T cells and IL-17A-producing CD4 cells in the colonic lamina propria (cLP), which was closely associated with changes in the intestinal microbial composition. Additionally, CBM 588 plays an important role in controlling antibiotic-induced gut inflammation through upregulation of anti-inflammatory lipid metabolites such as palmitoleic acid, 15d-prostaglandin J2, and protectin D1. This study reveals a previously unrecognized mechanism of CBM 588 and provides new insights into gut epithelial barrier protection with probiotics under conditions of antibiotic-induced dysbiosis.
Project description:A convergent stereoselective synthesis of the potent anti-inflammatory, proresolving and neuroprotective lipid mediator protectin D1 (2) has been achieved in 15% yield over eight steps. The key features were a stereocontrolled Evans-aldol reaction with Nagao's chiral auxiliary and a highly selective Lindlar reduction of internal alkyne 23, allowing the sensitive conjugated E,E,Z-triene to be introduced late in the preparation of 2. The UV and LC/MS-MS data of synthetic protectin D1 (2) matched those obtained from endogenously produced material.
Project description:In the present study, we investigated the effect of CBM 588 on lifespan and multiple-stress resistance using Caenorhabditis elegans as a model animal. When adult C. elegans were fed a standard diet of Escherichia coli OP50 or CBM 588, the lifespan of the animals fed CBM 588 was significantly longer than that of animals fed OP50. Moreover, the worms fed CBM 588 were more resistant to certain stressors, including infections with pathogenic bacteria, UV irradiation, and the metal stressor Cu2+. CBM 588 failed to extend the lifespan of the daf-2/IR, daf-16/FOXO and skn-1/Nrf2 mutants. Transcriptional profiling comparing CBM 588-fed and control-fed animals suggested that DAF-16-dependent class II genes were regulated by CBM 588. In conclusion, CBM 588 extends the lifespan of C. elegans probably through regulation of the insulin/IGF-1 signaling (IIS) pathway and the Nrf2 transcription factor, and CBM 588 improves resistance to several stressors in C. elegans. Overall design: Transcriptional profiling of eight-day-old worms that were fed OP50 or CBM 588 for five days, by deep sequencing, using Illumina HiSeq.
Project description:Protectin D1 is a specialized pro-resolving mediator with potent pro-resolving and anti-inflammatory effects in vivo in several human disease models. Herein the preparation of the first synthetic analog of protectin D1, named 22-F-PD1, is presented together with data from in vivo investigations. This analog showed potent pro-resolving and anti-inflammatory properties. These results inspired the preparation of the radiotracer 22-[18F]F-PD1-ME that was used in a positron emission tomography proof of concept study. Altogether, the findings presented contribute to new knowledge on the biomolecular properties of protectin D1 analogs. In addition, an improved formal synthesis of the metabolite 22-OH-PD1 is reported.
Project description:The IL-6/signal transducer and activator of transcription 3 (STAT3) pathway is a critical signaling pathway for colitis-associated colorectal cancer (CAC). Peroxisome proliferator-activated receptor (PPAR)-?, a lipid nuclear receptor, up-regulates IL-6. 15-Lipoxygenase-1 (15-LOX-1), which is crucial to production of lipid signaling mediators to terminate inflammation, down-regulates PPAR-?. 15-LOX-1 effects on IL-6/STAT3 signaling and CAC tumorigenesis have not been determined. We report that intestinally targeted transgenic 15-LOX-1 expression in mice inhibited azoxymethane- and dextran sodium sulfate-induced CAC, IL-6 expression, STAT3 phosphorylation, and IL-6/STAT3 downstream target (Notch3 and MUC1) expression. 15-LOX-1 down-regulation was associated with IL-6 up-regulation in human colon cancer mucosa. Reexpression of 15-LOX-1 in human colon cancer cells suppressed IL-6 mRNA expression, STAT3 phosphorylation, IL-6 promoter activity, and PPAR-? mRNA and protein expression. PPAR-? overexpression in colonic epithelial cells promoted CAC tumorigenesis in mice and increased IL-6 expression and STAT3 phosphorylation, whereas concomitant 15-LOX-1 expression in colonic epithelial cells (15-LOX-1-PPAR-?-Gut mice) suppressed these effects: the number of tumors per mouse (mean ± sem) was 4.22 ± 0.68 in wild-type littermates, 6.67 ± 0.83 in PPAR-?-Gut mice (P = 0.026), and 2.25 ± 0.25 in 15-LOX-1-PPAR-?-Gut mice (P = 0.0006). Identification of 15-LOX-1 suppression of PPAR-? to inhibit IL-6/STAT3 signaling-driven CAC tumorigenesis provides mechanistic insights that can be used to molecularly target CAC.
Project description:Persistent activation of the innate immune system greatly influences the risk for developing metabolic complications associated with obesity. In this study, we explored the therapeutic potential of the specialized proresolving mediator (SPM) resolvin D1 (RvD1) to actively promote the resolution of inflammation in human visceral adipose tissue from obese (Ob) patients. Using liquid chromatography-tandem mass spectrometry-based metabololipidomic analysis, we identified unbalanced production of SPMs (i.e., D- and E-series resolvins, protectin D1, maresin 1, and lipoxins) with respect to inflammatory lipid mediators (i.e., leukotriene B4 and PGs) in omental adipose tissue from Ob patients. In parallel, high-throughput transcriptomic analysis revealed a unique signature in this tissue that was characterized by overactivation of the IL-10 signaling pathway. Incubation of inflamed Ob visceral adipose tissues and human macrophages with RvD1 limited excessive activation of the IL-10 pathway by reducing phosphorylation of STAT proteins. Of interest, RvD1 blocked STAT-1 and its target inflammatory genes (i.e., CXCL9), as well as persistent STAT3 activation, without affecting the IL-10 anti-inflammatory response characterized by inhibition of IL-6, IL-1?, IL-8, and TNF-?. Furthermore, RvD1 promoted resolution by enhancing expression of the IL-10 target gene heme oxygenase-1 by mechanisms dependent on p38 MAPK activity. Together, our data show that RvD1 can tailor the quantitative and qualitative responses of human inflamed adipose tissue to IL-10 and provide a mechanistic basis for the immunoresolving actions of RvD1 in this tissue. These findings may have potential therapeutic implications in obesity-related insulin resistance and other metabolic complications.
Project description:Protectin D1 (PD1 (3)), a C22-dihydroxylated polyunsaturated fatty acid biosynthesized from all-Z-docosahexaenoic acid, belongs to the new family of endogenous mediators referred to as specialized pro-resolving lipid mediators. PD1 (3) is a natural product that displays potent anti-inflammatory properties together with pro-resolving actions including inhibition of polymorphonuclear leukocyte (PMN) infiltration and promotion of macrophage phagocytosis and efferocytosis. Given its potent endogenous actions, this compound has entered several clinical development programs. Little has been reported on the metabolism of PD1 (3). The synthesis and biological evaluations of the ?-22 monohydroxylated metabolite of PD1 (3), named herein 22-OH-PD1 (6), are presented. LC-MS/MS data of the free acid 6, obtained from hydrolysis of the synthetic methyl ester 7, matched data for the endogenously produced 22-OH-PD1 (6). Compound 6 exhibited potent pro-resolving actions by inhibiting PMN chemotaxis in vivo and in vitro comparable to its precursor PD1 (3) and decreased pro-inflammatory mediator levels in inflammatory exudates. The results reported herein provide new knowledge of the metabolism of the protectin class of specialized pro-resolving mediators.
Project description:The resolution of inflammation is an active process orchestrated by specialized proresolving lipid mediators (SPM) that limit the host response within the affected tissue; failure of effective resolution may lead to tissue injury. Because persistence of inflammatory signals is a main feature of chronic inflammatory conditions, including inflammatory bowel diseases (IBDs), herein we investigate expression and functions of SPM in intestinal inflammation. Targeted liquid chromatography-tandem mass spectrometry-based metabololipidomics was used to identify SPMs from n-3 polyunsaturated fatty acids in human IBD colon biopsies, quantifying a significant up-regulation of the resolvin and protectin pathway compared with normal gut tissue. Systemic treatment with protectin (PD)1n-3 DPA or resolvin (Rv)D5n-3 DPA protected against colitis and intestinal ischemia/reperfusion-induced inflammation in mice. Inhibition of 15-lipoxygenase activity reduced PD1n-3 DPA and augmented intestinal inflammation in experimental colitis. Intravital microscopy of mouse mesenteric venules demonstrated that PD1n-3 DPA and RvD5n-3 DPA decreased the extent of leukocyte adhesion and emigration following ischemia-reperfusion. These data were translated by assessing human neutrophil-endothelial interactions under flow: PD1n-3 DPA and RvD5n-3 DPA reduced cell adhesion onto TNF-?-activated human endothelial monolayers. In conclusion, we propose that innovative therapies based on n-3 DPA-derived mediators could be developed to enable antiinflammatory and tissue protective effects in inflammatory pathologies of the gut.
Project description:Critical conditions such as sepsis following infection or traumatic injury disturb the complex state of homeostasis that may lead to uncontrolled inflammation resulting in organ failure, shock and death. They are associated with endogenous mediators that control the onset of acute inflammatory response, but the central problem remains the complete resolution of inflammation. Omega-3 enriched lipid emulsions (?-3+ LEs) were used in experimental studies and clinical trials to establish homeostasis, yet with little understanding about their role on the resolution of inflammation and tissue regeneration. Here, we demonstrate that ?-3 lipid emulsions (LEs) orchestrate inflammation-resolution/regeneration mechanism during sterile peritonitis and murine polymicrobial sepsis. ?-3+ LEs recessed neutrophil infiltration, reduced pro-inflammatory mediators, reduced the classical monocyte and enhanced the non-classical monocytes/macrophages recruitment and finally increased the efferocytosis in sepsis. The actions of ?-3+ LE were 5-lipoxygenase (5-LOX) and 12/15-lipoxygenase (12/15-LOX) dependent. ?-3+ LEs shortened the resolution interval by 56%, stimulated the endogenous biosynthesis of resolution mediators lipoxin A4, protectin DX and maresin 1 and contributed to tissue regeneration. ?-3+ LEs protected against hypothermia and weight loss and enhanced survival in murine polymicrobial sepsis. We highlighted a role of ?-3+ LEs in regulating key mechanisms within the resolution terrain during murine sepsis. This might form the basis for a rational design of sepsis specific clinical nutrition.