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Autotaxin (ATX) converts lysophosphatidylcholine and sphingosyl-phosphorylcholine into lysophosphatidic acid and sphingosine 1-phosphate, respectively. Despite the pivotal function of ATX in lipid metabolism, mechanisms by which ATX regulates immune and inflammatory disorders remain elusive. Here, using myeloid cell lineage-restricted Atx knockout mice, we show that Atx deficiency disrupts membrane microdomains and lipid rafts, resulting in the inhibition of Toll-like receptor 4 (TLR4) complex formation and the suppression of adaptor recruitment, thereby inhibiting TLR4-mediated responses in macrophages. Accordingly, TLR4-induced innate immune functions, including phagocytosis and iNOS expression, are attenuated in Atx-deficient macrophages. Consequently, Atx-/- mice exhibit a higher bacterial prevalence in the intestinal mucosa compared to controls. When combined with global Il10-/- mice, which show spontaneous colitis due to the translocation of luminal commensal microbes into the mucosa, myeloid cell lineage-restricted Atx knockout accelerates colitis development compared to control littermates. Collectively, our data reveal that Atx deficiency compromises innate immune responses, thereby promoting microbe-associated gut inflammation.

BackgroundPhosphatase and tensin homolog (Pten) is capable of mediating microbe-induced immune responses in the gut. Thus, Pten deficiency in the intestine accelerates colitis development in Il10-/- mice. As some ambient pollutants inhibit Pten function and exposure to ambient pollutants may increase inflammatory bowel disease (IBD) incidence, it is of interest to examine how Pten inhibition could affect colitis development in genetically susceptible hosts.MethodsWith human colonic mucosa biopsies from pediatric ulcerative colitis and non-IBD control subjects, we assessed the mRNA levels of the PTEN gene and the gene involved in IL10 responses. The data from the human tissues were corroborated by treating Il10-/-, Il10rb-/-, and wild-type C57BL/6 mice with Pten-specific inhibitor VO-OHpic. We evaluated the severity of mouse colitis by investigating the tissue histology and cytokine production. The gut microbiome was investigated by analyzing the 16S ribosomal RNA gene sequence with mouse fecal samples.ResultsPTEN and IL10RB mRNA levels were reduced in the human colonic mucosa of pediatric ulcerative colitis compared with non-IBD subjects. Intracolonic treatment of the Pten inhibitor induced colitis in Il10-/- mice, characterized by reduced body weight, marked colonic damage, and increased production of inflammatory cytokines, whereas Il10rb-/- and wild-type C57BL/6 mice treated with the inhibitor did not develop colitis. Pten inhibitor treatment changed the fecal microbiome, with increased abundance of colitogenic bacteria Bacteroides and Akkermansia in Il10-/- mice.ConclusionsLoss of Pten function increases the levels of colitogenic bacteria in the gut, thereby inducing deleterious colitis in an Il10-deficient condition.

Intestinal wound healing is a new therapeutic goal for inflammatory bowel disease (IBD) as complete healing of the mucosa is the key element of clinical remission in IBD. Previous studies showed that termination of inflammation can be achieved by adding pro-resolving lipids like DHA and EPA exogenously. However, the roles of these lipids in mucosal healing have not been investigated. To recapitulate intestinal healing process, mice were received dextran sodium sulfate (DSS) for 7 days in the drinking water followed by regular tap water for 5 additional days. DSS-induced intestinal inflammation featuring body weight loss, histological tissue damage, increased cytokine production and infiltration of inflammatory cells was gradually reduced upon switching to water. To investigate whether endogenous lipids play a role in mucosal healing, the lipidomics analysis of mouse serum was performed. Reduced levels of arachidonic acid, the biosynthetic precursor of prostaglandin F (PGF)2α, 19H-PGF1α, the metabolite of prostacyclin, and 20H-PGF2α, the metabolite of PGF2α, suggest subsiding inflammation. In contrast, increased levels of an active metabolite of resolvin D1 along with decreased levels of its precursor DHA as well as decreased levels of the precursor of resolvin E, 18-hydroxy-eicosapentaenoic acid, suggest inauguration of mucosal healing by endogenous lipids. Furthermore, exogenously supplied fish oil enhanced the process even further. These results suggest the presence of mucosal healing regulated by endogenous pro-healing lipids and also indicate that the remission state of IBD could be prolonged by enhancing the levels of these lipids.

Inflammatory bowel disease (IBD) is a chronic inflammatory disease with increasing incidence and prevalence worldwide. Here we investigated the newly synthesized jasmonate analogue 2-hydroxyethyl 5-chloro-4,5-didehydrojasmonate (J11-Cl) for its anti-inflammatory effects on intestinal inflammation. First, to test whether J11-Cl can activate peroxisome proliferator-activated receptors (PPARs), we performed docking simulations because J11-Cl has a structural similarity with anti-inflammatory 15-deoxy-Δ(12,14)-prostaglandin J2 (15d-PGJ2), one of the endogenous ligands of PPARγ. J11-Cl bound to the ligand binding domain of PPARγ in the same manner as 15d-PGJ2 and rosiglitazone, and significantly increased transcriptional activity of PPARγ. In animal experiments, colitis was significantly reduced in mice with J11-Cl treatment, determined by analyses of survival rate, body weight changes, clinical symptoms, and histological evaluation. Moreover, J11-Cl decreased production of pro-inflammatory cytokines including IL-6, IL-8, and G-CSF as well as chemokines including chemokine (C-C motif) ligand (CCL)20, chemokine (C-X-C motif) ligand (CXCL)2, CXCL3, and chemokine (C-X3-C motif) ligand 1 (CX3CL1) in colon tissues, and LPS or TNF-α-stimulated macrophages and epithelial cells. In contrast, production of anti-inflammatory cytokines including IL-2 and IL-4 as well as the proliferative factor, GM-CSF, was increased by J11-Cl. Furthermore, inhibition of MAPKs and NF-κB activation by J11-Cl was also observed. J11-Cl reduced intestinal inflammation by increasing the transcriptional activity of PPARγ and modulating inflammatory signaling pathways. Therefore, our study suggests that J11-Cl may serve as a novel therapeutic agent against IBD.

Colonic epithelium is the first line of defense against various pathological offenses in the gut. Previous studies have shown that the peptides of the corticotropin-releasing hormone (CRH) family modulate vascular endothelial growth factor (VEGF)-A production in other cells. Here we sought to investigate whether CRH and urocortin (Ucn) 3 regulate VEGF-A secretion in colonocytes through CRH receptors and to elucidate the underlying mechanism of action. CRH and Ucn 3 significantly increased the expression levels of VEGF-A mRNA and protein through CRH receptor 1 and 2, respectively, in human colonic epithelial cells and primary mouse intestinal epithelial cells. Underlying mechanisms involve activation of adenylyl cyclase with subsequent increase of intracellular cAMP level and increased DNA binding activity of transcription factor CREB on VEGF-A promoter region. Finally, genetic deficiency of CREB decreased intestinal inflammation and VEGF-A expression in a dextran sodium sulfate-induced colitis model. These results show that activation of CRH receptors by CRH ligands stimulates VEGF-A expression in intestinal epithelial cells through the cAMP/CREB pathway. Since VEGF-A boosts inflammatory responses through angiogenesis, these data suggest that CREB may be a key effector of CRH and Ucn 3-dependent inflammatory angiogenesis.

Background & aimsEarly-onset ulcerative colitis, which is considered severe colonic inflammation that develops in infants and young children, can be caused by alterations in interleukin (IL)-10 signaling, although other factors are involved in its pathogenesis. We investigated whether loss of phosphatase and tensin homologue (PTEN), which regulates many important cell functions such as cell proliferation, cell survival, and Toll-like receptor (TLR) signaling pathways, contributes to the development of colitis in Il10(-/-) mice.MethodsWe generated Il10(-/-) mice (in C57BL/6 and C3H/HeJBir background strains) with disruption of Pten in the intestinal epithelium (Ints(ΔPten/ΔPten);Il10(-/-) mice) and Ints(ΔCont);Il10(-/-) (control) mice. Colon tissues were collected and histological, transmission electron microscopy, and gene expression analysis were performed. Fecal microbiota samples were analyzed by sequencing of 16S ribosomal RNA genes. We disrupted Tlr4 in Ints(ΔPten/ΔPten);Il10(-/-) mice. Lipopolysaccharide signaling via TLR4 was blocked by treating mice with polymyxin B.ResultsIl10(-/-) mice developed colitis when they were 6 to 7 months old, whereas Ints(ΔPten/ΔPten);Il10(-/-) mice developed severe colitis and colon tumors by the time they were 36 days old. Within 3 months of birth, 80% of Ints(ΔPten/ΔPten);Il10(-/-) mice developed severe colitis and colonic malignancy, whereas none of the Ints(ΔCont);Il10(-/-) mice had these phenotypes. Ints(ΔPten/ΔPten);Il10(-/-) mice had alterations in fecal microbiota compared with controls, such as increased proportions of Bacteroides species, which are gram negative. Disruption of Tlr4 or treating Ints(ΔPten/ΔPten);Il10(-/-) mice with polymyxin B delayed the development of colitis and reduced disease severity.ConclusionsDisruption of Pten in the intestinal epithelium of Il10(-/-) mice speeds the onset and increases the severity of colitis. Fecal microbiota from Ints(ΔPten/ΔPten);Il10(-/-) mice have increased proportions of Bacteroides species. Development of colitis is delayed and reduced by blocking TLR4 signaling. Ints(ΔPten/ΔPten);Il10(-/-) mice may be studied as a model for early-onset ulcerative colitis and used to identify new therapeutic targets.

Background & aimsEarly-onset ulcerative colitis, which is considered severe colonic inflammation that develops in infants and young children, can be caused by alterations in interleukin (IL)-10 signaling, although other factors are involved in its pathogenesis. We investigated whether loss of phosphatase and tensin homologue (PTEN), which regulates many important cell functions such as cell proliferation, cell survival, and Toll-like receptor (TLR) signaling pathways, contributes to the development of colitis in Il10(-/-) mice.MethodsWe generated Il10(-/-) mice (in C57BL/6 and C3H/HeJBir background strains) with disruption of Pten in the intestinal epithelium (Ints(ΔPten/ΔPten);Il10(-/-) mice) and Ints(ΔCont);Il10(-/-) (control) mice. Colon tissues were collected and histological, transmission electron microscopy, and gene expression analysis were performed. Fecal microbiota samples were analyzed by sequencing of 16S ribosomal RNA genes. We disrupted Tlr4 in Ints(ΔPten/ΔPten);Il10(-/-) mice. Lipopolysaccharide signaling via TLR4 was blocked by treating mice with polymyxin B.ResultsIl10(-/-) mice developed colitis when they were 6 to 7 months old, whereas Ints(ΔPten/ΔPten);Il10(-/-) mice developed severe colitis and colon tumors by the time they were 36 days old. Within 3 months of birth, 80% of Ints(ΔPten/ΔPten);Il10(-/-) mice developed severe colitis and colonic malignancy, whereas none of the Ints(ΔCont);Il10(-/-) mice had these phenotypes. Ints(ΔPten/ΔPten);Il10(-/-) mice had alterations in fecal microbiota compared with controls, such as increased proportions of Bacteroides species, which are gram negative. Disruption of Tlr4 or treating Ints(ΔPten/ΔPten);Il10(-/-) mice with polymyxin B delayed the development of colitis and reduced disease severity.ConclusionsDisruption of Pten in the intestinal epithelium of Il10(-/-) mice speeds the onset and increases the severity of colitis. Fecal microbiota from Ints(ΔPten/ΔPten);Il10(-/-) mice have increased proportions of Bacteroides species. Development of colitis is delayed and reduced by blocking TLR4 signaling. Ints(ΔPten/ΔPten);Il10(-/-) mice may be studied as a model for early-onset ulcerative colitis and used to identify new therapeutic targets.

N‐Acetylserotonin (NAS) is an intermediate in the melatonin biosynthetic pathway. We investigated the anti‐inflammatory activity of NAS by focusing on its chemical feature oxidizable to an electrophile. NAS was readily oxidized by reaction with HOCl, an oxidant produced in the inflammatory state. HOCl‐reacted NAS (Oxi‐NAS), but not NAS, activated the anti‐inflammatory nuclear factor erythroid 2‐related factor 2 (Nrf2)‐heme oxygenase (HO)‐1 pathway in cells. Chromatographic and mass analyses demonstrated that Oxi‐NAS was the iminoquinone form of NAS and could react with N‐acetylcysteine possessing a nucleophilic thiol to form a covalent adduct. Oxi‐NAS bound to Kelch‐like ECH‐associated protein 1, resulting in Nrf2 dissociation. Moreover, rectally administered NAS increased the levels of nuclear Nrf2 and HO‐1 proteins in the inflamed colon of rats. Simultaneously, NAS was converted to Oxi‐NAS in the inflamed colon. Rectal NAS mitigated colonic damage and inflammation. The anticolitic effects were significantly compromised by the coadministration of an HO‐1 inhibitor. [ABSTRACT FROM AUTHOR]

Angiogenesis is required for wound healing and repair, but dysregulated angiogenesis is involved in gastrointestinal inflammation. Bacillus polyfermenticus (B.P.) is a probiotic bacterium clinically used for a variety of intestinal disorders in East Asia. Here we investigated the effect of B.P. on angiogenesis of human intestinal microvascular endothelial cells (HIMECs) and wound healing in intestinal mucosa. Exposure of HIMECs to the conditioned medium of B.P. cultures (B.P. CM) increased cell migration, permeability, and tube formation. Production of the proangiogenic cytokine IL-8 was increased by B.P. CM, and neutralizing antibodies against IL-8 or IL-8 receptor CXCR2 reduced tube formation as well as actin stress fiber formation. B.P. CM also increased NF-[kappa]B activation, and inhibitors of NF-[kappa]B suppressed B.P. CM-induced tube formation and IL-8 production. Furthermore, B.P. facilitated recovery of mice from colitis as shown by increased body weight and reduced rectal bleeding and histological severity. B.P. also increased angiogenesis and mouse IL-8 production in the mucosal layer. Collectively, these results show that B.P. increases angiogenesis of HIMECs in a NF-[kappa]B/IL-8/CXCR2-dependent manner. Moreover, B.P. promotes angiogenesis in the mucosa during recovery of mice from colitis, suggesting that this probiotic may be clinically used to facilitate intestinal wound healing. inflammatory bowel diseases; wound healing; CXCR doi: 10.1152/ajpgi.00204.2009.

Although human consumption of Bacillus polyfermenticus provides several health benefits, the probiotic effect of this bacterium against colonic inflammation has not yet, to our knowledge, been studied. Therefore, we induced colitis in mice by oral or intrarectal administration of dextran sodium sulfate (DSS) or trinitrobenzenosulfonic acid (TNBS), respectively, and investigated the effect of B. polyfermenticus on colitis. We found that mice treated with DSS or TNBS along with B. polyfermenticus had reduced mortality and severity of colitis (weight loss, diarrhea, and mucosal damages) than mice treated with DSS or TNBS alone. B. polyfermenticus also reduced the expression of inflammatory molecules, including chemokine (C-X-C motif) ligand 1, intercellular adhesion molecule, and tumor necrosis factor-u, but enhanced the expression of the antiinflammatory cytokine interleukin-10 in the inflamed mouse colon. Moreover, B. polyfermenticus suppressed apoptosis both in vivo in inflamed colonic mucosa and in vitro in colonic epithelial cells stimulated with apoptosis-inducing agents (FasL or Clostridium difficile Toxin A) when the apoptotic response was determined by a terminal deoxynucleotidyl transferase dUTP nick end labeling assay and cleavage of poly(ADP-ribose) polymerase or caspase-3, respectively. Treating colonic epithelial cells with B. polyfermenticus-conditioned medium (BPCM) enhanced cell proliferation and induced the phosphoinositide 3-kinases/Akt signaling pathway, suggesting that this bacterium can promote epithelial cell proliferation. BPCM also promoted the migration of colonic epithelial cells. These data suggest that B. polyfermenticus ameliorates colonic inflammation by suppressing apoptosis and promoting epithelial cell proliferation and migration.