Annexin-1 and peptide derivatives are released by apoptotic cells and stimulate phagocytosis of apoptotic neutrophils by macrophages

The resolution of inflammation is a dynamically regulated process that may be subverted in many pathological conditions. Macrophage (Mphi) phagocytic clearance of apoptotic leukocytes plays an important role in the resolution of inflammation as this process prevents the exposure of tissues at the inflammatory site to the noxious contents of lytic cells. It is increasingly appreciated that endogenously produced mediators, such as lipoxins, act as potent regulators (nanomolar range) of the phagocytic clearance of apoptotic cells. In this study, we have investigated the intriguing possibility that apoptotic cells release signals that promote their clearance by phagocytes. We report that conditioned medium from apoptotic human polymorphonuclear neutrophils (PMN), Jurkat T lymphocytes, and human mesangial cells promote phagocytosis of apoptotic PMN by Mphi and THP-1 cells differentiated to a Mphi-like phenotype. This prophagocytic activity appears to be dose dependent, sensitive to the caspase inhibitor zVAD-fmk, and is associated with actin rearrangement and release of TGF-beta1, but not IL-8. The prophagocytic effect can be blocked by the formyl peptide receptor antagonist Boc2, suggesting that the prophagocytic factor(s) may interact with the lipoxin A(4) receptor, FPRL-1. Using nanoelectrospray liquid chromatography mass spectrometry and immunodepletion and immunoneutralization studies, we have ascertained that annexin-1 and peptide derivatives are putative prophagocytic factors released by apoptotic cells that promote phagocytosis of apoptotic PMN by M[phi] and differentiated THP-1 cells. These data highlight the role of annexin-1 and peptide derivatives in promoting the resolution of inflammation and expand on the therapeutic anti-inflammatory potential of annexin-1.     

Pentraxin-3 regulates the inflammatory activity of macrophages

Background and aimsPentraxin-3 (PTX3) reportedly has protective roles in atherosclerosis and myocardial infarction, and is a useful biomarker of vascular inflammation. However, the detailed functions of PTX3 in inflammation are yet to be elucidated. This study aimed to investigate the function of PTX3 in macrophages.MethodsPMA-treated THP-1 cell line (THP-1 macrophage) and monocyte-derived human primary macrophages were treated with recombinant PTX3. Cytokine and chemokine levels in the THP-1 culture medium were measured as well as monocyte chemoattractant protein (MCP-1) concentrations in the Raw 264.7 cell culture medium. PTX3-silenced apoptotic macrophages (THP-1 cell line) were generated to investigate the roles of PTX3 in phagocytosis.ResultsIn the presence of PTX3, macrophage interleukin-1β (IL-1β), tumor necrosis factor-alpha (TNF-α) and MCP-1 levels were reduced significantly (?39%, P=0.007; ?21%, P=0.008; and ?67%, P=0.0003, respectively), whilst activated transforming growth factor-β (TGF?β) was detected in the THP-1 macrophages (P=0.0004). Additionally, PTX3 induced Akt phosphorylation and reduced nuclear factor-kappa B (NF-κB) activation by 35% (P=0.002), which was induced by TNF-α in THP-1 macrophages. Furthermore, silencing of PTX3 in apoptotic cells resulted in increased macrophage binding, elevated expression rate of HLA-DR (+30%, P=0.015) and CD86 (+204%, P=0.004) positive cells, and induction of IL-1β (+36%, P=0.024) production. Conversely, adding recombinant PTX3 to macrophages reduced CD86 and HLA-DR expression in a dose-dependent manner.ConclusionsWe identified PTX3 as a novel regulator of macrophage activity, and this function suggests that PTX3 acts to resolve inflammation.     

Phospholipases D1 and D2 coordinately regulate macrophage phagocytosis

Phagocytosis is a fundamental feature of the innate immune system, required for antimicrobial defense, resolution of inflammation, and tissue remodeling. Furthermore, phagocytosis is coupled to a diverse range of cytotoxic effector mechanisms, including the respiratory burst, secretion of inflammatory mediators and Ag presentation. Phospholipase D (PLD) has been linked to the regulation of phagocytosis and subsequent effector responses, but the identity of the PLD isoform(s) involved and the molecular mechanisms of activation are unknown. We used primary human macrophages and human THP-1 promonocytes to characterize the role of PLD in phagocytosis. Macrophages, THP-1 cells, and other human myelomonocytic cells expressed both PLD1 and PLD2 proteins. Phagocytosis of complement-opsonized zymosan was associated with stimulation of the activity of both PLD1 and PLD2, as demonstrated by a novel immunoprecipitation-in vitro PLD assay. Transfection of dominant-negative PLD1 or PLD2 each inhibited the extent of phagocytosis (by 55-65%), and their combined effects were additive (reduction of 91%). PLD1 and PLD2 exhibited distinct localizations in resting macrophages and those undergoing phagocytosis, and only PLD1 localized to the phagosome membrane. The COS-7 monkey fibroblast cell line, which has been used as a heterologous system for the analysis of receptor-mediated phagocytosis, expressed PLD2 but not PLD1. These data support a model in which macrophage phagocytosis is coordinately regulated by both PLD1 and PLD2, with isoform-specific localization. Human myelomonocytic cell lines accurately model PLD-dependent signal transduction events required for phagocytosis, but the heterologous COS cell system does not.     

Long noncoding RNA THRIL promotes foam cell formation and inflammation in macrophages

Tumor necrosis factor-α (TNF-α) and heterogenous nuclear ribonucleoprotein L (hnRNPL)-related immunoregulatory lincRNA (THRIL) is a long noncoding RNA (lncRNA) involved in various inflammatory diseases. However, its role in atherosclerosis is not known. In this study, we aimed to investigate the function of THRIL in mediating macrophage inflammation and foam cell formation. The expression of THRIL was quantified in THP-1 macrophages after treatment with oxidized low-density lipoprotein (oxLDL). The effect of THRIL overexpression and knockdown on oxLDL-induced inflammatory responses and lipid accumulation was determined. THRIL-associated protein partners were identified by RNA pull-down and RNA immunoprecipitation assays. We show that THRIL is upregulated in macrophages after oxLDL treatment. Knockdown of THRIL blocks oxLDL-induced expression of interleukin-1β (IL-1β), IL-6, and TNF-α and lipid accumulation. Conversely, ectopic expression of THRIL enhances inflammatory gene expression and lipid deposition in oxLDL-treated macrophages. Moreover, THRIL depletion increases cholesterol efflux from macrophages and the expression of ATP-binding cassette transporter (ABC) A1 and ABCG1. FOXO1 is identified as a protein partner of THRIL and promotes macrophage inflammation and lipid accumulation. Furthermore, overexpression of FOXO1 restores lipid accumulation and inflammatory cytokine production in THRIL-depleted macrophages. In conclusion, our data suggest a model where THRIL interacts with FOXO1 to promote macrophage inflammation and foam cell formation. THRIL may represent a therapeutic target for atherosclerosis.     

Impaired phagocytosis in localized aggressive periodontitis: rescue by Resolvin E1

Resolution of inflammation is an active temporally orchestrated process demonstrated by the biosynthesis of novel proresolving mediators. Dysregulation of resolution pathways may underlie prevalent human inflammatory diseases such as cardiovascular diseases and periodontitis. Localized Aggressive Periodontitis (LAP) is an early onset, rapidly progressing form of inflammatory periodontal disease. Here, we report increased surface P-selectin on circulating LAP platelets, and elevated integrin (CD18) surface expression on neutrophils and monocytes compared to healthy, asymptomatic controls. Significantly more platelet-neutrophil and platelet-monocyte aggregates were identified in circulating whole blood of LAP patients compared with asymptomatic controls. LAP whole blood generates increased pro-inflammatory LTB4 with addition of divalent cation ionophore A23187 (5 μM) and significantly less, 15-HETE, 12-HETE, 14-HDHA, and lipoxin A(4). Macrophages from LAP subjects exhibit reduced phagocytosis. The pro-resolving lipid mediator, Resolvin E1 (0.1-100 nM), rescues the impaired phagocytic activity in LAP macrophages. These abnormalities suggest compromised resolution pathways, which may contribute to persistent inflammation resulting in establishment of a chronic inflammatory lesion and periodontal disease progression.     

Hepatitis C Virus Induces Interleukin-1β (IL-1β)/IL-18 in Circulatory and Resident Liver Macrophages

Hepatitis C virus (HCV)-mediated chronic liver disease is a global health problem, and inflammation is believed to be an important player in disease pathogenesis. HCV infection often leads to severe fibrosis/cirrhosis and hepatocellular carcinoma, although the mechanisms for advancement of disease are not fully understood. The proinflammatory cytokines interleukin-1β (IL-1β) and IL-18 have critical roles in establishment of inflammation. In this study, we examined induction of IL-1β/IL-18 secretion following HCV infection. Our results demonstrated that monocyte-derived human macrophages (THP-1) incubated with cell culture-grown HCV enhance the secretion of IL-1β/IL-18 into culture supernatants. A similar cytokine release was also observed for peripheral blood mononuclear cell (PBMC)-derived primary human macrophages and Kupffer cells (liver-resident macrophages) upon incubation with HCV. THP-1 cells incubated with HCV led to caspase-1 activation and release of proinflammatory cytokines. Subsequent studies demonstrated that HCV induces pro-IL-1β and pro-IL-18 synthesis via the NF-κB signaling pathway in macrophages. Furthermore, introduction of HCV viroporin p7 RNA into THP-1 cells was sufficient to cause IL-1β secretion. Together, our results suggested that human macrophages exposed to HCV induce IL-1β and IL-18 secretion, which may play a role in hepatic inflammation.     

Inulin stimulates phagocytosis of PMA-treated THP-1 macrophages by involvement of PI3-kinases and MAP kinases

Inulin is a polysaccharide that enhances various immune responses, mainly to T and B cells, natural killer cells, and macrophages in vivo and in vitro. Previous reports describe that inulin activates macrophages indirectly by affecting the alternative complement pathway. In this study, we examined the direct effect of inulin on PMA-treated THP-1 macrophages. Inulin treatment did not stimulate the proliferation of THP-1 macrophages at all. However, inulin treatment significantly increased phagocytosis of the polystyrene beads without the influence of serum. Doses of around 1 mg/mL had the maximal effect, and significant progression of phagocytosis occurred at times treated over 6 h. Inulin augmented phagocytosis not only with polystyrene beads but also with apoptotic cancer cells. The inulin-induced phagocytosis uptake was suppressed in Toll-like receptor (TLR) 4 mutated C3H/HeJ mice peritoneal macrophages. Moreover, inulin-induced THP-1 macrophage TNF-α secretion was inhibited using a blocking antibody specific to TLR4, suggesting that TLR4 is involved in the binding of inulin to macrophages. Furthermore, we used specific kinase inhibitors to assess the involvement of inulin-induced phagocytosis and revealed that phosphoinositide 3-kinase and mitogen-activated protein kinase, especially p38, participated in phagocytosis. These results suggest that inulin affects macrophages directly by involving the TLR4 signaling pathway and stimulating phagocytosis for enhancing immunomodulation.     

Basic calcium phosphate crystals induce monocyte/macrophage IL-1β secretion through the NLRP3 inflammasome in vitro

Basic calcium phosphate (BCP) crystals are associated with severe osteoarthritis and acute periarticular inflammation. Three main forms of BCP crystals have been identified from pathological tissues: octacalcium phosphate, carbonate-substituted apatite, and hydroxyapatite. We investigated the proinflammatory effects of these BCP crystals in vitro with special regard to the involvement of the NLRP3-inflammasome in THP-1 cells, primary human monocytes and macrophages, and mouse bone marrow-derived macrophages (BMDM). THP-1 cells stimulated with BCP crystals produced IL-1β in a dose-dependent manner. Similarly, primary human cells and BMDM from wild-type mice also produced high concentrations of IL-1β after crystal stimulation. THP-1 cells transfected with short hairpin RNA against the components of the NLRP3 inflammasome and mouse BMDM from mice deficient for NLRP3, apoptosis-associated speck-like protein, or caspase-1 did not produce IL-1β after BCP crystal stimulation. BCP crystals induced macrophage apoptosis/necrosis as demonstrated by MTT and flow cytometric analysis. Collectively, these results demonstrate that BCP crystals induce IL-1β secretion through activating the NLRP3 inflammasome. Furthermore, we speculate that IL-1 blockade could be a novel strategy to inhibit BCP-induced inflammation in human disease.     

Curcumin steers THP-1 cells under LPS and mTORC1 challenges toward phenotypically resting,low cytokine-producing macrophages

The persistent activation of intestinal mechanistic target of rapamycin complex 1 (mTORC1) triggered by mucosal stress has been linked to deregulation of the gut immune response resulting in intestinal inflammation and cell death. The present study investigated the regulatory properties of food-derived mTORC1 modulators, curcumin, and piperine, toward the polarization of stimulated macrophages and the differentiation of monocytes at two mTORC1 activity levels (baseline and elevated). To that end, we created stable human THP-1 monocytes exhibiting normal or constitutively active mTORC1. Curcumin or its combination with piperine, but not piperine alone, suppressed mTORC1 kinase activity, curtailed lipopolysaccharide-mediated inflammatory response of THP-1 macrophages, and repressed macrophage activation by inhibiting signaling pathways involved in M1 (mTORC1) and M2 (mTORC2 and cAMP response element binding protein) polarization. The effects of piperine in the curcumin/piperine combination were modest overall, indicating it was curcumin that modulated differentiating monocytes into acquiring a M0 macrophage phenotype characterized by low inflammatory cytokine output.     

氧化低密度脂蛋白对THP-1巨噬细胞吞噬和分泌功能的影响

目的:探讨氧化低密度脂蛋白(oxidized low density lipoprotein,ox-LDL)对巨噬细胞源性泡沫细胞吞噬功能和炎症相关因子分泌功能的影响。方法:利用佛波酯(phorbol ester,PMA)诱导THP-1细胞分化形成巨噬细胞,之后采用ox-LDL处理48小时后,诱导其形成泡沫细胞。利用中性红吞噬实验,分析泡沫细胞形成前后吞噬功能的变化;通过ELISA法,检测细胞培养上清中肿瘤坏死因子α(tumor necrosis factorα,TNF-α)含量,观察ox-LDL对THP-1巨噬细胞功能的影响。结果:细胞形态学结果表明,我们成功利用ox-LDL诱导THP-1巨噬细胞形成泡沫细胞;进一步发现ox-LDL诱导THP-1巨噬细胞表面的清道夫受体CD36表达升高,并促进细胞吞噬功能增加,进一步促进细胞内胆固醇含量显著升高(P0.05);同时,ox-LDL能够刺激巨噬细胞大量分泌TNF-α(P0.05)。结论:ox-LDL通过增强清道夫受体CD36表达,提高巨噬细胞的吞噬功能,引起大量胆固醇聚集,产生细胞毒性损伤,并促进TNF-α炎性因子的大量分泌。     

Akt2, but not Akt1 or Akt3 mediates pressure-stimulated serum-opsonized latex bead phagocytosis through activating mTOR and p70 S6 kinase

Monocytes and macrophages play critical roles in innate host defense and are sensitive to mechanical stimuli. Tissue pressure is often altered in association with inflammation or infection. Low pressure (20 mmHg), equivalent to normal tissue pressure, increases phagocytosis by primary monocytes and PMA-differentiated THP-1 macrophages, in part by FAK and ERK inhibition and p38 activation. PI-3K is required for macrophage phagocytosis, but whether PI-3K mediates pressure-stimulated phagocytosis is not known. Furthermore, little is known about the role played by the PI-3K downstream Kinases, Akt, and p70 S6 kinase (p70S6K) in modulating macrophage phagocytosis. Thus, we studied the contribution of PI-3K, Akt, and p70S6K to pressure-increased serum-opsonized bead phagocytosis. Pressure-induced p85 PI-3K translocation from cytosolic to membrane fractions and increased Akt activation by 36.1 +/- 12.0% in THP-1 macrophages. LY294002 or Akt inhibitor IV abrogated pressure-stimulated but not basal phagocytosis. Basal Akt activation was inhibited 90% by LY294002 and 70% by Akt inhibitor IV. Each inhibitor prevented Akt activation by pressure. SiRNA targeted to Akt1, Akt2, or Akt3 reduced Akt1, Akt2, and Akt3 expression by 50%, 45%, and 40%, respectively. However, only Akt2SiRNA abrogated the pressure-stimulated phagocytosis without affecting basal. Pressure also activated mTOR and p70S6K. mTORSiRNA and p70S6K inhibition by rapamycin or p70S6KSiRNA blocked pressure-induced, but not basal, phagocytosis. Changes in tissue pressure during inflammation may regulate macrophage phagocytosis by activation of PI-3K, which activates Akt2, mTOR, and p70S6K.     

Suppression of TNF-α and IL-1 signaling identifies a mechanism of homeostatic regulation of macrophages by IL-27

IL-27 is a pleiotropic cytokine with both activating and inhibitory functions on innate and acquired immunity. IL-27 is expressed at sites of inflammation in cytokine-driven autoimmune/inflammatory diseases, such as rheumatoid arthritis, psoriasis, inflammatory bowel disease, and sarcoidosis. However, its role in modulating disease pathogenesis is still unknown. In this study, we found that IL-27 production is induced by TNF-α in human macrophages (MΦ) and investigated the effects of IL-27 on the responses of primary human MΦ to the endogenous inflammatory cytokines TNF-α and IL-1. In striking contrast to IL-27-mediated augmentation of TLR-induced cytokine production, we found that IL-27 suppressed MΦ responses to TNF-α and IL-1β, thus identifying an anti-inflammatory function of IL-27. IL-27 blocked the proximal steps of TNF-α signaling by downregulating cell-surface expression of the signaling receptors p55 and p75. The mechanism of inhibition of IL-1 signaling was downregulation of the ligand-binding IL-1RI concomitant with increased expression of the receptor antagonist IL-1Ra and the decoy receptor IL-1RII. These findings provide a mechanism for suppressive effects of IL-27 on innate immune cells and suggest that IL-27 regulates inflammation by limiting activation of MΦ by inflammatory cytokines while preserving initial steps in host defense by augmenting responses to microbial products.     

Amyloid peptide-induced cytokine and chemokine expression in THP-1 monocytes is blocked by small inhibitory RNA duplexes for early growth response-1 messenger RNA

In Alzheimer's disease (AD) one finds increased deposition of A beta and also an increased presence of monocytes/macrophages in the vessel wall and activated microglial cells in the brain. AD patients show increased levels of proinflammatory cytokines by activated microglia. Here we used a human monocytic THP-1 cell line as a model for microglia to delineate the cellular signaling mechanism involved in amyloid peptides (A beta(1-40) and A beta(1-42))-induced expression of inflammatory cytokines and chemokines. We observed that A beta peptides at physiological concentrations (125 nM) increased mRNA expression of cytokines (TNF-alpha, and IL-1 beta) and chemokines (monocyte chemoattractant protein-1 (MCP-1), IL-8, and macrophage inflammatory protein-1 beta (MIP-1 beta)). The cellular signaling involved activation of c-Raf, extracellular signal-regulated kinase-1 (ERK-1)/ERK-2, and c-Jun N-terminal kinase, but not p38 mitogen-activated protein kinase. This is further supported by the data showing that A beta causes phosphorylation of ERK-1/ERK-2, which, in turn, activates Elk-1. Furthermore, A beta mediated a time-dependent increase in DNA binding activity of early growth response-1 (Egr-1) and AP-1, but not of NF-kappa B and CREB. Moreover, A beta-induced Egr-1 DNA binding activity was reduced >60% in THP-1 cells transfected with small interfering RNA duplexes for Egr-1 mRNA. We show that A beta-induced expression of TNF-alpha, IL-1 beta, MCP-1, IL-8, and MIP-1 beta was abrogated in Egr-1 small inhibitory RNA-transfected cells. Our results indicate that A beta-induced expression of cytokines (TNF-alpha and IL-1 beta) and chemokines (MCP-1, IL-8, and MIP-1 beta) in THP-1 monocytes involves activation of ERK-1/ERK-2 and downstream activation of Egr-1. The inhibition of Egr-1 by Egr-1 small inhibitory RNA may represent a potential therapeutic target to ameliorate the inflammation and progression of AD.     

IL-1β Production through the NLRP3 Inflammasome by Hepatic Macrophages Links Hepatitis C Virus Infection with Liver Inflammation and Disease

Chronic hepatitis C virus (HCV) infection is a leading cause of liver disease. Liver inflammation underlies infection-induced fibrosis, cirrhosis and liver cancer but the processes that promote hepatic inflammation by HCV are not defined. We provide a systems biology analysis with multiple lines of evidence to indicate that interleukin-1β (IL-1β) production by intrahepatic macrophages confers liver inflammation through HCV-induced inflammasome signaling. Chronic hepatitis C patients exhibited elevated levels of serum IL-1β compared to healthy controls. Immunohistochemical analysis of healthy control and chronic hepatitis C liver sections revealed that Kupffer cells, resident hepatic macrophages, are the primary cellular source of hepatic IL-1β during HCV infection. Accordingly, we found that both blood monocyte-derived primary human macrophages, and Kupffer cells recovered from normal donor liver, produce IL-1β after HCV exposure. Using the THP-1 macrophage cell-culture model, we found that HCV drives a rapid but transient caspase-1 activation to stimulate IL-1β secretion. HCV can enter macrophages through non-CD81 mediated phagocytic uptake that is independent of productive infection. Viral RNA triggers MyD88-mediated TLR7 signaling to induce IL-1β mRNA expression. HCV uptake concomitantly induces a potassium efflux that activates the NLRP3 inflammasome for IL-1β processing and secretion. RNA sequencing analysis comparing THP1 cells and chronic hepatitis C patient liver demonstrates that viral engagement of the NLRP3 inflammasome stimulates IL-1β production to drive proinflammatory cytokine, chemokine, and immune-regulatory gene expression networks linked with HCV disease severity. These studies identify intrahepatic IL-1β production as a central feature of liver inflammation during HCV infection. Thus, strategies to suppress NLRP3 or IL-1β activity could offer therapeutic actions to reduce hepatic inflammation and mitigate disease.     

Decreased alveolar macrophage apoptosis is associated with increased pulmonary inflammation in a murine model of pneumococcal pneumonia

Regulation of the inflammatory infiltrate is critical to the successful outcome of pneumonia. Alveolar macrophage apoptosis is a feature of pneumococcal infection and aids disease resolution. The host benefits of macrophage apoptosis during the innate response to bacterial infection are incompletely defined. Because NO is required for optimal macrophage apoptosis during pneumococcal infection, we have explored the role of macrophage apoptosis in regulating inflammatory responses during pneumococcal pneumonia, using inducible NO synthase (iNOS)-deficient mice. iNOS(-/-) mice demonstrated decreased numbers of apoptotic macrophages as compared with wild-type C57BL/6 mice following pneumococcal challenge, greater recruitment of neutrophils to the lung and enhanced expression of TNF-alpha. Pharmacologic inhibition of iNOS produced similar results. Greater pulmonary inflammation was associated with greater levels of early bacteremia, IL-6 production, lung inflammation, and mortality within the first 48 h in iNOS(-/-) mice. Labeled apoptotic alveolar macrophages were phagocytosed by resident macrophages in the lung and intratracheal instillation of exogenous apoptotic macrophages decreased neutrophil recruitment in iNOS(-/-) mice and decreased TNF-alpha mRNA in lungs and protein in bronchial alveolar lavage, as well as chemokines and cytokines including IL-6. These changes were associated with a lower probability of mice becoming bacteremic. This demonstrates the potential of apoptotic macrophages to down-regulate the inflammatory response and for the first time in vivo demonstrates that clearance of apoptotic macrophages decreases neutrophil recruitment and invasive bacterial disease during pneumonia.     

Resolvin d1 and aspirin-triggered resolvin d1 promote resolution of allergic airways responses

Asthma is a disease of airway inflammation that in most cases fails to resolve. The resolution of inflammation is an active process governed by specific chemical mediators, including D-series resolvins. In this study, we determined the impact of resolvin D1 (RvD1) and aspirin-triggered RvD1 (AT-RvD1) on the development of allergic airway responses and their resolution. Mice were allergen sensitized, and RvD1, AT-RvD1 (1, 10, or 100 ng), or vehicle was administered at select intervals before or after aerosol allergen challenge. RvD1 markedly decreased airway eosinophilia and mucus metaplasia, in part by decreasing IL-5 and IκBα degradation. For the resolution of established allergic airway responses, AT-RvD1 was even more efficacious than RvD1, leading to a marked decrease in the resolution interval for lung eosinophilia, decrements in select inflammatory peptide and lipid mediators, and more rapid resolution of airway hyperreactivity to methacholine. Relative to RvD1, AT-RvD1 resisted metabolic inactivation by macrophages, and AT-RvD1 significantly enhanced macrophage phagocytosis of IgG-OVA-coated beads in vitro and in vivo, a new proresolving mechanism for the clearance of allergen from the airways. In conclusion, RvD1 and AT-RvD1 can serve as important modulators of allergic airway responses by decreasing eosinophils and proinflammatory mediators and promoting macrophage clearance of allergen. Together, these findings identify D-series resolvins as potential proresolving therapeutic agents for allergic responses.     

Osteopontin Deficiency Accelerates Spontaneous Colitis in Mice with Disrupted Gut Microbiota and Macrophage Phagocytic Activity

Background

Osteopontin (OPN) is a multifunctional protein expressed in a variety of tissues and cells. Recent studies revealed increased OPN expression in the inflamed intestinal tissues of patients with inflammatory bowel disease (IBD). The role of OPN in the pathophysiology of IBD, however, remains unclear.

Aims

To investigate the role of OPN in the development of intestinal inflammation using a murine model of IBD, interleukin-10 knock out (IL-10 KO) mice.

Methods

We compared the development of colitis between IL-10 KO and OPN/IL-10 double KO (DKO) mice. OPN expression in the colonic tissues of IL-10 KO mice was examined by fluorescence in situ hybridization (FISH) analysis. Enteric microbiota were compared between IL-10 KO and OPN/IL-10 DKO mice by terminal restriction fragment length polymorphism analysis. The effect of OPN on macrophage phagocytic function was evaluated by phagocytosis assay.

Results

OPN/IL-10 DKO mice had an accelerated onset of colitis compared to IL-10 KO mice. FISH analysis revealed enhanced OPN synthesis in the colonic epithelial cells of IL-10 KO mice. OPN/IL-10 DKO mice had a distinctly different enteric bacterial profile with a significantly lower abundance of Clostridium subcluster XIVa and a greater abundance of Clostridium cluster XVIII compared to IL-10 KO mice. Intracellular OPN deletion in macrophages impaired phagocytosis of fluorescence particle-conjugated Escherichia coli in vitro. Exogenous OPN enhanced phagocytosis by OPN-deleted macrophages when administered at doses of 1 to 100 ng/ml, but not 1000 ng/ml.

Conclusions

OPN deficiency accelerated the spontaneous development of colitis in mice with disrupted gut microbiota and macrophage phagocytic activity.