Stimulates Macrophage Activation Research Articles

Role of histone deacetylase 3 (HDAC3) in adipose tissue metabolism and immunophenotype: Selected Abstract - SITeCS Congress 2022

Introduction: Obesity is associated with comorbidities such as cardiovascular disease and type 2 diabetes. HDAC3 regulates adipose tissue physiology (WAT), and its genetic inactivation causes metabolic reprogramming of white adipocytes toward browning. The aim of this work is to evaluate the effect of HDAC3 silencing at different stages of differentiation and investigate the influence of adipocyte metabolism on the immunophenotype of WAT. Materials and Methods: Following HDAC3 silencing in mesenchymal stem cells and mature adipocytes, adipocyte function, RNA, DNA and protein levels, and proliferation at the end of differentiation were analyzed. Visceral WAT immunophenotype (vWAT) of Hdac3 KO mice in WAT (Hdac3fatKO) and controls (FL) was performed by FACS. Results: Silencing HDAC3 in precursors amplifies the expression of genes and proteins that regulate differentiation, oxidative metabolism, browning and mitochondrial activity. Following silencing, we found increased 1)phosphorylation of AKT (1.64 fold change, P<0.0001), indicative of increased insulin signaling, and 2)proliferation, characteristic of the early phase of differentiation. Mitochondrial content was unchanged, but increased mitochondrial activity was observed in terms of maximal respiration (1.42 fold change, P=0.0151) and uncoupling of the electron transport chain (+11.6%, P<0.0001). No difference was observed following HDAC3 silencing in mature adipocytes. We hypothesized that the enhancement of oxidative metabolism may cause cellular damage or senescence and, consequently, the immunophenotype of vWAT might be affected by HDAC3 ablation. Analysis reveals an increase of macrophages (2.48 fold change, P=0.0311) in the vWAT of Hdac3fatKO mice polarizing toward the M2 population. Coculture of adipocytes with macrophages from bone marrow indicates that HDAC3 silencing in adipocytes stimulates macrophage activation. Conclusions: HDAC3 is a key factor in the WAT phenotype, and its inactivation triggers mechanisms that support browning. Early epigenetic events mediated by HDAC3 silencing are crucial in directing adipocyte precursors toward the oxidative phenotype. Finally, results obtained from ex vivo and in vitro studies suggest that specific factors produced by KO adipocytes may be involved in determining the observed immunophenotype. [FONDAZIONE CARIPLO 2015-0641]

Abstract 763: Minimally invasive colorectal resection is associated with significantly elevated plasma levels of extracellular cold inducible RNA binding protein for a month after colorectal resection & may promote the growth of residual metastases

Abstract Introduction: Cold-Inducible RNA-Binding Protein (CIRP) is similar to a class of stress-induced RNA binding proteins and is mostly found in the nucleus where it regulates transcription. In response to stress, CIRP moves to the cytoplasm where it regulates translation; a fraction of the cytoplasmic CIRP is released extracellularly (eCIRP). eCIRP is a damage-associated molecular pattern (DAMP) molecule that promotes inflammatory responses including TNF-alpha, IL-6, and HMGB production. Lymphocytes, macrophages, and PMN’s are activated after surgery and can generate eCIRP. Elevated eCIRP blood levels may impact macrophage function and induce immune tolerance that may promote tumor growth and progression. This study’s purpose was to measure plasma eCIRP levels before and for a month after MICR for CRC. Methods: CRC patients (pts) in an IRB approved data/tissue bank who underwent MICR for whom plasma was available were selected. Demographic, clinical and pathology data were reviewed. Plasma samples were collected preoperatively (preOp) and at 6 post-operative (Postop) time points (POD 1, 3, 7-13, 14-20, 21-27, 28-41); samples were processed and frozen until used. Late samples were bundled into 7-day blocks and considered as single time points. eCIRP levels were analyzed in duplicate using ELISA. The Wilcoxon paired t-test was used for analysis. Results: 83 CRC pts who underwent MICR were studied (colon 66%, rectal 34%; mean age 65); 26% had Hand assisted (HA) MIS (mean incision length[IL] 10.0±2.9cm) and 69% laparoscopic (LAP, mean IL 6.9±4.1cm) MICR. The cancer stage breakdown was; I, 30%; II, 32%; III, 35% and IV, 3%. The mean PreOp eCIRP level was 896.8± 757.0pg/ml; vs. preop levels increased mean plasma levels (p=<0.001) were noted on POD1 (2549± 2632pg/ml, n= 83), POD3 (1871± 1362pg/ml, n= 77), POD7-13 (1788± 1403pg/ml, n=57), POD14-20 (1473± 738.8pg/ml, n=30), and POD 21-27 (1681± 1375pg/ml, n= 21). With regard to MIS method, greater eCIRP increases were noted in the HA vs the Lap group from POD1-POD20 (p=0.01-0.04). Conclusions: Plasma eCIRP levels are elevated for a month after MICR for CRC (% change range 100-184) and peaked on POD1. The cause of the early increase may be the acute inflammatory response whereas the late elevations may be related to wound healing-related tissue remodeling. Higher mean levels in the HA vs Lap group suggests that the extent of surgical trauma impacts eCIRP levels postop. Previous studies noted increased post-MICR plasma levels of MCP-1, CHI3L1 and IL8 that stimulate macrophage activity. eCIRP, together with those proteins may promote macrophage recruitment and function and might promote the growth of residual tumor deposits postop. Further studies are warranted. Citation Format: Chandana S. K. Herath Mudiyanselage, Xiaohong Yan, Neil Mitra, Hansani N. Angammana, Doaa F. Morrar, Vincent Chia, Yanni Hedjar, Vesna Cekic, Jennifer L. Agnew, Richard L. Whelan. Minimally invasive colorectal resection is associated with significantly elevated plasma levels of extracellular cold inducible RNA binding protein for a month after colorectal resection & may promote the growth of residual metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 763.

Immunopotentiation mechanism of a mannose-modified peptide fragment containing a dominant antigenic epitope of Micropterus salmoides rhabdovirus (MSRV) glycoprotein after coupling to nanocarrier

Micropterus salmoides rhabavirus (MSRV) is an important pathogen that threatens the aquaculture of bass farming worldwide, and the efficacy of vaccines developed against this pathogen still needs to be improved. In order to further enhance the immune efficacy of existing vaccines, a mannose-modified nanovector vaccine SWCNTs-M-G2 was constructed by screening for peptide fragments encoding highly immunogenic antigenic epitopes (G2) and mannose modifications. Mannose receptor (MR) is abundantly expressed on the surface of Antigen-presenting cells (APCs) and plays a role in recognizing pathogens and presenting antigens. In this study, the effect of mannosylated modification on the immunopotentiation of highly immunogenic peptide fragments was investigated by using the characteristics of MR-specific recognition of mannose and APCs surface. In vitro macrophage assays showed that SWCNTs-M-G2 was efficiently phagocytosed and taken up by largemouth black bass head kidney macrophages, and effectively stimulated macrophage activation and upregulated the secretion of macrophage cytokines IL-1β, IL-8 and IL-12. In vivo, SWCNTs-M-G2 was also shown to be better taken up by immune organs such as spleen and head kidney, and induced increased expression of IL-1β, IL-8 and IL-12 in the immune organs of Micropterus salmoides, and promoted the expression of TCR and CD4 molecules in specific immune cells. Meanwhile, in vivo fluorescence imaging revealed that SWCNTs-M-G2 could be in the immune organ of fish for a longer period of time, contributing to the efficient uptake of antigens. Finally, the challenge test showed that SWCNTs-M-G2 could mediate the production of specific antibodies at a higher level than the unmodified carrier vaccine SWCNTs-G2 and showed a higher relative immune protection rate.

Excess Uric Acid Induces Gouty Nephropathy Through Crystal Formation: A Review of Recent Insights

Uric acid (UA) is the final product of purine metabolism in the human body, and impaired purine metabolism can increase the uric acid in serum, finally resulting in hyperuricemia (HUA). Current evidences suggest that urates might have antioxidant properties under certain circumstances, but most evidences suggest that urates promote inflammation. Hyperuricemia leads to the formation of urate crystals, which might be recognized as a red flag by the immune system. Such a response stimulates macrophage activation, leads to the activation of NOD-like receptor protein 3 (NLRP3) inflammasome vesicles, and ultimately the production and liberation of interleukin-1b (IL-1b) and interleukin-18 (IL-18), which can mediate inflammation, apoptosis and necroinflammation and cause an inflammatory cascade response. The kidney is one of the most commonly affected organs in HUA, which promotes the development of chronic kidney disease (CKD) by damaging endothelial cells, activating the renin-angiotensin system (RAS), and promoting inflammatory responses. Pharmacological interventions and lifestyle modifications are the primary means for controlling gout and lowering UA. The febuxostat is safe for CKD patients in the UA lowering therapy. Although dialysis can reduce UA levels, the application of drug is also necessary for dialysis patients. This article reviews the synthesis and metabolism of UA, etiology of HUA, the relationship between HUA and kidney disease, the treatment of gout and gouty nephropathy (GN).

Endothelial pannexin-1 channels modulate macrophage and smooth muscle cell activation in abdominal aortic aneurysm formation

Pannexin-1 (Panx1) channels have been shown to regulate leukocyte trafficking and tissue inflammation but the mechanism of Panx1 in chronic vascular diseases like abdominal aortic aneurysms (AAA) is unknown. Here we demonstrate that Panx1 on endothelial cells, but not smooth muscle cells, orchestrate a cascade of signaling events to mediate vascular inflammation and remodeling. Mechanistically, Panx1 on endothelial cells acts as a conduit for ATP release that stimulates macrophage activation via P2X7 receptors and mitochondrial DNA release to increase IL-1β and HMGB1 secretion. Secondly, Panx1 signaling regulates smooth muscle cell-dependent intracellular Ca2+ release and vascular remodeling via P2Y2 receptors. Panx1 blockade using probenecid markedly inhibits leukocyte transmigration, aortic inflammation and remodeling to mitigate AAA formation. Panx1 expression is upregulated in human AAAs and retrospective clinical data demonstrated reduced mortality in aortic aneurysm patients treated with Panx1 inhibitors. Collectively, these data identify Panx1 signaling as a contributory mechanism of AAA formation.

Limonene-carvacrol: A combination of monoterpenes with enhanced antileishmanial activity

BackgroundLeishmaniasis is a parasitosis with a wide incidence in developing countries. The drugs which are indicated for the treatment of this infection usually are able to promote high toxicity. PurposeA combination of limonene and carvacrol, monoterpenes present in plants with antiparasitic activity may constitute an alternative for the treatment of these diseases. MethodsIn this study, the antileishmania activity against Leishmania major, cytotoxicity tests, assessment of synergism, parasite membrane damage tests as well as molecular docking and immunomodulatory activity of limonene–carvacrol (Lim–Car) combination were evaluated. ResultsThe Lim–Car combination (5:0; 1:1; 1:4; 2:3; 3:2; 4:1 and 0:5) showed potential antileishmania activity, with mean inhibitory concentration (IC50) ranging from 5.8 to 19.0 μg.mL−1. They demonstrated mean cytotoxic concentration (CC50) ranging from 94.1 to 176.0 μg.mL−1, and did not show significant hemolytic effect. In the investigation of synergistic interaction, the 4:1 Lim–Car combination showed better fractional inhibitory concentration (FIC) index as well as better activity on amastigotes and IS. The samples caused considerable damage to the parasite membrane this monoterpene activity seems to be more related to Trypanothione Reductase (TryR) enzyme interaction, demonstrated in the molecular docking assay. In addition, the 4:1 Lim–Car combination stimulated macrophage activation, and showed at was the best association, with reduction of infection and infectivity of parasitized macrophages. ConclusionThe 4:1 Lim–Car combination appears to be a promising candidate as a monotherapeutic antileishmania agent.

Insulin-like growth factor binding protein 3 gene of golden pompano (TroIGFBP3) promotes antimicrobial immune defense

Insulin-like growth factor binding protein 3 (IGFBP3), an important member of the IGFBP family, plays an important biological role in regulating cellular proliferation, differentiation, growth, apoptosis, and innate immunity. However, studies concerning IGFBP3 in teleosts are very limited and IGFBP3 function remains unclear. In this study, we conducted both in vivo and in vitro functional analyses of an IGFBP3 (TroIGFBP3) from the teleost fish golden pompano (Trachinotus ovatus). TroIGFBP3 is composed of 286 amino acid residues and shares a high amino acid sequence similarity (50.18%–93.71%) with other IGFBP3 sequences in humans and teleosts. TroIGFBP3 was widely distributed in various tissues, with the highest expression in the liver. TroIGFBP3 expression was significantly upregulated following Vibrio harveyi infection. The results of in vitro assays showed that TroIGFBP3 could stimulate macrophage activation and promote peripheral blood leukocytes (PBLs) proliferation. Meanwhile, TroIGFBP3 overexpression significantly inhibited bacterial infection in fish tissues, whereas TroIGFBP3 knockdown resulted in increased bacterial dissemination and colonization in golden pompano tissues in vivo. Furthermore, recombinant TroIGFBP3 could inhibit cellular proliferation and promote apoptosis of mouse tumor cells. Taken together, these results indicated that TroIGFBP3 plays a significant role in innate antibacterial immunity and provides a theoretical foundation for investigating the function of IGFBP3 in fish immune response.

Disruption of hepatic small heterodimer partner induces dissociation of steatosis and inflammation in experimental nonalcoholic steatohepatitis

Disruption of hepatic small heterodimer partner induces dissociation of steatosis and inflammation in experimental nonalcoholic steatohepatitis

Growth hormone-mediated reprogramming of macrophage transcriptome and effector functions

Macrophages are an important component of the innate immune response. Priming and activation of macrophages is stimulated by cytokines (i.e IFNγ). However, growth hormone (GH) can also stimulate macrophage activation. Based on these observations, the goal of this work was to 1) to compare the transcriptome profile of macrophages activated in vitro with GH and IFNγ, and 2) to assess the impact of GH on key macrophage functional properties like reactive oxygen species (ROS) production and phagosomal proteolysis. To assess the global transcriptional and functional impact of GH on macrophage programming, bone marrow derived macrophages were treated with GH or IFNγ. Our data strongly support a potential link between GH, which wanes with age, and impaired macrophage function. The notable overlap of GH with IFNγ-induced pathways involved in innate immune sensing of pathogens and antimicrobial responses argue for an important role for GH in macrophage priming and maturation. By using functional assays that report on biochemical activities within the lumen of phagosomes, we have also shown that GH alters physiologically relevant processes such as ROS production and proteolysis. These changes could have far reaching impacts on antimicrobial capacity, signaling, and antigen presentation.

Intestinal dysbacteriosis activates tumor-associated macrophages to promote epithelial-mesenchymal transition of colorectal cancer.

In this study we investigated the association between intestinal dysbacteriosis with colorectal cancer progress and the underlying molecular mechanisms. Tumor progression was evaluated using xenograft mice model. The epithelial-mesenchymal transition (EMT) markers were quantified by both real-time PCR and immunoblotting. The serum content of IL-6 and TNF-α were measured with ELISA kits. Cell proliferation was determined by the Cell Counting Kit-8. Intestinal dysbacteriosis was successfully simulated by the administration of a large dose of antibiotics and was demonstrated to promote xenograft tumor growth and induce EMT. Accordingly, the serum concentrations of cytokines IL-6 and TNF-α were significantly increased. Furthermore, the production and secretion of IL-6 and TNF-α were remarkably elevated in macrophages isolated from intestinal dysbiotic mice in comparison with the normal counterparts, and conditioned medium from these was shown to significantly stimulate EMT process in HT29 cells in vitro. Macrophage depletion completely abrogated the pro-tumor effect of intestinal dysbacteriosis. Our results suggest that intestinal dysbacteriosis stimulates macrophage activation and subsequently induces EMT process via secreted pro-inflammatory cytokines IL-6 and TNF-α.

Critical Role of Plasmin in Macrophage Activation During Liver Injury

Activation of hepatic macrophages is critical for liver repair after injury. The mechanism by which liver injury stimulates macrophage activation is not fully understood. We tested the hypothesis that the fibrinolytic enzyme, plasmin, is critical for macrophage activation after liver injury. To test this hypothesis, mice were exposed to a hepatotoxic dose of acetaminophen followed by treatment with tranexamic acid (1200 mg/kg i.p., administered twice daily), a drug that inhibits the conversion of plasminogen to plasmin. Exposure of mice to acetaminophen stimulated rapid macrophage activation, increasing cytokine production and macrophage‐mediated phagocytosis of necrotic cells. This activation was reduced by plasmin inhibition, leading to impaired liver repair. Next, we determined whether plasmin directly activates macrophages. Treatment of either bone marrow‐derived macrophages or Kupffer cells with plasmin increased expression of the proinflammatory cytokines Cxcl1, Cxcl2, and tumor necrosis factor‐α in an Erk1/2 and p38‐dependent manner. Studies have indicated a role for high‐mobility group B1 protein (HMGB1), a damage‐associated molecular pattern molecule, in the activation of macrophages. Therefore, we determined whether HMGB1 affects plasmin‐mediated activation of macrophages. While HMGB1 alone at concentrations that are detected in the serum of acetaminophen‐treated mice did not increase expression of proinflammatory cytokines in macrophages, it synergistically enhanced plasmin‐mediated upregulation of cytokines. Furthermore, necrotic hepatocytes from wild‐type mice enhanced plasmin‐mediated activation of macrophages, whereas necrotic hepatocytes from hepatocyte‐specific HMGB1 knockout mice did not. Collectively, these studies demonstrate that plasmin is an important activator of macrophages after liver injury. Further characterization of this pathway could lead to the development of novel therapies aimed at enhancing macrophage‐mediated liver repair in patients suffering from acute liver injury.Support or Funding InformationNIH grant 2 R01 DK073566NIH Training Grant T32 ES007255This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Immunostimulatory effect of aqueous extract of Coriandrum sativum L. seed on macrophages.

Coriandrum sativum L. seed is generally used as a spice and crude drug. Although many functions of the various components in C. sativum L. seed have been reported, the immunostimulatory effect of water-soluble components in C. sativum L. seed has not been studied. In the present study, we focused on the immunostimulatory effect of C. sativum L. seed aqueous extract (CAE) on macrophages as a novel health function of C. sativum L. seed components. CAE significantly enhanced the production of tumor necrosis factor (TNF)-α and interleukin (IL)-6 in both RAW264.7 cells and peritoneal macrophages by enhancing the expression levels of these cytokine genes. CAE also stimulated nitric oxide (NO) production and the phagocytosis activity in RAW264.7 cells. We suggest that the activity of CAE is a result of the upregulation of mitogen-activated protein kinase and nuclear factor-κB cascades via TLR4. In addition, IL-6 production by peritoneal macrophages collected from CAE-administered mice was significantly enhanced, suggesting that CAE could stimulate macrophage activity in vivo. The findings of the present study suggest that CAE contains a novel water-soluble component with an immunostimulatory effect on macrophages. CAE would contribute to activating host defense against pathogens by stimulating the innate immunity. © 2017 Society of Chemical Industry.

Immunostimulatory effect of dried bonito extract on mouse macrophage cell lines and mouse primary peritoneal macrophages.

Dried bonito is a preserved food used in Japan, which contains abundant flavor ingredients and functional substances. We focused on the immunostimulatory effect of dried bonito extract (DBE) on mouse macrophage-like J774.1 cells, RAW264.7 cells, and mouse primary peritoneal macrophages. DBE significantly stimulated the production of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) by both J774.1 cells and peritoneal macrophages by enhancing the cytokine gene expression levels. In addition, DBE stimulated nitric oxide production by enhancing the expression of inducible nitric oxide synthase in RAW264.7 cells. DBE also increased the phagocytosis activity of J774.1 cells. Immunoblot analysis revealed that DBE has an immunostimulatory effect on macrophages through activation of mitogen-activated protein kinase and nuclear factor-κB cascades. TNF-α production enhanced by DBE was partially inhibited by treatment with TLR4 inhibitor TAK-242, whereas IL-6 production enhanced by DBE was almost inhibited. These results suggested that DBE is thought to strongly stimulate the TLR4 signaling pathway for macrophage activation, and its activation is also involved in other signaling. Finally, the phagocytosis activity of peritoneal macrophages from DBE-administered BALB/c mice increased significantly, suggesting that DBE has the potential to stimulate macrophage activity in vivo. In conclusion, these data indicated that DBE contributes to activating host defense against pathogens by activating innate immunity.

Immunostimulatory effect of spinach aqueous extract on mouse macrophage-like J774.1 cells and mouse primary peritoneal macrophages.

We herein report the immunostimulatory effect of spinach aqueous extract (SAE) on mouse macrophage-like J774.1 cells and mouse primary peritoneal macrophages. SAE significantly enhanced the production of interleukin (IL)-6 and tumor necrosis factor-α by both J774.1 cells and peritoneal macrophages by enhancing the expression levels of these cytokine genes. In addition, the phagocytosis activity of J774.1 cells was facilitated by SAE. Immunoblot analysis revealed that SAE activates mitogen-activated protein kinase and nuclear factor-κB cascades. It was found that SAE activates macrophages through not only TLR4, but also other receptors. The production of IL-6 was significantly enhanced by peritoneal macrophages from SAE-administered BALB/c mice, suggesting that SAE has a potential to stimulate macrophage activity in vivo. Taken together, these data indicate that SAE would be a beneficial functional food with immunostimulatory effects on macrophages.

Plasminogen activator inhibitor-1 stimulates macrophage activation through Toll-like Receptor-4

While inflammation is often associated with increased Plasminogen Activator Inhibitor-1 (PAI-1), the functional consequences of PAI-1 in inflammation have yet to be fully determined. The aim of this study was to establish the in vivo relevance of PAI-1 in inflammation. A mouse model of systemic inflammation was employed in wild-type (WT) and PAI-1 deficient (PAI-1−/−) mice. Mice survival, macrophage infiltration into the lungs, and plasma levels of pro-inflammatory cytokines were assessed after lipopolysaccharide (LPS) infusion. In vitro experiments were conducted to examine changes in LPS-induced inflammatory responses after PAI-1 exposure. PAI-1 was shown to regulate inflammation, in vivo, and affect macrophage infiltration into lungs. Further, PAI-1 activated macrophages, and increased pro-inflammatory cytokines at both the mRNA and protein levels in these cells. The effect of PAI-1 on macrophage activation was dose-dependent and LPS-independent. Proteolytic inhibitory activity and Lipoprotein Receptor-related Protein (LRP) and vitronectin (VN) binding functions, were not involved in PAI-1-mediated activation of macrophages. However, the effect of PAI-1 on macrophage activation was partially blocked by a TLR4 neutralizing antibody. Furthermore, PAI-1-induced Tumor Necrosis Factor-alpha (TNF-α) and Macrophage Inflammatory Protein-2 (MIP-2) expression was reduced in TLR4−/− macrophages compared to WT macrophages. These results demonstrate that PAI-1 is involved in the regulation of host inflammatory responses through Toll-like Receptor-4 (TLR4)-mediated macrophage activation.

Triamcinolone acetonide activates an anti-inflammatory and folate receptor-positive macrophage that prevents osteophytosis in vivo.

IntroductionTriamcinolone acetonide (TA) is used for osteoarthritis management to reduce pain, and pre-clinical studies have shown that TA limits osteophyte formation. Osteophyte formation is known to be facilitated by synovial macrophage activation. TA injections might influence macrophage activation and subsequently reduce osteophytosis. Although widely applied in clinical care, the mechanism through which TA exerts this effect remains unknown. In this animal study, we investigated the in vivo effects of TA injections on macrophage activation, osteophyte development and joint degeneration. Furthermore, in vitro macrophage differentiation experiments were conducted to further explain working mechanisms of TA effects found in vivo.MethodsOsteoarthritis was induced in rat knees using papain injections and a running protocol. Untreated and TA-treated animals were longitudinally monitored for 12 weeks with in vivo micro–computed tomography (μCT) to measure subchondral bone changes. Synovial macrophage activation was measured in vivo using folate receptor β (FRβ)-targeted single-photon emission computed tomography/computed tomography. Articular cartilage was analyzed at 6 and 12 weeks with ex vivo contrast-enhanced μCT and histology. To further explain the outcomes of our in vivo study, TA on macrophages was also studied in vitro. These cultured macrophages were either M1- or M2-activated, and they were analyzed using fluorescence-activated cell sorting for CD163 and FRβ expression as well as for messenger RNA (mRNA) expression of interleukin (IL)-10.ResultsOur in vivo study showed that intra-articular injections with TA strongly enhanced FRβ+ macrophage activation. Despite stimulated macrophage activation, osteophyte formation was fully prevented. There was no beneficial effect of TA against cartilage degradation or subchondral bone sclerosis. In vitro macrophage cultures showed that TA strongly induced monocyte differentiation towards CD163+ and FRβ+ macrophages. Furthermore, TA-stimulated M2 macrophages showed enhanced IL-10 expression at the mRNA level.ConclusionsTA injections potently induce a CD163+- and FRβ+-activated macrophage with anti-inflammatory characteristics such as reduced IL-10 production in vitro and lack of osteophytosis in vivo.

Alcohol stimulates macrophage activation through caspase-dependent hepatocyte derived release of CD40L containing extracellular vesicles

The mechanisms by which hepatocyte exposure to alcohol activates inflammatory cells such as macrophages in alcoholic liver disease (ALD) are unclear. The role of released nano-sized membrane vesicles, termed extracellular vesicles (EV), in cell-to-cell communication has become increasingly recognized. We tested the hypothesis that hepatocytes exposed to alcohol may increase EV release to elicit macrophage activation. Primary hepatocytes or HepG2 hepatocyte cell lines overexpressing ethanol-metabolizing enzymes alcohol dehydrogenase (HepG2(ADH)) or cytochrome P450 2E1 (HepG2(Cyp2E1)) were treated with ethanol and EV release was quantified with nanoparticle tracking analysis. EV mediated macrophage activation was monitored by analysing inflammatory cytokines and macrophage associated mRNA expression, immunohistochemistry, biochemical serum alanine aminotransferase and triglycerides analysis in our in vitro macrophage activation and in vivo murine ethanol feeding studies. Ethanol significantly increased EV release by 3.3-fold from HepG2(Cyp2E1) cells and was associated with activation of caspase-3. Blockade of caspase activation with pharmacological or genetic approaches abrogated alcohol-induced EV release. EV stimulated macrophage activation and inflammatory cytokine induction. An unbiased microarray-based approach and antibody neutralization experiments demonstrated a critical role of CD40 ligand (CD40L) in EV mediated macrophage activation. In vivo, wild-type mice receiving a pan-caspase, Rho kinase inhibitor or with genetic deletion of CD40 (CD40(-/-)) or the caspase-activating TRAIL receptor (TR(-/-)), were protected from alcohol-induced injury and associated macrophage infiltration. Moreover, serum from patients with alcoholic hepatitis showed increased levels of CD40L enriched EV. In conclusion, hepatocytes release CD40L containing EV in a caspase-dependent manner in response to alcohol exposure which promotes macrophage activation, contributing to inflammation in ALD.

PPE26 induces TLR2-dependent activation of macrophages and drives Th1-type T-cell immunity by triggering the cross-talk of multiple pathways involved in the host response.

The pathophysiological functions and the underlying molecular basis of PE /PPE proteins of M. tuberculosis remain largely unknown. In this study, we focused on the link between PPE26 and host response. We demonstrated that PPE26 can induce extensive inflammatory responses in macrophages through triggering the cross-talk of multiple pathways involved in the host response, as revealed by iTRAQ-based subcellular quantitative proteomics. We observed that PPE26 is able to specifically bind to TLR2 leading to the subsequent activation of MAPKs and NF-κB signaling. PPE26 functionally stimulates macrophage activation by augmenting pro-inflammatory cytokine production (TNF-α, IL-6 and IL-12 p40) and the expression of cell surface markers (CD80, CD86, MHC class I and II). We observed that PPE26-treated macrophages effectively polarizes naïve CD4(+) T cells to up-regulate CXCR3 expression, and to secrete IFN-γ and IL-2, indicating PPE26 contributes to the Th1 polarization during the immune response. Importantly, rBCG::PPE26 induces stronger antigen-specific TNF-α and IFN-γ activity, and higher levels of the Th1 cytokines TNF-α and IFN-γ comparable to BCG. Moreover, PPE26 effectively induces the reciprocal expansion of effector/memory CD4(+)/CD8(+) CD44(high)CD62L(low) T cells in the spleens of mice immunized with this strain. These results suggest that PPE26 may be a TLR2 agonist that stimulates innate immunity and adaptive immunity, indicating that PPE26 is a potential antigen for the rational design of an efficient vaccine against M. tuberculosis.

Characterization of polysaccharide from longan pulp as the macrophage stimulator

A 44.7-kDa polysaccharide LPIIa from longan pulp was mainly composed of →6)-Glc-(1→, →5)-Ara-(1→, →4)-Man-(1→ and →6)-Gal-(1→. It stimulated macrophage activation partlyviaTLR4 and TLR2, followed by p38 MAPK- and NF-κB-dependent signaling pathways.

Modeling early events in Francisella tularensis pathogenesis.

Computational models can provide valuable insights into the mechanisms of infection and be used as investigative tools to support development of medical treatments. We develop a stochastic, within-host, computational model of the infection process in the BALB/c mouse, following inhalational exposure to Francisella tularensis SCHU S4. The model is mechanistic and governed by a small number of experimentally verifiable parameters. Given an initial dose, the model generates bacterial load profiles corresponding to those produced experimentally, with a doubling time of approximately 5 h during the first 48 h of infection. Analytical approximations for the mean number of bacteria in phagosomes and cytosols for the first 24 h post-infection are derived and used to verify the stochastic model. In our description of the dynamics of macrophage infection, the number of bacteria released per rupturing macrophage is a geometrically-distributed random variable. When combined with doubling time, this provides a distribution for the time taken for infected macrophages to rupture and release their intracellular bacteria. The mean and variance of these distributions are determined by model parameters with a precise biological interpretation, providing new mechanistic insights into the determinants of immune and bacterial kinetics. Insights into the dynamics of macrophage suppression and activation gained by the model can be used to explore the potential benefits of interventions that stimulate macrophage activation.