RUNX3 Regulates Intercellular Adhesion Molecule 3 (ICAM-3) Expression during Macrophage Differentiation and Monocyte Extravasation

The adhesion molecule ICAM-3 belongs to the immunoglobulin gene superfamily and functions as a ligand for the β2 integrins LFA-1, Mac-1 and αdβ2. The expression of ICAM-3 is restricted to cells of the hematopoietic lineage. We present evidences that the ICAM-3 gene promoter exhibits a leukocyte-specific activity, as its activity is significantly higher in ICAM-3+ hematopoietic cell lines. The activity of the ICAM-3 gene promoter is dependent on the occupancy of RUNX cognate sequences both in vitro and in vivo, and whose integrity is required for RUNX responsiveness and for the cooperative actions of RUNX with transcription factors of the Ets and C/EBP families. Protein analysis revealed that ICAM-3 levels diminish upon monocyte-derived macrophage differentiation, monocyte transendothelial migration and dendritic cell maturation, changes that correlate with an increase in RUNX3. Importantly, disruption of RUNX-binding sites led to enhanced promoter activity, and small interfering RNA-mediated reduction of RUNX3 expression resulted in increased ICAM-3 mRNA levels. Altogether these results indicate that the ICAM-3 gene promoter is negatively regulated by RUNX transcription factors, which contribute to the leukocyte-restricted and the regulated expression of ICAM-3 during monocyte-to-macrophage differentiation and monocyte extravasation

Expression of DC-SIGN and DC-SIGNR on human sinusoidal endothelium: a role for capturing hepatitis C virus particles.

Hepatic sinusoidal endothelial cells are unique among endothelial cells in their ability to internalize and process a diverse range of antigens. DC-SIGNR, a type 2 C-type lectin expressed on liver sinusoids, has been shown to bind with high affinity to hepatitis C virus (HCV) E2 glycoprotein. DC-SIGN is a closely related homologue reported to be expressed only on dendritic cells and a subset of macrophages and has similar binding affinity to HCV E2 glycoprotein. These receptors function as adhesion and antigen presentation molecules. We report distinct patterns of DC-SIGNR and DC-SIGN expression in human liver tissue and show for the first time that both C-type lectins are expressed on sinusoidal endothelial cells. We confirmed that these receptors are functional by demonstrating their ability to bind HCV E2 glycoproteins. Although these lectins on primary sinusoidal cells support HCV E2 binding, they are unable to support HCV entry. These data support a model where DC-SIGN and DC-SIGNR on sinusoidal endothelium provide a mechanism for high affinity binding of circulating HCV within the liver sinusoids allowing subsequent transfer of the virus to underlying hepatocytes, in a manner analogous to DC-SIGN presentation of human immunodeficiency virus on dendritic cells

Folate‐targeted nanoparticles show efficacy in the treatment of inflammatory arthritis

Objective To investigate the uptake of a poly(amidoamine) dendrimer (generation 5 [G5]) nanoparticle covalently conjugated to polyvalent folic acid (FA) as the targeting ligand into macrophages, and to investigate the activity of an FA‐ and methotrexate (MTX)–conjugated dendrimer (G5‐FA‐MTX) as a therapeutic for the inflammatory disease of arthritis. Methods In vitro studies were performed in macrophage cell lines and in isolated mouse macrophages to check the cellular uptake of fluorescence‐tagged G5‐FA nanoparticles, using flow cytometry and confocal microscopy. In vivo studies were conducted in a rat model of collagen‐induced arthritis to evaluate the therapeutic potential of G5‐FA‐MTX. Results Folate‐targeted dendrimer bound and internalized in a receptor‐specific manner into both folate receptor β–expressing macrophage cell lines and primary mouse macrophages. The conjugate G5‐FA‐MTX acted as a potent antiinflammatory agent and reduced arthritis‐induced parameters of inflammation such as ankle swelling, paw volume, cartilage damage, bone resorption, and body weight decrease. Conclusion The use of folate‐targeted nanoparticles to specifically target MTX into macrophages may provide an effective clinical approach for antiinflammatory therapy in rheumatoid arthritis.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/86938/1/30459_ftp.pd

Role of the Mannose Receptor (CD206) in Innate Immunity to Ricin Toxin

The entry of ricin toxin into macrophages and certain other cell types in the spleen and liver results in toxin-induced inflammation, tissue damage and organ failure. It has been proposed that uptake of ricin into macrophages is facilitated by the mannose receptor (MR; CD206), a C-type lectin known to recognize the oligosaccharide side chains on ricin’s A (RTA) and B (RTB) subunits. In this study, we confirmed that the MR does indeed promote ricin binding, uptake and killing of monocytes in vitro. To assess the role of MR in the pathogenesis of ricin in vivo, MR knockout (MR−/−) mice were challenged with the equivalent of 2.5× or 5× LD50 of ricin by intraperitoneal injection. We found that MR−/− mice were significantly more susceptible to toxin-induced death than their age-matched, wild-type control counterparts. These data are consistent with a role for the MR in scavenging and degradation of ricin, not facilitating its uptake and toxicity in vivo

Primary Hyperparathyroidism Influences the Expression of Inflammatory and Metabolic Genes in Adipose Tissue

Background: Primary hyperparathyroidism (PHPT) is characterised by increased production of parathyroid hormone (PTH) resulting in elevated serum calcium levels. The influence on bone metabolism with altered bone resorption is the most studied clinical condition in PHPT. In addition to this, patients with PHPT are at increased risk of non-skeletal diseases, such as impaired insulin sensitivity, arterial hypertension and increased risk of death by cardiovascular diseases (CVD), possibly mediated by a chronic low-grade inflammation. The aim of this study was to investigate whether adipose tissue reflects the low-grade inflammation observed in PHPT patients. Methodology/Principal Findings: Subcutaneous fat tissue from the neck was sampled from 16 non-obese patients with PHPT and from 16 patients operated for benign thyroid diseases, serving as weight-matched controls. RNA was extracted and global gene expression was analysed with Illumina BeadArray Technology. We found 608 differentially expressed genes (q-value,0.05), of which 347 were up-regulated and 261 were down-regulated. Gene ontology analysis showed that PHPT patients expressed increased levels of genes involved in immunity and defense (e.g. matrix metallopeptidase 9, S100 calcium binding protein A8 and A9, CD14, folate receptor 2), and reduced levels of genes involved in metabolic processes. Analysis of transcription factor binding sites present in the differentially expressed genes corroborated the up-regulation of inflammatory processes. Conclusions/Significance: Our findings demonstrate that PHPT strongly influences gene regulation in fat tissue, which may result in altered adipose tissue function and release of pathogenic factors that increase the risk of CVD

Supernatant from Bifidobacterium Differentially Modulates Transduction Signaling Pathways for Biological Functions of Human Dendritic Cells

International audienceBACKGROUND:Probiotic bacteria have been shown to modulate immune responses and could have therapeutic effects in allergic and inflammatory disorders. However, the signaling pathways engaged by probiotics are poorly understood. We have previously reported that a fermentation product from Bifidobacterium breve C50 (BbC50sn) could induce maturation, high IL-10 production and prolonged survival of DCs via a TLR2 pathway. We therefore studied the roles of mitogen-activated protein kinases (MAPK), glycogen synthase kinase-3 (GSK3) and phosphatidylinositol 3-kinase (PI3K) pathways on biological functions of human monocyte-derived DCs treated with BbC50sn.METHODOLOGY/PRINCIPAL FINDINGS:DCs were differentiated from human monocytes with IL-4 and GM-CSF for 5 days and cultured with BbC50sn, lipopolysaccharide (LPS) or Zymosan, with or without specific inhibitors of p38MAPK (SB203580), ERK (PD98059), PI3K (LY294002) and GSK3 (SB216763). We found that 1) the PI3K pathway was positively involved in the prolonged DC survival induced by BbC50sn, LPS and Zymosan in contrast to p38MAPK and GSK3 which negatively regulated DC survival; 2) p38MAPK and PI3K were positively involved in DC maturation, in contrast to ERK and GSK3 which negatively regulated DC maturation; 3) ERK and PI3K were positively involved in DC-IL-10 production, in contrast to GSK3 that was positively involved in DC-IL-12 production whereas p38MAPK was positively involved in both; 4) BbC50sn induced a PI3K/Akt phosphorylation similar to Zymosan and a p38MAPK phosphorylation similar to LPS.CONCLUSION/SIGNIFICANCE:We report for the first time that a fermentation product of a bifidobacteria can differentially activate MAPK, GSK3 and PI3K in order to modulate DC biological functions. These results give new insights on the fine-tuned balance between the maintenance of normal mucosal homeostasis to commensal and probiotic bacteria and the specific inflammatory immune responses to pathogen bacteria

Differential Macrophage Polarization Promotes Tissue Remodeling and Repair in a Model of Ischemic Retinopathy

Diabetic retinopathy is the leading cause of visual loss in individuals under the age of 55. Umbilical cord blood (UCB)–derived myeloid progenitor cells have been shown to decrease neuronal damage associated with ischemia in the central nervous system. In this study we show that UCB-derived CD14+ progenitor cells provide rescue effects in a mouse model of ischemic retinopathy by promoting physiological angiogenesis and reducing associated inflammation. We use confocal microscopy to trace the fate of injected human UCB-derived CD14+ cells and PCR with species-specific probes to investigate their gene expression profile before and after injection. Metabolomic analysis measures changes induced by CD14+ cells. Our results demonstrate that human cells differentiate in vivo into M2 macrophages and induce the polarization of resident M2 macrophages. This leads to stabilization of the ischemia-injured retinal vasculature by modulating the inflammatory response, reducing oxidative stress and apoptosis and promoting tissue repair

Generation of Novel Bone Forming Cells (Monoosteophils) from the Cathelicidin-Derived Peptide LL-37 Treated Monocytes

Bone generation and maintenance involve osteoblasts, osteoclasts, and osteocytes which originate from unique precursors and rely on key growth factors for differentiation. However, an incomplete understanding of bone forming cells during wound healing has led to an unfilled clinical need such as nonunion of bone fractures. Since circulating monocytes are often recruited to sites of injury and may differentiate into various cell types including osteoclasts, we investigated the possibility that circulating monocytes in the context of tissue injury may also contribute to bone repair. In particular, we hypothesized that LL-37 (produced from hCAP-18, cathelicidin), which recruits circulating monocytes during injury, may play a role in bone repair.Treatment of monocytes from blood with LL-37 for 6 days resulted in their differentiation to large adherent cells. Growth of LL-37-differentiated monocytes on osteologic discs reveals bone-like nodule formation by scanning electron microscopy (SEM). In vivo transplantation studies in NOD/SCID mice show that LL-37-differentiated monocytes form bone-like structures similar to endochondral bone formation. Importantly, LL-37-differentiated monocytes are distinct from conventional monocyte-derived osteoclasts, macrophages, and dendritic cells and do not express markers of the mesenchymal stem cells (MSC) lineage, distinguishing them from the conventional precursors of osteoblasts. Furthermore, LL-37 differentiated monocytes express intracellular proteins of both the osteoblast and osteoclast lineage including osteocalcin (OC), osteonectin (ON), bone sialoprotein II (BSP II), osteopontin (OP), RANK, RANKL, MMP-9, tartrate resistant acid phosphatase (TRAP), and cathepsin K (CK).Blood derived monocytes treated with LL-37 can be differentiated into a novel bone forming cell that functions both in vitro and in vivo. We propose the name monoosteophil to indicate their monocyte derived lineage and their bone forming phenotype. These cells may have wide ranging implications in the clinic including repair of broken bones and treatment of osteoporosis