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Biomedical subjects

Alberto Mantovani

Publications and source records attributed to Alberto Mantovani.

At least 109 records · Page 6Linked to original sources
PubMed2003

Characterization of the long pentraxin PTX3 as a TNFalpha-induced secreted protein of adipose cells.

Exposure of preadipocytes to long-chain fatty acids induces the expression of several markers of adipocyte differentiation. In an attempt to identify novel genes and proteins that are regulated by fatty acids in preadipocytes, we performed a substractive hybridization screening and identified PTX3, a protein of the pentraxin family. PTX3 mRNA expression is transient during adipocyte differentiation of clonal cell lines and is absent in fully differentiated cells. Stable overexpression of PTX3 in preadipocytes has no effect on adipocyte differentiation. In line with this, PTX3 mRNA is expressed in the stromal-vascular fraction of adipose tissue, but not in the adipocyte fraction; however, in 3T3-F442A adipocytes, the PTX3 gene can be reinduced by tumor necrosis factor alpha (TNFalpha) in a dose-dependent manner. This effect is accompanied by PTX3 protein secretion from both 3T3-F442A adipocytes and explants of mouse adipose tissue. PTX3 mRNA levels are found to be higher in adipose tissue of genetically obese mice versus control mice, consistent with their increased TNFalpha levels. In conclusion, PTX3 appears as a TNFalpha-induced protein that provides a new link between chronic low-level inflammatory state and obesity.

3T3 Cells↗
PubMed2003

Cutting edge: scavenging of inflammatory CC chemokines by the promiscuous putatively silent chemokine receptor D6.

In an effort to define the actual function of the promiscuous putatively silent chemokine receptor D6, transfectants were generated in different cell types. Engagement of D6 by inflammatory CC chemokines elicited no calcium response nor chemotaxis, but resulted in efficient agonist internalization and degradation. Also in lymphatic endothelium, where this receptor is expressed in vivo, D6 did not elicit cellular responses other than ligand internalization and degradation. In particular, no evidence was obtained for D6-mediated transcytosis of chemokines in the apical-to-basal or basal-to-apical directions. These results indicate that D6 acts as an inflammatory chemokine scavenging nonactivatory decoy receptors and suggest that in lymphatic vessels D6 may function as a gatekeeper for inflammatory CC chemokines, by clearing them and preventing excessive diffusion via afferent lymphatics to lymph nodes.

Animals↗
PubMed2003

The long pentraxin PTX3 is synthesized in IgA glomerulonephritis and activates mesangial cells.

The long pentraxin PTX3 has been recently involved in amplification of the inflammatory reactions and regulation of innate immunity. In the present study we evaluated the expression and role of PTX3 in glomerular inflammation. PTX3 expression was investigated in the IgA, type I membranoproliferative, and diffuse proliferative lupus glomerulonephritis, which are characterized by inflammatory and proliferative lesions mainly driven by resident mesangial cells, and in the membranous glomerulonephritis and the focal segmental glomerular sclerosis, where signs of glomerular inflammation are usually absent. We found an intense staining for PTX3 in the expanded mesangial areas of renal biopsies obtained from patients with IgA glomerulonephritis. The pattern of staining was on glomerular mesangial and endothelial cells. Scattered PTX3-positive cells were also detected in glomeruli of type I membranoproliferative glomerulonephritis. The concomitant expression of CD14 suggests an inflammatory origin of these cells. Normal renal tissue and biopsies from patients with the other glomerular nephropathies studied were mainly negative for PTX3 expression in glomeruli. However, PTX3-positive cells were detected in the interstitium of nephropathies showing inflammatory interstitial injury. In vitro, cultured human mesangial cells synthesized PTX3 when stimulated with TNF-alpha and IgA and exhibited specific binding for recombinant PTX3. Moreover, stimulation with exogenous PTX3 promoted mesangial cell contraction and synthesis of the proinflammatory lipid mediator platelet-activating factor. In conclusion, we provide the first evidence that mesangial cells may both produce and be a target for PTX3. The detection of this long pentraxin in the renal tissue of patients with glomerulonephritis suggests its potential role in the modulation of glomerular and tubular injury.

Animals↗
PubMed2003

Role of the MyD88 transduction signaling pathway in endothelial activation by antiphospholipid antibodies.

Antiphospholipid syndrome (APS) is an autoimmune disease characterized by the persistent presence of antiphospholipid antibodies (aPLs) and recurrent thrombosis or fetal loss. The thrombophilic state has been partially related to the induction of a proinflammatory and procoagulant endothelial cell (EC) phenotype induced by anti-beta(2)-glycoprotein I (beta(2)-GPI) antibodies that bind beta(2)-GPI expressed on the EC surface. Anti-beta(2)-GPI antibody binding has been shown to induce nuclear factor-kappa B (NF-kappa B) translocation leading to a proinflammatory EC phenotype similar to that elicited by interaction with microbial products (lipopolysaccharide [LPS]) and proinflammatory cytokines (interleukin 1 beta [IL-1 beta], tumor necrosis factor alpha [TNF-alpha]). However, the upstream signaling events are not characterized yet. To investigate the endothelial signaling cascade activated by anti-beta(2)-GPI antibodies, we transiently cotransfected immortalized human microvascular endothelial cells (HMEC-1) with dominant-negative constructs of different components of the pathway (Delta TRAF2, Delta TRAF6, Delta MyD88) together with reporter genes (NF-kappa B luciferase and pCMV-beta-galactosidase). Results showed that both human anti-beta(2)-GPI IgM monoclonal antibodies as well as polyclonal affinity-purified anti-beta(2)-GPI IgG display a signaling cascade comparable to that activated by LPS or IL-1. Delta TRAF6 and Delta MyD88 significantly abrogate antibody-induced as well as IL-1- or LPS-induced NF-kappa B activation, whereas Delta TRAF2 (involved in NF-kappa B activation by TNF) does not affect it. Moreover, anti- beta(2)-GPI antibodies and LPS followed the same time kinetic of IL-1 receptor-activated kinase (IRAK) phosphorylation, suggesting an involvement of the toll-like receptor (TLR) family. Our findings demonstrate that anti-beta(2)-GPI antibodies react with their antigen likely associated to a member of the TLR/IL-1 receptor family on the EC surface and directly induce activation.

Adaptor Proteins, Signal Transducing↗
PubMed2003

Production of the soluble pattern recognition receptor PTX3 by myeloid, but not plasmacytoid, dendritic cells.

PTX3 is a prototypic of long pentraxin consisting of an N-terminal portion coupled to a C-terminal pentraxin domain, the latter related to short pentraxins (C-reactive protein and serum amyloid P component). PTX3 is a soluble pattern recognition receptor, which plays a non-redundant role in resistance against selected pathogens and in female fertility. The present study was designed to analyze the production of PTX3 by human dendritic cells (DC) and to define the role of different innate immunity receptors in its induction. Human monocyte-derived DC produced copious amounts of PTX3in response to microbial ligands engaging different members of the Toll-like receptor (TLR) family (TLR1 through TLR6), whereas engagement of the mannose receptor had no substantial effect. DC werebetter producers of PTX3 than monocytes and macrophages. Freshly isolated peripheral blood myeloid DC produced PTX3 in response to diverse microbial stimuli. In contrast, plasmacytoid DC exposed to influenza virus or to CpG oligodeoxynucleotides engaging TLR9, did not produce PTX3. PTX3-expressing DC were present in inflammatory lymph nodes from HIV-infected patients. These results suggest that DC of myelomonocytic origin are a major source of PTX3, a molecule which facilitates pathogen recognition and subsequent activation of innate and adaptive immunity.

C-Reactive Protein↗
PubMed2003

Biochemical and functional characterization of the interaction between pentraxin 3 and C1q.

Pentraxin 3 (PTX3) is a recently characterized member of the pentraxin family of acute-phase proteins produced during inflammation. Classical short pentraxins, C-reactive protein, and serum amyloid P component can bind to C1q and thereby activate the classical complement pathway. Since PTX3 can also bind C1q, the present study was designed to define the interaction between PTX3 and C1q and to examine the functional consequences of this interaction. A dose-dependent binding of both C1q and the C1 complex to PTX3 was observed. Experiments with recombinant globular head domains of human C1q A, B, and C chains indicated that C1q interacts with PTX3 via its globular head region. Binding of C1q to immobilized PTX3 induced activation of the classical complement pathway as assessed by C4 deposition. Furthermore, PTX3 enhanced C1q binding and complement activation on apoptotic cells. However, in the fluid-phase, pre-incubation of PTX3 with C1q resulted in inhibition of complement activation by blocking the interaction of C1q with immunoglobulins. These results indicate that PTX3 can both inhibit and activate the classical complement pathway by binding C1q, depending on the way it is presented. PTX3 may therefore be involved in the regulation of the innate immune response.

Apoptosis↗
PubMed2003

Activation of signal transducer and activator of transcription 3 in rat liver after heat shock and reperfusion stress.

Changes in transcription factors (TFs) accompany many types of cell stresses. By using electrophoretic mobility assays we show that the DNA binding of signal transducer and activator of transcription 3 (STAT3) is activated in rat liver by heat shock and ischemia-reperfusion. Northern blot and Western blot analysis reveal an increase of the mRNA and protein level of this transcription factor. Under both conditions the phosphorylation of pre-existing STAT3 is prompt and precedes the increase in the STAT3 protein. The activation: (1) is functional, i.e. is followed by the transcription of the target gene alpha(1)-acid glycoprotein (2) is strongly inhibited by pretreatment with the interleukin-1 receptor antagonist before heat shock but only slightly by pretreatment before ischemia-reperfusion (3) might, at least in part, be mediated by a cytokine cascade involving also interleukin-6. These results are consistent with the hypothesis that different kinds of stress can activate a number of common TFs.

Animals↗
PubMed2003

IL-6: a regulator of the transition from neutrophil to monocyte recruitment during inflammation.

The origin of the Toll-like family of receptors pre-dates the evolutionary split between the plant and animal kingdoms. These receptors are remarkably conserved across the taxonomic kingdoms and have fundamental roles in triggering immune responses. How they trigger such responses, and how these mechanisms arose in evolution, is a topic of extensive debate. We postulate a surveillance model: these receptors "keep watch" for both endogenous and exogenous molecules that indicate tissue inquiry, infection and remodeling. Furthermore, we suggest that the first Toll-like family receptors that arose in evolution might have acted in both development and immunity by recognizing the degradation of endogenous macromolecules.

Animals↗
PubMed2003

Tuning of innate immunity and polarized responses by decoy receptors.

After the identification of the interleukin (IL)-1 type II receptor as the prototype, decoy receptors have been identified for a number of members of the IL-1/IL-18, TNF, IL-10 and IL-13 receptor families. Moreover, the silent receptor D6 is a promiscuous decoy and scavenger receptor of inflammatory chemokines. The IL-1 decoy receptor is regulated by pro- and anti-inflammatory signals and its levels may serve as a readout of the activation of anti-inflammatory pathways, for instance by glucocorticoid hormones. Decoy receptors represent a strategy to tune inflammatory and polarized adaptive responses.

Cytokines↗
PubMed2002

Hazard identification and risk assessment of endocrine disrupting chemicals with regard to developmental effects.

Endocrine Disrupting Chemicals (EDC) may affect mammalian development either indirectly (by affecting implantation, placenta, lactation, etc.) or directly, altering the maturation of target tissues. Studies on reproductive and developmental effects are likely to provide the critical information for risk assessment of most EDC, when proper endpoints are investigated. Toxicity testing guidelines, and especially the two-generation test, are currently being updated with regard to risk assessment of EDC. Particular emphasis is put on selection and validation of the parameters most sensitive to given chemicals and/or mechanisms of action; therefore, a screening battery it is important to address further testing. Under this respect the future use of gene arrays might be a most promising development, when more data will be available. Further points to be considered in a biologically-based assessment of developmental risks include: the identification of critical effects also through the evaluation of dose-response curves; the assessment of the actual exposure and different susceptibility of target tissues; the identification of possible factors of susceptibility and gene-environment interactions; the elaboration of models to estimate the possible additional risk deriving from combined exposures to EDC having similar mechanisms and/or targets.

Animals↗
PubMed2002

Non-redundant role of the long pentraxin PTX3 in anti-fungal innate immune response.

Pentraxins are a superfamily of conserved proteins that are characterized by a cyclic multimeric structure. The classical short pentraxins, C-reactive protein (CRP) and serum amyloid P component (SAP), are acute-phase proteins produced in the liver in response to inflammatory mediators. Short pentraxins regulate innate resistance to microbes and the scavenging of cellular debris and extracellular matrix components. In contrast, long pentraxins have an unrelated, long amino-terminal domain coupled to the carboxy-terminal pentraxin domain, and differ, with respect to short pentraxins, in their gene organization, chromosomal localization, cellular source, and in their stimuli-inducing and ligand-recognition ability. To investigate the in vivo function of the long pentraxin PTX3, we generated mice deficient in Ptx3 by homologous recombination. Ptx3-null mice were susceptible to invasive pulmonary aspergillosis. Ptx3 binds selected microbial agents, including conidia of Aspergillus fumigatus, and we found that susceptibility of Ptx3-null mice was associated with defective recognition of conidia by alveolar macrophages and dendritic cells, as well as inappropriate induction of an adaptive type 2 response. Thus, the long pentraxin Ptx3 is a secreted pattern-recognition receptor that has a non-redundant role in resistance to selected microbial agents, in particular to the opportunistic fungal pathogen Aspergillus fumigatus.

Animals↗
PubMed2002

Long pentraxin PTX3 upregulates tissue factor expression in human endothelial cells: a novel link between vascular inflammation and clotting activation.

Inflammation is a major contributing factor to atherosclerotic plaque development and ischemic heart disease. PTX3 is a long pentraxin that was recently found to be increased in patients with acute myocardial infarction. Because tissue factor (TF), the in vivo trigger of blood coagulation, plays a dominant role in thrombus formation after plaque rupture, we tested the possibility that PTX3 could modulate TF expression. Human umbilical vein endothelial cells, incubated with endotoxin (lipopolysaccharide) or the inflammatory cytokines interleukin-1beta and tumor necrosis factor-alpha, expressed TF. The presence of PTX3 increased TF activity and antigen severalfold in a dose-dependent fashion. PTX3 exerted its effect at the transcription level, inasmuch as the increased levels of TF mRNA, mediated by the stimuli, were enhanced in its presence. The increase in mRNA determined by PTX3 originated from an enhanced nuclear binding activity of the transacting factor c-Rel/p65, which was mediated by the agonists and measured by electrophoretic mobility shift assay. The mechanism underlying the increased c-Rel/p65 activity resided in an enhanced degradation of the c-Rel/p65 inhibitory protein IkappaBalpha. In the area of vascular injury, during the inflammatory response, cell-mediated fibrin deposition takes place. Our results suggest that PTX3, by increasing TF expression, potentially plays a role in thrombogenesis and ischemic vascular disease.

Acute-Phase Proteins↗
PubMed2002

Production of the long pentraxin PTX3 in advanced atherosclerotic plaques.

Elevated plasma levels of the pentraxin protein family member C-reactive protein (CRP) are associated with increased risk of cardiovascular disease in both healthy and high-risk subjects. The long pentraxin family member, pentraxin 3 (PTX3), was recently described. Like CRP, PTX3 is induced by acute inflammatory stimuli and is increased in the blood of patients with acute myocardial infarction. Unlike CRP, it is expressed in a wide range of cell types, but not in hepatocytes. In this study, we have investigated the expression of PTX3 in atherosclerosis. Immunohistochemical staining of advanced atherosclerotic lesions revealed strong expression of PTX3. In contrast, no PTX3 expression was observed in nonatherosclerotic internal mammary arteries. By staining serial sections with cell type- and PTX3-specific antibodies, we observed that PTX3 was produced principally by macrophages and endothelial cells. Infrequent expression by smooth muscle cells was also observed. Our results suggest that PTX3 may contribute to the pathogenesis of atherosclerosis.

Acute-Phase Proteins↗
PubMed2002

Stimulation of toll-like receptor 4 expression in human mononuclear phagocytes by interferon-gamma: a molecular basis for priming and synergism with bacterial lipopolysaccharide.

In human monocytes and macrophages, interferon-gamma (IFNgamma) augmented mRNA and surface expression of toll-like receptor 4 (TLR4), a crucial component of the signaling receptor complex for bacterial lipopolysaccharide (LPS). Expression of the accessory component MD-2 and of the adapter protein MyD88 was also increased. LPS increased TLR4 mRNA levels, but concomitantly decreased its surface expression. IFNgamma counteracted the LPS-induced downregulation of TLR4. IFNgamma-primed monocytes showed increased responsiveness to LPS in terms of phosphorylation of the interleukin-1 receptor-associated kinase (IRAK; immediately downstream of the MyD88 adapter protein), NF-kB DNA binding activity, and, accordingly, of cytokine (tumor necrosis factor alpha [TNFalpha] and interleukin-12 [IL-12]) production. These results suggest that enhanced TLR4 expression underlies the long-known priming by IFNgamma of mononuclear phagocytes for pathogen recognition and killing as well as its synergism with LPS in macrophage activation.

Antigens, Surface↗
PubMed2002

Neutrophils are indispensable for hematopoietic stem cell mobilization induced by interleukin-8 in mice.

The CXC chemokine interleukin-8 (IL-8/CXCL8) induces rapid mobilization of hematopoietic progenitor cells (HPCs). Previously we showed that mobilization could be prevented completely in mice by pretreatment with neutralizing antibodies against the beta2-integrin LFA-1 (CD11a). In addition, murine HPCs do not express LFA-1, indicating that mobilization requires a population of accessory cells. Here we show that polymorphonuclear cells (PMNs) serve as key regulators in IL-8-induced HPC mobilization. The role of PMNs was studied in mice rendered neutropenic by administration of a single injection of antineutrophil antibodies. Absolute neutropenia was observed up to 3-5 days with a rebound neutrophilia at day 7. The IL-8-induced mobilizing capacity was reduced significantly during the neutropenic phase, reappeared with recurrence of the PMNs, and was increased proportionally during the neutrophilic phase. In neutropenic mice, the IL-8-induced mobilizing capacity was restored by the infusion of purified PMNs but not by infusion of mononuclear cells. Circulating metalloproteinase gelatinase B (MMP-9) levels were detectable only in neutropenic animals treated with PMNs in combination with IL-8, showing that in vivo activated PMNs are required for the restoration of mobilization. However, IL-8-induced mobilization was not affected in MMP-9-deficient mice, indicating that MMP-9 is not indispensable for mobilization. These data demonstrate that IL-8-induced mobilization of HPCs requires the in vivo activation of circulating PMNs.

Animals↗
PubMed2002

Identification of biologically active chemokine isoforms from ascitic fluid and elevated levels of CCL18/pulmonary and activation-regulated chemokine in ovarian carcinoma.

Chemokines are important in leukocyte homeostasis, inflammation, angiogenesis, and metastasis. Here, the molecular diversity of chemokines present in ovarian carcinoma was studied by purifying the proteins to homogeneity from ascitic fluid. Biologically active intact CCL2 and processed CXCL8, CCL3, and CCL18 isoforms were recovered. CCL7 and CCL20 were also purified, but their levels were 10-fold lower compared with CXCL8, CCL2, and CCL3 and even 100-fold lower than the amounts of CCL18 isolated. In ascitic fluids from patients with ovarian carcinoma (n = 12), significantly higher levels of CXCL8 and CCL18 (2.0 versus 0.7 ng/ml (p = 0.01) and 120 versus 44 ng/ml (p = 0.0002), respectively) were detected compared with those in nonovarian carcinoma patients (n = 12). In contrast to CXCL8, CCL18 was not inducible in carcinoma cell lines. Immunostaining showed CCL18 expression in tumor-infiltrating cells with monocyte/macrophage morphology but not in the ovarian carcinoma cells. Our data demonstrate that biochemically heterogenous but biologically active forms of several chemokines are present at different concentrations in ovarian carcinoma ascitic fluid. This points to a delicate balance of chemokines in epithelial ovarian cancer and to a potentially major role for CXCL8 and CCL18 in this tumor.

Ascites↗