Biomedical subjects

Alberto Mantovani

Publications and source records attributed to Alberto Mantovani.

At least 73 records · Page 4Linked to original sources

PubMed2004

Increased survival, proliferation, and migration in metastatic human pancreatic tumor cells expressing functional CXCR4.

In this study, we have evaluated 11 pancreatic tumor cell lines and tumor cells from surgical samples of patients with pancreatic adenocarcinoma for expression of the chemokine receptor CXCR4. Six of 11 cell lines expressed detectable mRNA of CXCR4, with three cell lines (AsPC1, Capan1, and Hs766T) having substantial amounts of transcripts. Expression was higher in lines derived from metastatic lesions compared with those derived from primary tumors. Different inflammatory cytokines did not modify expression, whereas IFN-gamma down-regulated and hypoxia up-regulated CXCR4 transcripts. Transcript expression was associated with surface expression in pancreatic carcinoma cell lines. All surgical carcinoma samples tested expressed higher levels of CXCR4 than normal pancreatic ducts, which were used as reference tissue. The chemokine CXCL12 induced chemotaxis in CXCR4-positive pancreatic carcinoma cell lines, which was inhibited by anti-CXCR4 monoclonal antibody and by the antagonist AMD3100. Transendothelial migration, Matrigel invasion, and activation of matrix metalloproteases were also enhanced by CXCL12. In CXCR4-positive cell lines, CXCL12 stimulated cell proliferation. The cell line Hs766T produces high levels of CXCL12, and addition of the CXCR4 antagonist AMD3100 partially inhibited proliferation, indicating an autocrine loop. Moreover, the addition of exogenous CXCL12 inhibited apoptosis induced by serum starvation. These results indicate that the CXCR4 receptor is frequently expressed in metastatic pancreatic tumor cells. CXCR4 not only stimulates cell motility and invasion but also promotes survival and proliferation. Strategies to target CXCR4 expressed on tumor cells may be of benefit in patients with pancreatic cancer.

Adenocarcinoma↗

PubMed2004

Gene expression profile activated by the chemokine CCL5/RANTES in human neuronal cells.

Differentiated human NT2-N neurons were shown to express CCR5 and CXCR4 chemokine receptor mRNA and protein, and to be responsive to the chemokines CCL5 and CXCL12. Using cDNA microarray technology, CCL5 was found to induce a distinct transcriptional program, with reproducible induction of 46 and 9 genes after 2 and 8 hr of treatment, respectively. Conversely, downregulation of 20 and 7 genes was observed after 2 and 8 hr of treatment, respectively. Modulation of a selected panel of CCL5-responsive genes was also confirmed by quantitative RT-PCR and Western blot and compared to gene expression changes induced by CXCL12 treatment. Gene clustering identified distinct functional subsets of CCL5-responsive molecules, and a significant number of expressed sequence tags encoding unknown genes. CCL5-responsive genes comprise a significant number of enzymes, transcription factors, and miscellaneous molecules involved in neuronal survival and differentiation, including neurite outgrowth and synaptogenesis. Our results suggest that CCL5 biological functions might go beyond its recognized chemotactic activity in the central nervous system, in particular with regard to the control of neural plasticity events both during development and in postnatal life.

Blotting, Western↗

PubMed2004

Prognostic significance of the long pentraxin PTX3 in acute myocardial infarction.

BACKGROUND: Inflammation has a pathogenetic role in acute myocardial infarction (MI). Pentraxin-3 (PTX3), a long pentraxin produced in response to inflammatory stimuli and highly expressed in the heart, was shown to peak in plasma approximately 7 hours after MI. The aim of this study was to assess the prognostic value of PTX3 in MI compared with the best-known and clinically relevant biological markers. METHODS AND RESULTS: In 724 patients with MI and ST elevation, PTX3, C-reactive protein (CRP), creatine kinase (CK), troponin T (TnT), and N-terminal pro-brain natriuretic peptide (NT-proBNP) were assayed at entry, a median of 3 hours, and the following morning, a median of 22 hours from symptom onset. With respect to outcome events occurring over 3 months after the index event, median PTX3 values were 7.08 ng/mL in event-free patients, 16.12 ng/mL in patients who died, 9.12 ng/mL in patients with nonfatal heart failure, and 6.88 ng/mL in patients with nonfatal residual ischemia (overall P<0.0001). Multivariate analysis including CRP, CK, TnT, and NT-proBNP showed that only age > or =70 years (OR, 2.11; 95% CI, 1.04 to 4.31), Killip class >1 at entry (OR, 2.20; 95% CI, 1.14 to 4.25), and PTX3 (>10.73 ng/mL) (OR, 3.55; 95% CI, 1.43 to 8.83) independently predicted 3-month mortality. Biomarkers predicting the combined end point of death and heart failure in survivors were the highest tertile of PTX3 and of NT-proBNP and a CK ratio >6. CONCLUSIONS: In a representative contemporary sample of patients with MI with ST elevation, the acute-phase protein PTX3 but not the liver-derived short pentraxin CRP or other cardiac biomarkers (NT-proBNP, TnT, CK) predicted 3-month mortality after adjustment for major risk factors and other acute-phase prognostic markers.

Aged↗

PubMed2004

Defective dendritic cell migration and activation of adaptive immunity in PI3Kgamma-deficient mice.

Gene-targeted mice were used to evaluate the role of the gamma isoform of phosphoinositide 3-kinase (PI3Kgamma) in dendritic cell (DC) migration and induction of specific T-cell-mediated immune responses. DC obtained from PI3Kgamma-/- mice showed a reduced ability to respond to chemokines in vitro and ex vivo and to travel to draining lymph nodes under inflammatory conditions. PI3Kgamma-/- mice had a selective defect in the number of skin Langerhans cells and in lymph node CD8alpha- DC. Furthermore, PI3Kgamma-/- mice showed a defective capacity to mount contact hypersensitivity and delayed-type hypersensitivity reactions. This defect was directly related to the reduced ability of antigen-loaded DC to migrate from the periphery to draining lymph nodes. Thus, PI3Kgamma plays a nonredundant role in DC trafficking and in the activation of specific immunity. Therefore, PI3Kgamma may be considered a new target to control exaggerated immune reactions.

Animals↗

PubMed2004

Noncompetitive allosteric inhibitors of the inflammatory chemokine receptors CXCR1 and CXCR2: prevention of reperfusion injury.

The chemokine CXC ligand 8 (CXCL8)/IL-8 and related agonists recruit and activate polymorphonuclear cells by binding the CXC chemokine receptor 1 (CXCR1) and CXCR2. Here we characterize the unique mode of action of a small-molecule inhibitor (Repertaxin) of CXCR1 and CXCR2. Structural and biochemical data are consistent with a noncompetitive allosteric mode of interaction between CXCR1 and Repertaxin, which, by locking CXCR1 in an inactive conformation, prevents signaling. Repertaxin is an effective inhibitor of polymorphonuclear cell recruitment in vivo and protects organs against reperfusion injury. Targeting the Repertaxin interaction site of CXCR1 represents a general strategy to modulate the activity of chemoattractant receptors.

Allosteric Regulation↗

PubMed2004

Distinct transcriptional programs activated by interleukin-10 with or without lipopolysaccharide in dendritic cells: induction of the B cell-activating chemokine, CXC chemokine ligand 13.

To understand the modulation of dendritic cell (DC) function by IL-10, gene expression profiling was performed by using Affymetrix technology (Santa Clara, CA) in human monocyte-derived DC treated with IL-10, alone or in combination with LPS. The modulation of selected genes was validated by real-time PCR, Northern blot, and protein production. IL-10 regulated in DC the expression of a limited number of genes, including IL-7, the receptors for transferrin and vitamin D(3), structural matrix proteins, and signal transduction elements. The combined treatment with LPS plus IL-10 modulated a number of genes comparable to LPS alone, but the expression profiles were distinct. As expected, IL-10 suppressed the expression of several LPS-inducible proinflammatory molecules. Among genes uniquely modulated by the concomitant treatment with LPS plus IL-10, phosphatidylinositol 3-kinase gamma was down-regulated while the suppressor of cytokine signaling 3, signaling lymphocytic activation molecule, regulator of G protein signaling 16, and the chemokine, CXC chemokine ligand (CXCL) 13, were up-regulated. Overall, four distinct transcriptional programs were identified, related to: 1) control of immunity and inflammation; 2) tuning of cytokine receptor and G protein-coupled receptor signaling; 3) remodeling of extracellular matrix; and 4) B cell function and lymphoid tissue neogenesis. Among the latter genes, we further demonstrate that IL-10 synergizes with TLR ligands for the production of functionally active B cell-attracting chemokine, CXCL13, in both myeloid and plasmacytoid DC. This novel finding reveals that IL-10 sustains humoral immunity by inducing the production in APCs of the chemokine, CXCL13, which amplifies B cell recruitment and promotes lymphoid tissue neogenesis.

Chemokine CXCL13↗

PubMed2004

Differential recognition and scavenging of native and truncated macrophage-derived chemokine (macrophage-derived chemokine/CC chemokine ligand 22) by the D6 decoy receptor.

The promiscuous D6 receptor binds several inflammatory CC chemokines and has been recently proposed to act as a chemokine-scavenging decoy receptor. The present study was designed to better characterize the spectrum of CC chemokines scavenged by D6, focusing in particular on CCR4 ligands and analyzing the influence of NH(2)-terminal processing on recognition by this promiscuous receptor. Using D6 transfectants, it was found that D6 efficiently bound and scavenged most inflammatory CC chemokines (CCR1 through CCR5 agonists). Homeostatic CC chemokines (CCR6 and CCR7 agonists) were not recognized by D6. The CCR4 agonists CC chemokine ligand 17 (CCL17) and CCL22 bound to D6 with high affinity. CCL17 and CCL22 have no agonistic activity for D6 (chemotaxis and calcium fluxes), but were rapidly scavenged, resulting in reduced chemotactic activity on CCR4 transfectants. CD26 mediates NH(2) terminus processing of CCL22, leading to the production of CCL22 (3-69) and CCL22 (5-69) that do not interact with CCR4. These NH(2)-terminal truncated forms of CCL22 were not recognized by D6. The results presented in this study show that D6 recognizes and scavenges a wide spectrum of inflammatory CC chemokines, including the CCR4 agonists CCL22 and CCL17. However, this promiscuous receptor is not engaged by CD26-processed, inactive, CCL22 variants. By recognizing intact CCL22, but not its truncated variants, D6 expressed on lymphatic endothelial cells may regulate the traffic of CCR4-expressing cells, such as dendritic cells.

Animals↗

PubMed2004

beta-Arrestin-dependent constitutive internalization of the human chemokine decoy receptor D6.

Seven transmembrane receptors mediate diverse physiological responses including hormone action, olfaction, neurotransmission, and chemotaxis. Human D6 is a non-signaling seven-transmembrane receptor expressed on lymphatic endothelium interacting with most inflammatory CC-chemokines resulting in their rapid internalization. Here, we demonstrate that this scavenging activity is mediated by continuous internalization and constant surface expression of the receptor, a process involving the clathrin-coated pit-dependent pathway. D6 constitutively associates with the cytoplasmic adaptor beta-arrestin, and this interaction is essential for D6 internalization. An acidic region, but not the putative phosphorylation sites in the cytoplasmic tail of D6, is critical for receptor interaction with beta-arrestin and subsequent internalization. Neither the native D6 nor mutants uncoupled from beta-arrestin activate any G-protein-mediated signaling pathways. Therefore, D6 may be considered a decoy receptor structurally adapted to perform chemokine scavenging.

Amino Acid Sequence↗

PubMed2004

Selective recognition of fibroblast growth factor-2 by the long pentraxin PTX3 inhibits angiogenesis.

The long pentraxin PTX3 is a soluble pattern recognition receptor produced by monocytes and endothelial cells that plays a nonredundant role in inflammation. Several pathologic conditions are characterized by local production of both PTX3 and the angiogenic fibroblast growth factor-2 (FGF2). Here, solid-phase binding assays demonstrated that PTX3 binds with high affinity to FGF2 but not to a panel of cytokines and growth factors, including FGF1, FGF4, and FGF8. Accordingly, PTX3 prevented (125)I-FGF2 binding to endothelial cell receptors, leading to specific inhibition of FGF2-induced proliferation. PTX3 hampered also the motogenic activity exerted by endogenous FGF2 on a wounded endothelial cell monolayer. Moreover, PTX3 cDNA transduction in FGF2-transformed endothelial cells inhibited their autocrine FGF2-dependent proliferation and morphogenesis in vitro and their capacity to generate vascular lesions when injected in nude mice. Finally, PTX3 suppressed neovascularization triggered by FGF2 in the chick embryo chorioallantoic membrane with no effect on physiologic angiogenesis. In contrast, the short pentraxin C-reactive protein was a poor FGF2 ligand/antagonist. These results establish the selective binding of a member of the pentraxin superfamily to a growth factor. PTX3/FGF2 interaction may modulate angiogenesis in various physiopathologic conditions driven by inflammation, innate immunity, and/or neoplastic transformation.

Angiogenesis Inhibitors↗

PubMed2004

PTX3 plays a key role in the organization of the cumulus oophorus extracellular matrix and in in vivo fertilization.

PTX3 is a prototypic long pentraxin that plays a non-redundant role in innate immunity against selected pathogens and in female fertility. Here, we report that the infertility of Ptx3(-/-) mice is associated with severe abnormalities of the cumulus oophorus and failure of in vivo, but not in vitro, oocyte fertilization. PTX3 is produced by mouse cumulus cells during cumulus expansion and localizes in the matrix. PTX3 is expressed in the human cumulus oophorus as well. Cumuli from Ptx3(-/-) mice synthesize normal amounts of hyaluronan (HA), but are unable to organize it in a stable matrix. Exogenous PTX3 restores a normal cumulus phenotype. Incorporation in the matrix of inter-alpha-trypsin inhibitor is normal in Ptx3(-/-) cumuli. PTX3 does not interact directly with HA, but it binds the cumulus matrix hyaladherin tumor necrosis factor alpha-induced protein 6 (TNFAIP6, also known as TSG6) and thereby may form multimolecular complexes that can cross-link HA chains. Thus, PTX3 is a structural constituent of the cumulus oophorus extracellular matrix essential for female fertility.

Animals↗

PubMed2004

Intestinal inflammation in mice deficient in Tir8, an inhibitory member of the IL-1 receptor family.

TIR8, also known as single Ig IL-1-related receptor, is a member of the IL-1 receptor/Toll-like receptor (TLR) superfamily, which acts as an intracellular decoy for components of the signaling pathway. Here we report that Tir8 has a unique pattern of expression, which includes mucosal tissues and dendritic cells (DC). Tir8-deficient DC showed increased cytokine production in response to TLR agonists (lipopolysaccharide, CpG oligodeoxynucleotides). Tir8-deficient mice had normal susceptibility to systemic lipopolysaccharide toxicity and to i.p. or s.c. inflammation. However, Tir8-deficient mice were more susceptible to intestinal inflammation. Thus, TIR8 represents a negative pathway of regulation of the IL-1 receptor/TLR system, expressed in epithelial cells and DC, crucial for tuning inflammation in the gastrointestinal tract.

Animals↗

PubMed2004

The contribution of the Toll-like/IL-1 receptor superfamily to innate and adaptive immunity to fungal pathogens in vivo.

In vitro studies have indicated the importance of Toll-like receptor (TLR) signaling in response to the fungal pathogens Candida albicans and Aspergillus fumigatus. However, the functional consequences of the complex interplay between fungal morphogenesis and TLR signaling in vivo remain largely undefined. In this study we evaluate the impact of the IL-1R/TLR/myeloid differentiation primary response gene 88 (MyD88)-dependent signaling pathway on the innate and adaptive Th immunities to C. albicans and A. fumigatus in vivo. It was found that 1) the MyD88-dependent pathway is required for resistance to both fungi; 2) the involvement of the MyD88 adapter may occur through signaling by distinct members of the IL-1R/TLR superfamily, including IL-1R, TLR2, TLR4, and TLR9, with the proportional role of the individual receptors varying depending on fungal species, fungal morphotypes, and route of infection; 3) individual TLRs and IL-1R activate specialized antifungal effector functions on neutrophils, which correlates with susceptibility to infection; and 4) MyD88-dependent signaling on dendritic cells is crucial for priming antifungal Th1 responses. Thus, the finding that the innate and adaptive immunities to C. albicans and A. fumigatus require the coordinated action of distinct members of the IL-1R/TLR superfamily acting through MyD88 makes TLR manipulation amenable to the induction of host resistance to fungi.

Adaptor Proteins, Signal Transducing↗

PubMed2004

The prototypic tissue pentraxin PTX3, in contrast to the short pentraxin serum amyloid P, inhibits phagocytosis of late apoptotic neutrophils by macrophages.

OBJECTIVE: Phagocytosis of apoptotic cells can be facilitated by complement components and short pentraxins, such as serum amyloid P (SAP). In contrast, the long pentraxin PTX3 was shown to inhibit phagocytosis of apoptotic Jurkat cells by dendritic cells and to bind late apoptotic polymorphonuclear leukocytes (PMNs). Recently, levels of the pentraxin PTX3 were shown to parallel disease activity in small-vessel vasculitis, which is often characterized by leukocytoclasia, a persistence of leukocyte remnants in the vessel wall. We undertook this study to test our hypothesis that PTX3 inhibits phagocytosis of late apoptotic PMNs by macrophages, thereby leading to their accumulation in the vessel wall. METHODS: Macrophages were allowed to phagocytose late apoptotic or secondary necrotic PMNs that were incubated with or without PTX3 for 30 minutes prior to phagocytosis. Phagocytosis was allowed to occur in the presence of 30% normal human serum with or without SAP and with or without depletion of complement. To discriminate between an inhibitory effect of PTX3 on binding and the internalization of apoptotic PMNs into macrophages, internalization was blocked by cytochalasin B. RESULTS: SAP and complement were both necessary for effective in vitro phagocytosis. In contrast, PTX3 inhibited phagocytosis in a dose-dependent manner, from 11% inhibition at 6.25 microg/ml to almost complete inhibition at 100 microg/ml. Furthermore, PTX3 partly affected binding of apoptotic PMNs to macrophages. CONCLUSION: PTX3, in contrast to SAP and complement, inhibits phagocytosis of late apoptotic PMNs by monocyte-derived macrophages in a dose-dependent manner. Therefore, PTX3 can play a role in the development of leukocytoclasia by affecting the clearance of apoptotic PMNs, thereby inducing their accumulation in the vessel wall.

Apoptosis↗

PubMed2004

IL-8 induces a specific transcriptional profile in human neutrophils: synergism with LPS for IL-1 production.

IL-8 is an inflammatory CXC chemokine involved in neutrophil recruitment and activation in various inflammatory conditions. The transcriptional profile induced by IL-8 in human neutrophils was analyzed using high-density oligonucleotide arrays and compared with that of the prototypic phagocyte activator LPS. As expected, LPS induced a major effect on the cell transcriptome, upregulating 116 (0.93%) and downregulating 70 (0.56%) of the transcripts. IL-8 induced a less profound modulation of the cell transcriptome, with upregulation of 30 (0.25%) and downregulation of 6 (0.04%) of the transcripts. Although the two proinflammatory mediators induced partially overlapping transcriptional profiles (50.0% of IL-8-responsive genes were concordantly regulated by LPS), IL-8 also modulated a significant number of genes unresponsive to LPS, including soluble mediators, membrane receptors, signaling molecules, and regulators of transcription and translation. A set of IL-8-inducible genes was related to cell motility, possibly a strategy to prepare for migration into tissues. Analysis of the IL-8-responsive gene IL-1beta at the protein level revealed that transcript induction was not followed by protein production. Neutrophils stimulated with IL-8, however, showed a significant increase in IL-1beta secretion after subsequent exposure to LPS. Thus, the effect of IL-8 at the transcriptional level could provide a synergistic effect with microbial products for neutrophil activation.

Gene Expression↗

PubMed2004

Modified atherogenic lipoproteins induce expression of pentraxin-3 by human vascular smooth muscle cells.

Inflammation is a critical contributing factor to the development and the progression of atherosclerosis. Recently, the acute-phase protein pentraxin-3 (PTX3), which has C-terminal sequence homology with the classic pentraxin C-reactive protein (CRP), was described to be increased in patients with myocardial infarction. In this study, we have investigated the capacity of human primary vascular smooth muscle cells (VSMC), derived from arterial specimens of ten different patients, to express PTX3 after incubation with atherogenic lipoproteins. Enzymatically degraded LDL (E-LDL), which is present in human early lesions, mediated a rapid cholesterol loading and foam cell transformation of primary VSMC, which was paralleled by a marked dose- and time-dependent expression of PTX3 mRNA and release of the acute-phase protein. Expression of PTX3 mRNA was delayed and remained almost undetectable for up to 6 h of incubation with E-LDL. However, during extended exposure to E-LDL for more than 24 h, PTX3 mRNA expression increased by more than 15-fold in VSMC foam cells, which was reflected by a concomitant release of up to 211 ng/ml PTX3 protein. We provide evidence for marked expression of PTX3 by VSMC induced by degraded lipoproteins, which may lead to an in situ vascular acute-phase reaction, contributing to the inflammatory pathogenesis of atherosclerosis.

Acute-Phase Reaction↗

PubMed2004

Selective induction of pentraxin 3, a soluble innate immune pattern recognition receptor, in infectious episodes in patients with haematological malignancy.

Pentraxins are a superfamily of conserved proteins induced in response to microbial and inflammatory stimuli. Members of this family include C-reactive protein (CRP) and serum amyloid P component, collectively known as the classical short pentraxins, and the more recently discovered pentraxin 3 (PTX3), a member of the closely related subfamily of the long pentraxins. PTX3 has been shown to be produced in response to microbial infections, and highly elevated levels were reported in patients with sepsis. In this study, PTX3 levels were evaluated in sera of a group of patients with haematological malignancy. Our findings indicate that serum PTX3 was elevated in only 1/11 afebrile episodes, despite evidence of mucositis (median 1.39), in 10/10 episodes of blood stream or target organ infections (median 7.2) but, surprisingly, was normal in 5/5 episodes of invasive aspergillosis (median 1.39). The data suggest that serum PTX3 levels are elevated selectively in response to infection. These disparate responses require further study.

C-Reactive Protein↗

PubMed2004

Tumour-associated macrophages as a prototypic type II polarised phagocyte population: role in tumour progression.

Macrophages are versatile, plastic cells which respond to micro-environmental signals with distinct functional programmes. In the tumour microenvironment, tumour-associated macrophages (TAM) polarise towards a type II phenotype, oriented to the promotion of tissue remodelling and repair. As polarised type II macrophages, TAM are a key component of the inflammatory circuits that promote tumour progression.

Antigens, Neoplasm↗

PubMed2004

The partial head decondensation test is a new, quick method to assess acrosome status in human spermatozoa.

OBJECTIVE: To develop a fast method for assessing acrosome status in human spermatozoa. DESIGN: Development of a new in vitro test to assess acrosome reaction in human spermatozoa. SETTING: Academic medical institution. PATIENT(S): Normozoospermic subjects. INTERVENTION(S): Spermatozoa were isolated from fresh semen samples, capacitated, and stimulated or not with P or ionomycin. Acrosome reactions were evaluated by phase-contrast microscopy after a brief sperm incubation in a decondensing solution. The results were compared with those obtained by scanning electron microscopy and fluoresceinated lectin staining. MAIN OUTCOME MEASURE(S): Percentage of intact acrosomes. RESULT(S): The new procedure allowed intact acrosomes to be easily identified and quantified by phase-contrast microscopy. In unstimulated and ionomycin-treated spermatozoa, a very good agreement was found among the new test, scanning electron microscopy, and fluoresceinated lectin staining. In P-treated spermatozoa, the proposed method allowed a significantly higher percentage of reacted acrosomes to be resolved, likely due to its ability to detect the very initial stages of the acrosome reaction. CONCLUSION(S): The new test allows acrosome-intact and acrosome-reacted spermatozoa to be unambiguously singled out and quantified. The method is rapid, reliable, sensitive, and easy to perform, which makes it of profitable use in both basic research and diagnostic practice.

Acrosome Reaction↗