Biomedical subjects

Maria Elena Remoli

Publications and source records attributed to Maria Elena Remoli.

8 recordsLinked to original sources

PubMed2006

NF-{kappa}B is required for STAT-4 expression during dendritic cell maturation.

The transcription factor STAT-4 plays a pivotal role in the IL-12-mediated development of naive CD4+ T cells into the Th1 phenotype. Initially thought to be restricted to the lymphoid lineage, STAT-4 was subsequently shown to be expressed in the myeloid compartment, mainly in activated monocytes, macrophages, and dendritic cells (DC). Here, we have studied STAT-4 in human monocyte-derived DC, and we demonstrated that its expression can be induced by multiple stimuli, such as the ligands for TLR-4, TLR-2, and TLR-3, different pathogens, CD40 ligand, and the proinflammatory cytokines TNF-alpha and IL-1beta. It is interesting that we found that STAT-4 is tyrosine-phosphorylated in response to type I IFN but not IL-12 in human mature DC. Cloning and functional analysis of the STAT-4 promoter showed that a NF-kappaB binding site, localized at -969/-959 bp upstream of the transcriptional start site, is involved in the regulation of this gene in primary human DC. EMSAs using a probe containing this NF-kappaB binding sequence and chromatin immunoprecipitation indicated that p65/p50 and p50/p50 dimers were the main NF-kappaB/Rel proteins involved in STAT-4 gene expression in maturing DC. The mutation of this kappaB site or the overexpression of the repressor IkappaBalpha exerted an inhibitory effect on a STAT-4 promoter-driven reporter as well as on STAT-4 expression. Altogether, these results indicate that STAT-4 can be finely tuned along with DC maturation through NF-kappaB activation and that its induction may be involved in preparing the DC to be receptive to the cytokine environment present in lymphoid organs.

Base Sequence↗

PubMed2006

Differential responsiveness to IFN-alpha and IFN-beta of human mature DC through modulation of IFNAR expression.

In human monocyte-derived dendritic cells (DC), infection with Mycobacterium tuberculosis and viruses or stimulation with Toll-like receptor type 3 and 4 agonists causes the release of type I interferon (IFN). Here, we describe that the IFN-beta released upon stimulation with lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (poly I:C) is responsible for a rapid and sustained signal transducer and activator of transcription 1 and 2 activation and expression of IFN-stimulated genes, such as the transcription factor IFN regulatory factor 7 and the chemokine CXC chemokine ligand 10. The autocrine production of IFN-beta from LPS and poly I:C-matured DC (mDC) induced a temporary saturation of the response to type I IFN and a marked decline in the level of the two IFN receptor (IFNAR) subunits. It is interesting that we found that upon clearing of the released cytokines, LPS-stimulated DC reacquired full responsiveness to IFN-beta but only partial responsiveness to IFN-alpha, and their maturation process was unaffected. Monitoring of surface and total levels of the receptor subunits showed that maximal expression of IFNAR2 resumed within 24 h of clearing, and IFNAR1 expression remained low. Thus, mDC can modulate their sensitivity to two IFN subtypes through a differential regulation of the IFNAR subunits.

Antibodies, Monoclonal↗

PubMed2006

Infection of human dendritic cells with a Mycobacterium tuberculosis sigE mutant stimulates production of high levels of interleukin-10 but low levels of CXCL10: impact on the T-cell response.

The Mycobacterium tuberculosis genome encodes 13 sigma factors. We have previously shown that mutations in some of these transcriptional activators render M. tuberculosis sensitive to various environmental stresses and can attenuate the virulence phenotype. In this work, we focused on extracytoplasmic factor sigmaE and studied the effects induced by the deletion of its structural gene (sigE) in the infection of human monocyte-derived dendritic cells (MDDC). We found that the wild-type M. tuberculosis strain (H37Rv), the sigE mutant (ST28), and the complemented strain (ST29) were able to infect dendritic cells (DC) to similar extents, although at 4 days postinfection a reduced ability to grow inside MDDC was observed for the sigE mutant ST28. After mycobacterium capture, the majority of MDDC underwent full maturation and expressed both inflammatory cytokines, such as tumor necrosis factor alpha, and the regulatory cytokines interleukin-12 (IL-12), IL-18, and beta interferon (IFN-beta). Conversely, a higher level of production of IL-10 was observed in ST28-infected MDDC compared to H37Rv- or ST29-infected cell results. However, in spite of the presence of IL-10, supernatants from ST28-infected DC induced IFN-gamma production by T cells similarly to those from H37Rv-infected DC culture. On the other hand, IL-10 impaired CXCL10 production in sigE mutant-infected DC and, indeed, its neutralization restored CXCL10 secretion. In line with these results, supernatants from ST28-infected cells showed a decreased capability to recruit CXCR3+ CD4+ T cells compared to those obtained from H37Rv-infected DC culture. Thus, our findings suggest that the sigE mutant-induced secretion of IL-10 inhibits CXCL10 expression and, in turn, the recruitment of activated-effector cells involved in the formation of granulomas.

Bacterial Proteins↗

PubMed2006

Human dendritic cells following Aspergillus fumigatus infection express the CCR7 receptor and a differential pattern of interleukin-12 (IL-12), IL-23, and IL-27 cytokines, which lead to a Th1 response.

Aspergillus fumigatus is the most prevalent airborne fungal pathogen and causes fatal invasive aspergillosis in immunocompromised patients. Given the essential role of dendritic cells (DC) in initiating and regulating immune responses, we investigated the impact of A. fumigatus conidial infection on human DC. A. fumigatus conidia were rapidly internalized and induced the release of tumor necrosis factor alpha within the first 8 h. After A. fumigatus infection, the majority of DC underwent full maturation, although CCR7 expression was observed only in DC that had internalized the conidia. Additionally, the analysis of regulatory cytokines showed that infected DC simultaneously produced interleukin-12p70 (IL-12p70) and significant amounts of IL-10. IL-10 neutralization was not able to further increase IL-12p70 production from infected DC. Whereas the central role of IL-12 in the generation of Th1 cells has long been appreciated, recently two other members of the IL-12 family, IL-23 and IL-27, were reported to play important roles in the regulation of gamma interferon (IFN-gamma) production from naïve and memory T cells. A. fumigatus-infected DC were also able to express high levels of IL-23p19 and low levels of IL-27p28 at later stages of infection. According to this expression pattern, A. fumigatus-infected DC were able to prime IFN-gamma production of naïve T cells. Thus, this study on the expression of the new IL-12 family members controlling the Th1 response sheds light on a novel aspect of the contribution of DC to anti-Aspergillus immunity.

Aspergillosis↗

PubMed2005

Astrocytes produce dendritic cell-attracting chemokines in vitro and in multiple sclerosis lesions.

As a result of their close association with the blood-brain barrier, astrocytes play an important role in regulating the homing of different leukocyte subsets to the inflamed central nervous system (CNS). In this study, we investigated whether human astrocytes produce chemokines that promote the migration of myeloid dendritic cells (DCs). By reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, we show that cultured human astrocytes stimulated with interleukin-1beta and tumor necrosis factor produce CCL2, CCL3, CCL4, CCL5, CCL20, and CXCL12 that act on immature DCs, but not CCL19 and CCL21, 2 chemokines specific for mature DCs. Compared with controls, supernatants of cytokine-stimulated astrocytes are more effective in promoting the migration of immature monocyte-derived DCs (iMDDCs). Desensitization of CXCR4 (receptor for CXCL12), CCR1-3-5 (shared receptors for CCL3-4-5), and CCR6 (receptor for CCL20) on iMDDC reduces cell migration toward astrocyte supernatants, indicating that astrocytes release biologically relevant amounts of iMDDC-attracting chemokines. By immunohistochemistry, we show that CXCL12 and, to a lesser extent, CCL20 are expressed by reactive astrocytes in multiple sclerosis lesions. These data lend support to the idea that astrocyte-derived chemokines may contribute to immature DC recruitment to the inflamed CNS.

Antigens, CD↗

PubMed2004

Viral infection and Toll-like receptor agonists induce a differential expression of type I and lambda interferons in human plasmacytoid and monocyte-derived dendritic cells.

In humans, the type I interferon (IFN) family consists of 13 IFN-alpha subtypes, IFN-beta and IFN-omicron the newly discovered IFN-like family consists of IFN-lambda1, -lambda2 and -lambda3. We have investigated the expression of type I and lambda IFN genes following virus infections or Toll-like receptor (TLR) triggering in monocyte-derived DC (MDDC) and plasmacytoid DC (pDC). We found that all IFN-alpha, -beta, -omicron and -lambda subtypes are expressed in influenza-virus-infected MDDC or pDC. Conversely, differential type I IFN gene transcription was induced in MDDC and pDC stimulated by specific TLR agonists. TLR-9 stimulation by CpG DNA induced the expression of all IFN-alpha, -beta, -omicron and -lambda subtypes in pDC, whereas TLR-4 stimulation by LPS, or TLR-3 stimulation by poly I:C, induced only IFN-beta and IFN-lambda gene expression in MDDC. The expression pattern of IFN regulatory factor (IRF)-5 and IRF-7 in MDDC and pDC was also determined. IRF-5 was constitutively expressed in the two DC subsets whereas IRF-7 was constitutive in pDC but its expression was induced along MDDC maturation. Overall, our data indicate that the coordinated expression of IFN-lambda with IFN-beta would be of crucial importance for the maturation of DC.

Cell Line↗

PubMed2003

IFN-alpha beta released by Mycobacterium tuberculosis-infected human dendritic cells induces the expression of CXCL10: selective recruitment of NK and activated T cells.

We recently reported that dendritic cells (DC) infected with Mycobacterium tuberculosis (Mtb) produce Th1/IFN-gamma-inducing cytokines, IFN-alpha beta and IL-12. In the present article, we show that maturing Mtb-infected DC express high levels of CCR7 and they become responsive to its ligand CCL21. Conversely, CCR5 expression was rapidly lost from the cell surface following Mtb infection. High levels of CCL3 and CCL4 were produced within 8 h after infection, which is likely to account for the observed CCR5 down-modulation on Mtb-infected DC. In addition, Mtb infection stimulated the secretion of CXCL9 and CXCL10. Interestingly, the synthesis of CXCL10 was mainly dependent on the Mtb-induced production of IFN-alpha beta. Indeed, IFN-alpha beta neutralization down-regulated CXCL10 expression, whereas the expression of CXCL9 appeared to be unaffected. The chemotactic activity of the Mtb-infected DC supernatants was evaluated by migration assays using activated NK, CD4(+), and CD8(+) cells that expressed both CCR5 and CXCR3. Mtb-induced expression of CCL3, CCL4, CXCL9, and CXCL10 was involved in the stimulation of NK and T cell migration. In accordance with the data on the IFN-alpha beta-induced expression of CXCL10, neutralization of IFN-alpha beta significantly reduced the chemotactic activity of the supernatant from Mtb-infected DC. This indicates that IFN-alpha beta may modulate the immune response through the expression of CXCL10, which along with CXCL9, CCL3, and CCL4 participates in the recruitment and selective homing of activated/effector cells, which are known to accumulate at the site of Mtb infection and take part in the formation of the granulomas.

CD4-Positive T-Lymphocytes↗

PubMed2002

Selective expression of type I IFN genes in human dendritic cells infected with Mycobacterium tuberculosis.

Type I IFN regulates different aspects of the immune response, inducing a cell-mediated immunity. We have recently shown that the infection of dendritic cells (DC) with Mycobacterium tuberculosis (Mtb) induces IFN-alpha. In this work we have monitored a rapid induction of IFN-beta followed by the delayed production of the IFN-alpha1 and/or -alpha13 subtypes. The Mtb infection rapidly activates the NF-kappaB complex and stimulates the phosphorylation of IFN regulatory factor (IRF)-3, events known to induce IFN-beta expression in viral infection. In turn, the autocrine production of IFN-beta induces the IFN-stimulated genes that contain binding sites for activated STATs in their promoters. Among the IFN-stimulated genes induced in DC through STAT activation are IRF-1 and IRF-7. The expression of IRF-1 appears to be dependent on the sequential activation of NF-kappaB and STAT-1. Once expressed, IRF-1 may further stimulate the transcription of IFN-beta. Induction of IRF-7 is also regulated at the transcriptional level through the binding of phosphorylated STAT-1 and STAT-2, forming the IFN-stimulated gene factor-3 complex. In turn, the IRF-1 and IRF-7 expression appears to be required for the delayed induction of the IFN-alpha1/13 genes. Although correlative, our results strongly support the existence of a cascade of molecular events in Mtb-infected DC. Upon infection, constitutively expressed NF-kappaB and IRF-3 are activated and likely contribute to the rapid IFN-beta expression. In turn, IFN-beta-induced IRF-1 and IRF-7 may cooperate toward induction of IFN-alpha1/13 if infection persists and these factors are activated.

Cells, Cultured↗