Papers by Lena Alexopoulou
Toll-like receptor 3 signaling inhibits double-stranded RNA-induced apoptosis of macrophages (168.21)
The Journal of Immunology
Most viruses produce dsRNA during their infection. Recognition of dsRNA by TLR3, PKR, RIG-I, or M... more Most viruses produce dsRNA during their infection. Recognition of dsRNA by TLR3, PKR, RIG-I, or MDA-5 initiates innate immune responses, RIG-I and MDA-5 are sensors for dsRNA to induce proinflammatory cytokines or type I interferon, whereas PKR mediates dsRNA-induced apoptosis. Here, we reported that TLR3 deficiency enhances apoptosis of macrophages in response to poly(I:C) as well as Japanese encephalitis virus (JEV). Unexpectedly, TLR3 deficiency markedly increased the phosphorylation of PKR, and inhibition of PKR by RNAi or a PKR-specific inhibitory oligonucleotide (ION) reduced poly(I:C)-induced apoptosis of TLR3-/- macrophages. Stimulation of TLR3 by poly(I:C) activates PI3K and NF-κB signaling pathways, and inhibition of either PI3K or NF-κB enhanced the poly(I:C)-induced apoptosis of macrophages. Further analysis revealed that TLR3 deficiency, as well as inhibition of PI3K or NF-κB, markedly induced p58IPK, a PKR inhibitor, and inhibition of p58IPK by RNAi reduced the apoptos...
Frontiers in Immunology
Sjögren’s syndrome (SS) is a chronic systemic autoimmune disease that affects the salivary and la... more Sjögren’s syndrome (SS) is a chronic systemic autoimmune disease that affects the salivary and lacrimal glands, as well as other organ systems like the lungs, kidneys and nervous system. SS can occur alone or in combination with another autoimmune disease, such as systemic lupus erythematosus (SLE) or rheumatoid arthritis. The etiology of SS is unknown but recent studies have revealed the implication of the activation of innate immune receptors, including Toll-like receptors (TLRs), mainly through the detection of endogenous nucleic acids, in the pathogenesis of systemic autoimmune diseases. Studies on SS mouse models suggest that TLRs and especially TLR7 that detects single-stranded RNA of microbial or endogenous origin can drive the development of SS and findings in SS patients corroborate those in mouse models. In this review, we will give an overview of the function and signaling of nucleic acid-sensing TLRs, the interplay of TLR7 with TLR8 and TLR9 in the context of autoimmunit...
Alveolar macrophages (AM) are tissue resident macrophages of the lung that can be expanded in cul... more Alveolar macrophages (AM) are tissue resident macrophages of the lung that can be expanded in culture, but it is unknown to what extent culture affects their in vivo identity. Here we show that long-term ex vivo expanded mouse AM (exAM) maintain core AM gene expression but show culture adaptations related to adhesion, metabolism and proliferation. Strikingly, even after several months in culture exAM reacquired full transcriptional and epigenetic identity upon transplantation into the lung and could self-maintain in the natural niche long-term. Changes in open chromatin regions (OCR) observed in culture were fully reversible in transplanted exAM (texAM) and resulted in a gene expression profile indistinguishable from resident AM. Our results demonstrate that long-term proliferation of AM in culture does not compromise cellular identity in vivo. The demonstrated robustness of exAM identity provides new opportunities for mechanistic analysis and highlights the therapeutic potential of...
Atherosclerosis Supplements, 2011
Recognition of double-stranded RNA and activation of NF-κB by Toll-like receptor 3
Nature, 2001
Toll-like receptors (TLRs) are a family of innate immune-recognition receptors that recognize mol... more Toll-like receptors (TLRs) are a family of innate immune-recognition receptors that recognize molecular patterns associated with microbial pathogens, and induce antimicrobial immune responses. Double-stranded RNA (dsRNA) is a molecular pattern associated with viral infection, because it is produced by most viruses at some point during their replication. Here we show that mammalian TLR3 recognizes dsRNA, and that activation of the receptor induces the activation of NF-kappaB and the production of type I interferons (IFNs). TLR3-deficient (TLR3-/-) mice showed reduced responses to polyinosine-polycytidylic acid (poly(I:C)), resistance to the lethal effect of poly(I:C) when sensitized with d-galactosamine (d-GalN), and reduced production of inflammatory cytokines. MyD88 is an adaptor protein that is shared by all the known TLRs. When activated by poly(I:C), TLR3 induces cytokine production through a signalling pathway dependent on MyD88. Moreover, poly(I:C) can induce activation of NF-kappaB and mitogen-activated protein (MAP) kinases independently of MyD88, and cause dendritic cells to mature.
European Journal of Immunology, 2016
HAL is a multidisciplinary open access archive for the deposit and dissemination of scientific re... more HAL is a multidisciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés. Distributed under a Creative Commons Attribution-NonCommercial| 4.0 International License
TNF- a transgenic and knockout models of CNS inflammation and degeneration
J Neuroimmunol, 1997
Tumour necrosis factor-α (TNF-α) plays a central role in inflammatory events including those taki... more Tumour necrosis factor-α (TNF-α) plays a central role in inflammatory events including those taking place in the central nervous system (CNS), and has been implicated as a key pathogenicc mediator in several human inflammatory, infectious and autoimmune CNS disorders. Using transgenic and gene knockout mice we have investigated the role of deregulated TNF-α production in the CNS. We show that the overexpression of wild-type murine or human TNF-α transgenes by resident CNS astrocytes or neurons in sufficient to trigger a neurological disorder characterised by ataxia, seizures and paresis, with histopathological features of chronic CNS inflammation and white matter degeneration. Furthermore, we show that transmembrane human TNF-α is sufficient to trigger CNS inflammation and degeneration when overexpressed by astrocytes but not by neurons, indicating that target cells mediating the neuroinflammatory activities of TNF-α localise in the vicinity of astrocytes rather than neurons. Our results establish that both soluble and transmembrane molecular forms of TNF-α can play critical roles in vivo in the pathogenesis of CNS inflammation and demyelination, and validate TNF-α transgenic and mutant mice as important models for the further study of related human CNS diseases.
Reich-Recognition of double-stranded RNA and activation of NF-kappaB hart
Nature Medicine, 2004
West Nile virus (WNV), a mosquito-borne single-stranded (ss)RNA flavivirus, causes human disease ... more West Nile virus (WNV), a mosquito-borne single-stranded (ss)RNA flavivirus, causes human disease of variable severity. We investigated the involvement of Toll-like receptor (Tlr) 3, which recognizes viral double-stranded (ds)RNA, on WNV infection. Tlr3-deficient (Tlr3-/-) mice were more resistant to lethal WNV infection and had impaired cytokine production and enhanced viral load in the periphery, whereas in the brain, viral load, inflammatory responses and neuropathology were reduced compared to wild-type mice. Peripheral WNV infection led to a breakdown of the blood-brain barrier and enhanced brain infection in wildtype but not in Tlr3-/mice, although both groups were equally susceptible upon intracerebroventricular administration of the virus. Tumor necrosis factor-α receptor 1 signaling is vital for blood-brain barrier compromise upon Tlr3 stimulation by dsRNA or WNV. Collectively, WNV infection leads to a Tlr3-dependent inflammatory response, which is involved in brain penetration of the virus and neuronal injury.
Nature Medicine, 2002
The Lyme disease vaccine is based on the outer-surface lipoprotein (OspA) of the pathogen Borreli... more The Lyme disease vaccine is based on the outer-surface lipoprotein (OspA) of the pathogen Borrelia burgdorferi, and 95% of vaccine recipients develop substantial titers of antibodies against OspA. Here, we identified seven individuals with very low antibody titers after vaccination (low responders). The macrophages of low responders produced less tumor necrosis factor-α and interleukin-6 after OspA stimulation and had lower cell-surface expression of Toll-like receptor (TLR) 1 as compared to normal cells, but normal expression of TLR2. TLRs activate innate responses to pathogens, and TLR2 recognizes lipoproteins and peptidoglycan (PGN). After OspA immunization, mice genetically deficient in either TLR2 (TLR2-/-) or TLR1 (TLR1-/-) produced low titers of antibodies against OspA. Notably, macrophages from TLR2-/mice were unresponsive to OspA and PGN, whereas those from TLR1-/mice responded normally to PGN but not to OspA. These data indicate that TLR1 and TLR2 are required for lipoprotein recognition and that defects in the TLR1/2 signaling pathway may account for human hyporesponsiveness to OspA vaccination.
Nature, 2005
programmes are never initiated 28. However, inactivation of Pax6 later in development, at the ret... more programmes are never initiated 28. However, inactivation of Pax6 later in development, at the retinal progenitor stage, results in loss of ability of this committed but undifferentiated cell type to maintain pluripotentiality 29. Pax5, which can also interact with Grg4 (ref. 14), may play similar roles in lymphocyte development 30. Adult resident stem cells have been identified in a large number of organs and provide exciting potential for tissue regeneration. A fundamental understanding of the molecular programmes regulating both differentiation and maintenance of the undifferentiated state will be required to harness this potential. Our work characterizes a critical regulatory circuit that exemplifies conservation of genetic programmes between embryonic neural crest development and adult melanocyte stem cell function. Additional nodal checkpoints, with parallel transcriptional circuits, are likely to exist in other embryonic and adult stem cells. A Methods Immunohistochemistry Immunohistochemistry was performed on paraffin-embedded tissue fixed in 4% paraformaldehyde. Antigen was exposed using Bull's Eye reagent (Biocare Medical) and heated in a pressure cooker. Antibodies utilized were Pax3 (polyclonal sera or monoclonal supernatant,
TNF-α transgenic and knockout models of CNS inflammation and degeneration
Journal of Neuroimmunology, 1997
Tumour necrosis factor-α (TNF-α) plays a central role in inflammatory events including those taki... more Tumour necrosis factor-α (TNF-α) plays a central role in inflammatory events including those taking place in the central nervous system (CNS), and has been implicated as a key pathogenicc mediator in several human inflammatory, infectious and autoimmune CNS disorders. Using transgenic and gene knockout mice we have investigated the role of deregulated TNF-α production in the CNS. We show that
A Novel Bitriazolyl Acyclonucleoside Endowed with Dual Antiproliferative and Immunomodulatory Activity
Journal of Medicinal Chemistry, 2012
A novel bitriazolyl acyclonucleoside was discovered to exhibit powerful antiproliferative effects... more A novel bitriazolyl acyclonucleoside was discovered to exhibit powerful antiproliferative effects on different cancer cell lines through caspase-dependent apoptosis and at the same time stimulate the immune response in dendritic cells via Toll-like receptor 7 (TLR7) signaling. This promising compound with dual anticancer and immunomodulatory activity may represent a new generation of highly efficacious drug candidates for use in cancer therapy.
Inflammatory Bowel Diseases, 2004
U lcerative Colitis and Crohn's disease, collectively termed inflammatory bowel disease (IBD), ar... more U lcerative Colitis and Crohn's disease, collectively termed inflammatory bowel disease (IBD), are chronic inflammatory diseases of the gastrointestinal tract. The diseases are characterized by bloody diarrhea, abdominal pain, weight loss, fatigue, and increased risk of gastrointestinal malignancy. Understanding the molecular pathogenesis of IBD is critical toward developing new therapies. IBD appears to result from an inappropriate response to normal gastrointestinal flora. The normal mucosal immune system represents a balance between aggressive cells and regulatory cells that can suppress the activity of the former. This balance can be disturbed by either an enhanced effector cell response of the innate or adaptive immune system, or a deficiency of a regulatory mechanism. We will explore the role of innate immunity and apoptosis and how defects can lead to the imbalance seen in IBD.
YIA 1 Toll-like receptor 3 expression is increased in atherosclerosis and confers protection against early atherosclerotic lesion development and weight gain in apolipoprotein E deficient mice
Heart, 2011
ABSTRACT Introduction Previous studies have assigned detrimental roles to toll-like receptors (TL... more ABSTRACT Introduction Previous studies have assigned detrimental roles to toll-like receptors (TLR) in cardiovascular disease. Recently we described a novel protective role for TLR3 in vivo in intimal hyperplasia. Aim Using human and murine systems we investigated the consequence of TLR3 signalling in atherosclerosis. Methods and Results We compared the responses of human atheroma-derived smooth muscle cells (AthSMC) and control aortic smooth muscle cells (AoSMC) to various TLR ligands. AthSMC exhibited a specific increase in TLR3 expression and TLR3-dependent functional responses. Interestingly, exposure to the TLR3 ligand poly (I:C) induced both pro- and anti-inflammatory gene expression in vitro in AthSMC and in vivo in vascular tissues. We examined the role of TLR3 signalling in atherosclerosis in vivo using 15- and 30-week ApoE-/- and ApoE-/-TLR3-/- mice fed a chow diet. Aortic root lesion area was significantly increased in ApoE-/-TLR3-/- mice compared to ApoE-/- mice at 15- (p=0.035) but not 30-weeks of age (p>0.05) suggesting that TLR3 is protective in early but not more advanced stages of lesion development. The absence of an exogenous viral stimulus implicates an endogenous vasculoprotective TLR3 ligand. TLR3 deficiency did not affect lesional macrophage or collagen content nor cholesterol levels. Intriguingly, high-fat feeding for 15-weeks induced increased weight gain in ApoE-/-TLR3-/- mice compared to ApoE-/- mice (p=0.047). Conclusions Collectively, our data describe for the first time a protective role for TLR signalling in atherosclerosis and suggest that an endogenous TLR3 ligand may mediate the observed protection. Moreover, genetic deletion of TLR3 leads to susceptibility to high-fat diet induced obesity.
A murine transmembrane tumor necrosis factor (TNF) transgene induces arthritis by cooperative p55/p75 TNF receptor signaling
European Journal of Immunology, 1997
The arthritogenic activities of tumor necrosis factor (TNF) and its p55TNF-receptor (R) have been... more The arthritogenic activities of tumor necrosis factor (TNF) and its p55TNF-receptor (R) have been well documented in experimental animal models of arthritis, and in transgenic mice expressing wild-type or mutant transmembrane human TNF proteins in their joints. In this study we show that chronic inflammatory arthritis also develops in transgenic mice made to overexpress a mutant transmembrane from of the murine TNF protein (muTNF delta 1-12) which is known to utilize efficiently both the p55 and the p75TNFR. Cross-breeding of the transgene into a TNF knockout background did not alter development of disease. Analysis of TNF bioactivity in sera from lipopolysaccharide-stimulated mice or ex vivo macrophage cultures demonstrated that the muTNF delta 1-12 protein accumulates on the cell surface and is not processed to bioactive soluble TNF, indicating that transmembrane TNF is by itself sufficient to mediate pathogenesis of arthritis. Furthermore, using TNFR knockout mice, it is shown that development of transmembrane TNF-mediated arthritis requires the presence of the p55TNFR but is significantly delayed in the absence of the p75TNFR, suggesting a positive cooperation between the two TNFR in the arthritogenic process. These results indicate that blocking the activities of both soluble and transmembrane TNF may be required to effectively neutralize the pathogenic potential of this cytokine in arthritis.
In vivo evidence for a functional role of both tumor necrosis factor (TNF) receptors and transmembrane TNF in experimental hepatitis
European Journal of Immunology, 1997
The significance of tumor necrosis factor receptor 1 (TNFR1) for TNF function in vivo is well doc... more The significance of tumor necrosis factor receptor 1 (TNFR1) for TNF function in vivo is well documented, whereas the role of TNFR2 so far remains obscure. In a model of concanavalin A (Con A)‐induced, CD4+ T cell‐dependent experimental hepatitis in mice, in which TNF is a central mediator of apoptotic and necrotic liver damage, we now provide evidence for an essential in vivo function of TNFR2 in this pathophysiological process. We demonstrate that a cooperation of TNFR1 and TNFR2 is required for hepatotoxicity as mice deficient of either receptor were resistant against Con A. A significant role of TNFR2 for Con A‐induced hepatitis is also shown by the enhanced sensitivity of transgenic mice overexpressing the human TNFR2. The ligand for cytotoxic signaling via both TNF receptors is the precursor of soluble TNF, i.e. transmembrane TNF. Indeed, transmembrane TNF is sufficient to mediate hepatic damage, as transgenic mice deficient in wild‐type soluble TNF but expressing a mutated no...
European Journal of Immunology, 2006
Using targeted mutagenesis in mice, we have blocked shedding of endogenous murine TNF by deleting... more Using targeted mutagenesis in mice, we have blocked shedding of endogenous murine TNF by deleting its cleavage site. Mutant mice produce physiologically regulated levels of transmembrane TNF (tmTNF), which suffice to support thymocyte proliferation but cannot substitute for the hepatotoxic activities of wild‐type TNF following LPS/D‐galactosamine challenge in vivo and are not sufficient to support secondary lymphoid organ structure and function. Notably, however, tmTNF is capable of exerting anti‐Listerial host defenses while remaining inadequate to mediate arthritogenic functions, as tested in the tristetraprolin‐deficient model of TNF‐dependent arthritis. Most interestingly, in the EAE model of autoimmune demyelination, tmTNF suppresses disease onset and progression and retains the autoimmune suppressive properties of wild‐type TNF. Together, these results indicate that tmTNF preserves a subset of the beneficial activities of TNF while lacking detrimental effects. These data suppo...
Cytokine & Growth Factor Reviews, 1996
Tumour necrosis factors have been classically studied as molecules central to the pathogenesis of... more Tumour necrosis factors have been classically studied as molecules central to the pathogenesis of infectious, inflammatory and autoimmune diseases. The recent generation of mice deficient in TNF~, LT~, or their receptors, has provided exciting new insights into the physiological role of these molecules in the development of secondary lymphoid tissues and in the organisation of the humoral immune response.
Contribution of TLR7 and TLR9 signaling to the susceptibility of MyD88-deficient mice to myocarditis
Autoimmunity, 2010
Toll-like receptors (TLRs) are evolutionary conserved molecules that recognize various microbial ... more Toll-like receptors (TLRs) are evolutionary conserved molecules that recognize various microbial components and host-derived agonists from damaged cells and play a central role in innate and adaptive immunity. It has been reported that MyD88, the adaptor molecule downstream of all TLRs, except TLR3, is essential for initiation of experimental autoimmune myocarditis (EAM). To determine the role of the intracellular TLRs in EAM, TLR3(-/-), TLR7(-/-), and TLR9(-/-) mice were immunized with cardiac alpha-myosin heavy chain peptide (MyHC-alpha) in Complete Freund's Adjuvant (CFA) and their EAM scores and associated immunological responses were compared to wild-type (WT) and MyD88(-/-) mice. MyD88(-/-) mice were completely resistant to EAM and had a profound defect in all the parameters we tested. Myocardial cellular infiltration and in vitro proliferation of MyHC-alpha-restimulated splenocytes were markedly reduced in TLR7(-/-) mice, while TLR3(-/-) and TLR9(-/-) mice showed similar inflammatory cell infiltration in the heart-like WT mice. Thus, the resistance of MyD88(-/-) mice to EAM can be attributed to a certain degree to TLR7 signaling. Moreover, upon murine cytomegalovirus-induced myocarditis, we found that the severity of myocardial inflammation was higher in TLR9(-/-) and MyD88(-/-) mice compared with WT, TLR3(-/-), or TLR7(-/-) mice and paralleled the ability of the mice to fight the viral infection.
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Papers by Lena Alexopoulou