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Bastian Höchst,
Frank A Schildberg,
Pia Sauerborn,
Yvonne A Gäbel,
Heidrun Gevensleben,
Diane Goltz,
Lukas C Heukamp,
Matthias Ballmaier,
Friederike Gieseke,
Ingo Müller, Christian Kurts,
Percy A Knolle,
Linda Diehl
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ABSTRACT: BACKGROUND: Myeloid derived suppressor cells (MDSCs) are a heterogeneous population of cells associated with the suppression of immunity. However, little is known about how or where MDSCs are induced and from which cells they originate. The liver is known for its immune regulatory functions. Here, we investigated the capacity of human hepatic stellate cells (HSCs) to transform peripheral blood monocytes into MDSCs. METHODS: We cultured freshly isolated human monocytes from healthy donors on primary human HSCs or an HSC cell-line and characterized the phenotype and function of resulting CD14(+)HLA-DR(-/low) monocytes by flow cytometry, quantitative PCR and functional assays. We analyzed the molecular mechanisms underlying the induction and function of the CD14(+)HLA-DR(-/low) cells by using blocking antibodies or knock-down technology. RESULTS: Mature peripheral blood monocytes co-cultured with HSCs down-regulated HLA-DR and developed a phenotypic and functional profile similar to MDSCs. Only activated but not freshly isolated HSCs were capable of inducing CD14(+)HLA-DR(-/low) cells. Such CD14(+)HLA-DR(-/low) monocyte-derived MDSCs suppressed T-cell proliferation in an arginase-1 dependent fashion. HSC-induced development of CD14(+)HLA-DR(-/low) monocyte-derived MDSCs was not mediated by soluble factors, but required physical interaction and was abrogated by blocking CD44. CONCLUSIONS: Our study shows that activated human HSCs convert mature peripheral blood monocytes into MDSCs. As HSCs are activated during chronic inflammation the subsequent local induction of MDSCs may prevent ensuing excessive liver injury. HSC-induced MDSCs functionally and phenotypically resemble those isolated from liver cancer patients. Thus, our data suggest that local generation of MDSCs by liver-resident HSCs may contribute to immune suppression during inflammation and cancer in the liver.
Journal of Hepatology 05/2013; · 9.26 Impact Factor
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Li-Rung Huang,
Dirk Wohlleber,
Florian Reisinger,
Craig N Jenne,
Ru-Lin Cheng,
Zeinab Abdullah,
Frank A Schildberg,
Margarete Odenthal,
Hans-Peter Dienes,
Nico van Rooijen,
Edgar Schmitt,
Natalio Garbi,
Michael Croft, Christian Kurts,
Paul Kubes,
Ulrike Protzer,
Mathias Heikenwalder,
Percy A Knolle
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ABSTRACT: Chronic infection is difficult to overcome because of exhaustion or depletion of cytotoxic effector CD8(+) T cells (cytotoxic T lymphoytes (CTLs)). Here we report that signaling via Toll-like receptors (TLRs) induced intrahepatic aggregates of myeloid cells that enabled the population expansion of CTLs (iMATEs: 'intrahepatic myeloid-cell aggregates for T cell population expansion') without causing immunopathology. In the liver, CTL proliferation was restricted to iMATEs that were composed of inflammatory monocyte-derived CD11b(+) cells. Signaling via tumor-necrosis factor (TNF) caused iMATE formation that facilitated costimulation dependent on the receptor OX40 for expansion of the CTL population. The iMATEs arose during acute viral infection but were absent during chronic viral infection, yet they were still induced by TLR signaling. Such hepatic expansion of the CTL population controlled chronic viral infection of the liver after vaccination with DNA. Thus, iMATEs are dynamic structures that overcome regulatory cues that limit the population expansion of CTLs during chronic infection and can be used in new therapeutic vaccination strategies.
Nature Immunology 04/2013; · 26.01 Impact Factor
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ABSTRACT: This unit describes a simple way to induce pyelonephritis in female mice using uropathogenic Escherichia coli (UPEC). Methods for culturing and preparing working stocks of UPEC are provided. Protocols to measure the bacterial load in the murine kidney following the establishment of pyelonephritis by determining the bacterial colony forming units (CFU) are also included. This assay can be performed with kidney homogenates if the bacterial load is the sole read-out or if cellular intactness is not required for subsequent assays, or from kidney single cell suspensions prepared by enzymatic digestion. A support protocol describes methods for isolating cells for flow cytometry, ex vivo culture, and other assays. Curr. Protoc. Immunol. 101:15.23.1-15.23.9. © 2013 by John Wiley & Sons, Inc.
Current protocols in immunology / edited by John E. Coligan ... [et al.] 04/2013; Chapter 15:Unit15.23.
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Jan P Böttcher,
Oliver Schanz,
Dirk Wohlleber,
Zeinab Abdullah,
Svenja Debey-Pascher,
Andrea Staratschek-Jox,
Bastian Höchst,
Silke Hegenbarth,
Jessica Grell,
Andreas Limmer,
Imke Atreya,
Markus F Neurath,
Dirk H Busch,
Edgar Schmitt,
Peter van Endert,
Waldemar Kolanus, Christian Kurts,
Joachim L Schultze,
Linda Diehl,
Percy A Knolle
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ABSTRACT: Development of CD8(+) T cell (CTL) immunity or tolerance is linked to the conditions during T cell priming. Dendritic cells (DCs) matured during inflammation generate effector/memory T cells, whereas immature DCs cause T cell deletion/anergy. We identify a third outcome of T cell priming in absence of inflammation enabled by cross-presenting liver sinusoidal endothelial cells. Such priming generated memory T cells that were spared from deletion by immature DCs. Similar to central memory T cells, liver-primed T cells differentiated into effector CTLs upon antigen re-encounter on matured DCs even after prolonged absence of antigen. Their reactivation required combinatorial signaling through the TCR, CD28, and IL-12R and controlled bacterial and viral infections. Gene expression profiling identified liver-primed T cells as a distinct Neuropilin-1(+) memory population. Generation of liver-primed memory T cells may prevent pathogens that avoid DC maturation by innate immune escape from also escaping adaptive immunity through attrition of the T cell repertoire.
Cell reports. 03/2013;
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ABSTRACT: Although the spleen is a major site where immune tolerance to circulating innocuous antigens occurs, the kidney also contributes. Circulating antigens smaller than albumin are constitutively filtered and concentrated in the kidney and reach the renal lymph node by lymphatic drainage, where resident dendritic cells (DCs) capture them and induce tolerance of specific cytotoxic T cells through unknown mechanisms. Here, we found that the coinhibitory cell surface receptor programmed death 1 (PD-1) on cytotoxic T cells mediates to their tolerance. Renal lymph node DCs of the CD8(+) XCR1(+) subset, which depend on the transcription factor Batf3, expressed the PD-1 cognate ligand PD-L1. Batf3-dependent DCs in the renal lymph node presented antigen that had been concentrated in the kidney and used PD-L1 to induce apoptosis of cytotoxic T cells. In contrast, T cell tolerance in the spleen was independent of PD-1, PD-L1, and Batf3. In summary, these results clarify how the kidney/renal lymph node system tolerizes the immune system against circulating innocuous antigens.
Journal of the American Society of Nephrology 02/2013; · 9.66 Impact Factor
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ABSTRACT: Professional antigen-presenting cells such as dendritic cells (DCs) and macrophages internalize extracellular antigens, process them intracellularly, and present the resulting antigen-derived peptides in the context of MHC I or MHC II molecules. Since the intracellular routing of the antigen determines whether antigens are presented on MHC I or MHC II molecules, a profound analysis of the intracellular distribution of the internalized antigens is of high interest. Here, we describe an immunofluorescence protocol to monitor the intracellular routing of the model-antigen Ovalbumin in bone marrow-derived dendritic cells (BM-DCs). This protocol describes a procedure to stain such cells with antibodies against different endosomal markers, such as EEA1 and LAMP1, and can be easily adopted to other antigen-presenting cells or antigens.
Methods in molecular biology (Clifton, N.J.) 01/2013; 960:371-377.
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ABSTRACT: Isolation and characterization of antigen-containing endosomes remains difficult utilizing standard purification techniques. Here, we describe a method, which allows isolation of antigen-loaded endosomes, that is based on flow cytometrical analysis and sorting. We specifically isolated antigen-containing endosomes from cells that had taken up fluorochrome-labeled ovalbumin via mannose receptor-mediated endocytosis. The protocol described here allows for the isolation of pure fractions of ovalbumin-containing endosomes and the extraction of proteins from these endosomes for analysis by western blot. Importantly, this protocol can be easily extended to other fluorochrome-labeled antigens and to primary antigen-presenting cells like bone marrow-derived dendritic cells.
Methods in molecular biology (Clifton, N.J.) 01/2013; 960:379-388.
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ABSTRACT: Introduction: Standardized intestinal manipulation (IM) leads to generalized postoperative ileus (POI). Previously, we demonstrated that POI is immune-mediated. Aim of this study was to investigate the role of secondary lymphoid organs (mesenteric lymph nodes/MLN, GALT) in IM-mediated POI by employing lymphatic-deficient mice (aly/aly, MLN ex) or by administration of immunosuppressants FTY720. Materials and Methods: Jejunal and colonic muscularis from wild-type (WT) mice as control, from aly/aly mice, from FTY720 treated mice (daily dose 1.0 mg/kg mouse i.p. starting 3 days before surgical procedure) and mice after removal of mesenteric lymph nodes prior to IM (MLN ex mice) were obtained after IM or sham operation. The POI was analyzed by measuring the gastrointestinal transit time, colonic transit time, infiltration of myeloperoxidase positive cells, and circular smooth muscle contractility. Furthermore, the mRNA levels of inflammatory cytokines (IL-6, TNF-α, MIP-1α) were determined by Taqman®-PCR. Results: We observed a significantly reduced upregulation of proinflammatory cytokines (IL-6, TNF-α, MIP-1α) in colonic muscularis of lymphatic-deficient mice and FTY720 treated mice. The colonic, but not the jejunal contractility was significantly improved in these mice. Additionally, the colonic inflammation and the colonic transit time were significantly improved. In summary, the lack of secondary lymphoid organs cancelled the POI. Conclusion: These data demonstrate that secondary lymphoid organs are involved in the propagation of POI. FTY720 indirectly affects POI by inhibiting migration of activated T cells from the jejunum and adjacent secondary lymphoid organs to the colon. These findings support the crucial role of the adaptive immune system in POI.
AJP Gastrointestinal and Liver Physiology 12/2012; · 3.43 Impact Factor
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Jan-Hendrik Riedel,
Hans-Joachim Paust,
Jan-Eric Turner,
André P Tittel,
Christian Krebs,
Erik Disteldorf,
Claudia Wegscheid,
Gisa Tiegs,
Joachim Velden,
Hans-Willi Mittrücker,
Natalio Garbi,
Rolf A K Stahl,
Oliver M Steinmetz, Christian Kurts,
Ulf Panzer
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ABSTRACT: Immature renal dendritic cells (DCs) are protective early in murine crescentic GN, but the mechanisms underlying this protection are unknown. Here, depletion of DCs reduced the recruitment of invariant natural killer T (iNKT) cells, which attenuate GN, into the kidney in the early stage of experimental crescentic GN. More than 90% of renal iNKT cells expressed the chemokine receptor CXCR6, and renal DCs produced high amounts of the cognate ligand CXCL16 early after induction of nephritis, suggesting that renal DC-derived CXCL16 might attract protective CXCR6(+) iNKT cells. Consistent with this finding, CXCR6-deficient mice exhibited less iNKT cell recruitment and developed nephritis that was more severe, similar to the aggravated nephritis observed in mice depleted of immature DCs. Finally, adoptive transfer of CXCR6-competent NKT cells ameliorated nephritis. Taken together, these results suggest an immunoprotective mechanism involving immature DCs, CXCL16, CXCR6, and regulatory iNKT cells, which might stimulate the development of new therapeutic strategies for GN.
Journal of the American Society of Nephrology 11/2012; · 9.66 Impact Factor
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Zeinab Abdullah,
Martin Schlee,
Susanne Roth,
Mobarak Abu Mraheil,
Winfried Barchet,
Jan Böttcher,
Torsten Hain,
Sergej Geiger,
Yoshihiro Hayakawa,
Jörg H Fritz,
Filiz Civril,
Karl-Peter Hopfner, Christian Kurts,
Jürgen Ruland,
Gunther Hartmann,
Trinad Chakraborty,
Percy A Knolle
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ABSTRACT: Immunity against infection with Listeria monocytogenes is not achieved from innate immune stimulation by contact with killed but requires viable Listeria gaining access to the cytosol of infected cells. It has remained ill-defined how such immune sensing of live Listeria occurs. Here, we report that efficient cytosolic immune sensing requires access of nucleic acids derived from live Listeria to the cytoplasm of infected cells. We found that Listeria released nucleic acids and that such secreted bacterial RNA/DNA was recognized by the cytosolic sensors RIG-I, MDA5 and STING thereby triggering interferon β production. Secreted Listeria nucleic acids also caused RIG-I-dependent IL-1β-production and inflammasome activation. The signalling molecule CARD9 contributed to IL-1β production in response to secreted nucleic acids. In conclusion, cytosolic recognition of secreted bacterial nucleic acids by RIG-I provides a mechanistic explanation for efficient induction of immunity by live bacteria.
The EMBO Journal 10/2012; · 9.20 Impact Factor
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Dirk Wohlleber,
Hamid Kashkar,
Katja Gärtner,
Marianne K Frings,
Margarete Odenthal,
Silke Hegenbarth,
Carolin Börner,
Bernd Arnold,
Günter Hämmerling,
Bernd Nieswandt,
Nico van Rooijen,
Andreas Limmer,
Karin Cederbrant,
Mathias Heikenwalder,
Manolis Pasparakis,
Ulrike Protzer,
Hans-Peter Dienes, Christian Kurts,
Martin Krönke,
Percy A Knolle
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ABSTRACT: Viruses can escape cytotoxic T cell (CTL) immunity by avoiding presentation of viral components via endogenous MHC class I antigen presentation in infected cells. Cross-priming of viral antigens circumvents such immune escape by allowing noninfected dendritic cells to activate virus-specific CTLs, but they remain ineffective against infected cells in which immune escape is functional. Here, we show that cross-presentation of antigen released from adenovirus-infected hepatocytes by liver sinusoidal endothelial cells stimulated cross-primed effector CTLs to release tumor necrosis factor (TNF), which killed virus-infected hepatocytes through caspase activation. TNF receptor signaling specifically eliminated infected hepatocytes that showed impaired anti-apoptotic defense. Thus, CTL immune surveillance against infection relies on two similarly important but distinct effector functions that are both MHC restricted, requiring either direct antigen recognition on target cells and canonical CTL effector function or cross-presentation and a noncanonical effector function mediated by TNF.
Cell reports. 08/2012; 2(3):478-87.
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ABSTRACT: Dendritische Zellen (DZ) verknüpfen das angeborene und das erworbene Immunsystem und haben einen fundamentalen Einfluss sowohl auf Infektabwehr als auch auf immunvermittelte Krankheiten. In der menschlichen Niere wurden Dendritische Zellen erstmals 1981 beschrieben, doch erst in den vergangenen Jahren konnten durch Tiermodelle weitergehende Einsichten über die funktionelle Rolle dieser Zellen in Homöostase und Nierenerkrankungen erhalten werden. Renale DZ besitzen eine wichtige Wächterrolle zum Schutz gegen Nierenschaden und -infektion. In akuten immunvermittelten Krankheiten agieren sie primär antiinflammatorisch, erlangen jedoch proinflammatorische Funktionalität, wenn die renale Entzündung chronisch wird. Dann reifen sie und können Effektor-T-Zellen zur Produktion proinflammatorischer Zytokine anregen, womit sie zur Progression der Nephritis beitragen. Die Möglichkeit, neuerdings zwischen Untertypen muriner und humaner DZ unterscheiden zu können, könnte neue Wege zur Beurteilung von humanen Nierenbiopsien eröffnen. Dieser Übersichtsartikel stellt den momentanen Kenntnisstand über renale DZ vor, wobei der Schwerpunkt auf ihre Funktion in Glomerulo- und Pyelonephritiden gelegt wurde.
Nieren- und Hochdruckkrankheiten 08/2012; 41(8):325-332.
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Jan-Eric Turner,
Christian Krebs,
Andre P Tittel,
Hans-Joachim Paust,
Catherine Meyer-Schwesinger,
Sabrina B Bennstein,
Oliver M Steinmetz,
Immo Prinz,
Tim Magnus,
Thomas Korn,
Rolf A K Stahl, Christian Kurts,
Ulf Panzer
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ABSTRACT: The Th17 immune response appears to contribute to the pathogenesis of human and experimental crescentic GN, but the cell types that produce IL-17A in the kidney, the mechanisms involved in its induction, and the IL-17A-mediated effector functions that promote renal tissue injury are incompletely understood. Here, using a murine model of crescentic GN, we found that CD4(+) T cells, γδ T cells, and a population of CD3(+)CD4(-)CD8(-)γδT cell receptor(-)NK1.1(-) T cells all produce IL-17A in the kidney. A time course analysis identified γδ T cells as a major source of IL-17A in the early phase of disease, before the first CD4(+) Th17 cells arrived. The production of IL-17A by renal γδ T cells depended on IL-23p19 signaling and retinoic acid-related orphan receptor-γt but not on IL-1β or IL-6. In addition, depletion of dendritic cells, which produce IL-23 in the kidney, reduced IL-17A production by renal γδ T cells. Furthermore, the lack of IL-17A production in γδ T cells, as well as the absence of all γδ T cells, reduced neutrophil recruitment into the kidney and ameliorated renal injury. Taken together, these data suggest that γδ T cells produce IL-17A in the kidney, induced by IL-23, promoting neutrophil recruitment, and contributing to the immunopathogenesis of crescentic GN.
Journal of the American Society of Nephrology 07/2012; 23(9):1486-95. · 9.66 Impact Factor
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ABSTRACT: The mechanisms by which regulatory T cells (T(regs)) suppress autoantibody production are unclear. Here we have addressed this question using transgenic mice expressing model antigens in the kidney. We report that T(regs) were essential and sufficient to suppress autoreactive B cells in an antigen-specific manner and to prevent them from producing autoantibodies. Most of this suppression was mediated through the inhibitory cell-surface-molecule programmed death-1 (PD-1). Suppression required PD-1 expression on autoreactive B cells and expression of the two PD-1 ligands on T(regs). PD-1 ligation inhibited activation of autoreactive B cells, suppressed their proliferation, and induced their apoptosis. Intermediate PD-1(+) cells, such as T helper cells, were dispensable for suppression. These findings demonstrate in vivo that T(regs) use PD-1 ligands to directly suppress autoreactive B cells, and they identify a previously undescribed peripheral B-cell tolerance mechanism against tissue autoantigens.
Proceedings of the National Academy of Sciences 06/2012; 109(26):10468-73. · 9.68 Impact Factor
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Bastian Höchst,
Frank A Schildberg,
Jan Böttcher,
Christina Metzger,
Sebastian Huss,
Andreas Türler,
Markus Overhaus,
Andreas Knoblich,
Berthold Schneider,
Dimitrios Pantelis, Christian Kurts,
Jörg C Kalff,
Percy Knolle,
Linda Diehl
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ABSTRACT: Immunity against cancer is impeded by local mechanisms promoting development of tumor-specific T cell tolerance, such as regulatory T cells, myeloid-derived suppressor cells, or immunosuppressive factors in the tumor microenvironment. The release of soluble antigens, such as carcinoembryonic antigen (CEA) from colorectal carcinoma (CRC) cells, has been investigated for diagnostic purposes, but not for its immunological consequences. Here, we address the question of whether soluble CEA influences tumor-specific immunity. Mice were injected with soluble CEA protein, and CEA-specific CD8 T cells were analyzed for their phenotype and functionality by means of restimulation ex vivo or antitumor efficacy in vivo. We furthermore characterized the CD8 T cell population in peripheral blood mononuclear cell (PBMCs) from healthy donors and colorectal carcinoma patients. In mice, circulating CEA was preferentially taken up in a mannose receptor-dependent manner and cross-presented by liver sinusoidal endothelial cells, but not dendritic cells, to CD8 T cells. Such systemically circulating CEA promoted tolerization of CEA-specific CD8 T cells in the endogenous T cell repertoire through the coinhibitory molecule B7H1. These CD8 T cells were not deleted but were rendered nonresponsive to antigen-specific stimulation and failed to control growth of CEA-expressing tumor cells. These nonresponsive CD8 T cells were phenotypically similar to central memory T cells being CD44(high) CD62L(high) CD25(neg) . We found T cells with a similar phenotype in PBMCs of healthy donors and at increased frequency also in patients with colorectal carcinoma. Conclusion: Our results provide evidence for the existence of an unrecognized tumor immune escape involving cross-presentation of systemically circulating tumor antigens that may influence immunotherapy of cancer. (HEPATOLOGY 2012).
Hepatology 05/2012; · 11.66 Impact Factor
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Matthias Zehner,
Judith Rauen,
Achmet I Chasan,
Maria Embgenbroich,
Marcel G Camps,
Sylvia Kaden,
Albert Haas, Christian Kurts,
Elmar Endl,
Marc Beyer,
Hermann-Josef Gröne,
Ferry Ossendorp,
Sven Burgdorf
European Journal of Immunology 05/2012; 42(8):2187-90. · 5.10 Impact Factor
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Stephanie Hucke,
Juliane Floßdorf,
Berit Grützke,
Ildiko R Dunay,
Kathrin Frenzel,
Johannes Jungverdorben,
Bettina Linnartz,
Matthias Mack,
Michael Peitz,
Oliver Brüstle, Christian Kurts,
Thomas Klockgether,
Harald Neumann,
Marco Prinz,
Heinz Wiendl,
Percy Knolle,
Luisa Klotz
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ABSTRACT: During central nervous system autoimmunity, interactions between infiltrating immune cells and brain-resident cells are critical for disease progression and ultimately organ damage. Here, we demonstrate that local cross-talk between invading autoreactive T cells and auto-antigen-presenting myeloid cells within the central nervous system results in myeloid cell activation, which is crucial for disease progression during experimental autoimmune encephalomyelitis, the animal model of multiple sclerosis. This T cell-mediated licensing of central nervous system myeloid cells triggered astrocytic CCL2-release and promoted recruitment of inflammatory CCR2(+)-monocytes, which are the main effectors of disease progression. By employing a cell-specific knockout model, we identify the nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) in myeloid cells as key regulator of their disease-determining interactions with autoreactive T cells and brain-resident cells, respectively. LysM-PPARγ(KO) mice exhibited disease exacerbation during the effector phase of experimental autoimmune encephalomyelitis characterized by enhanced activation of central nervous system myeloid cells accompanied by pronounced local CCL2 production and inflammatory monocyte invasion, which finally resulted in increased demyelination and neuronal damage. Pharmacological PPARγ activation decreased antigen-specific T cell-mediated licensing of central nervous system myeloid cells, reduced myeloid cell-mediated neurotoxicity and hence dampened central nervous system autoimmunity. Importantly, human monocytes derived from patients with multiple sclerosis clearly responded to PPARγ-mediated control of proinflammatory activation and production of neurotoxic mediators. Furthermore, PPARγ in human monocytes restricted their capacity to activate human astrocytes leading to dampened astrocytic CCL2 production. Together, interference with the disease-promoting cross-talk between central nervous system myeloid cells, autoreactive T cells and brain-resident cells represents a novel therapeutic approach that limits disease progression and lesion development during ongoing central nervous system autoimmunity.
Brain 03/2012; 135(Pt 5):1586-605. · 9.46 Impact Factor
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ABSTRACT: Studies of mechanisms responsible for the persistence of hepatitis B virus (HBV) infection have been hindered by a lack of appropriate animal models. HBV genomes can be delivered to livers of mice using hydrodynamic injection or high doses of an adenoviral vector; these lead to clearance of HBV. We found that infection of immunocompetent mice with low doses of an adenoviral vector resulted in persistent HBV infection; the mice neither underwent seroconversion to production of antibodies against HBV nor developed a strong HBV-specific effector T-cell response. As in patients with chronic HBV infection, DNA vaccination failed to generate T cells that cleared infection. This model of persistent HBV infection could be used to study the pathogenesis of chronic HBV infection and develop new therapeutic strategies.
Gastroenterology 03/2012; 142(7):1447-50.e3. · 11.68 Impact Factor
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ABSTRACT: Transgenic mice expressing the diphtheria toxin receptor (DTR) in specific cell types are key tools for functional studies in several biological systems. B6.FVB-Tg(Itgax-DTR/EGFP)57Lan/J (CD11c.DTR) and B6.Cg-Tg(Itgax-DTR/OVA/EGFP)1Gjh/Crl (CD11c.DOG) mice express the DTR in CD11c(+) cells, allowing conditional depletion of dendritic cells. We report that dendritic-cell depletion in these models caused polymorphonuclear neutrophil (PMN) release from the bone marrow, which caused chemokine-dependent neutrophilia after 6-24 h and increased bacterial clearance in a mouse pyelonephritis model. We present a transgenic mouse line, B6.Cg-Tg(Itgax-EGFP-CRE-DTR-LUC)2Gjh/Crl (CD11c.LuciDTR), which is unaffected by early neutrophilia. However, CD11c.LuciDTR and CD11c.DTR mice showed late neutrophilia 72 h after dendritic cell depletion, which was independent of PMN release and possibly resulted from increased granulopoiesis. Thus, the time point of dendritic cell depletion and the choice of DTR transgenic mouse line must be considered in experimental settings where neutrophils may be involved.
Nature Methods 02/2012; 9(4):385-90. · 19.28 Impact Factor
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ABSTRACT: The kidney is a complex organ, requiring the contributions of multiple cell types to perform its various functions. Within this system the dendritic cell has been demonstrated to play a key role in maintaining the immunological balance of the kidney. In this methods paper we aim to identify the best method for isolating murine renal dendritic cells.
The efficiency of isolating dendritic cells from enzymatically digested renal parenchyma by density centrifugation, MACS and FACS was compared.
Density centrifugation enriched dendritic cells by only approximately two fold. However, MACS and FACS resulted in a much higher purity (80% versus 95% respectively).
Although FACS gave the highest purity, MACS is the optimal method for isolating dendritic cells given cost and time factors. Isolation of a homogeneous population of renal dendritic cells will enable the molecular and functional dissection of these cells in both homeostasis and disease models.
Nephrology 02/2012; 17(4):364-71. · 1.31 Impact Factor
