an-experimental-model-to-study-tuberculosis-malaria-coinfection-upon

An Experimental Model to Study Tuberculosis-Malaria Coinfection upon Natural Transmission of <i>Mycobacterium tuberculosis</i> and <i>Plasmodium berghei</i>

Coinfections naturally occur due to the geographic overlap of distinct types of pathogenic organisms. Concurrent infections most likely modulate the ...

Leukocyte recruitment to the pregnant mouse uterus has been suggested to be associated with highly regulated expression of distinct patterns of vascular adhesion receptors. One of the most striking observations is the combined expression of mucosal addressin cell adhesion molecule-1 (MAdCAM-1) and P-selectin by maternal vessels of the vascular zone during the critical period of initial placenta development. The predominant cell population within these vessels is of the monocyte/macrophage lineage and expresses the mucosal integrin alpha4beta7, which represents the ligand for MAdCAM-1; neutrophils and lymphocytes are rare. To directly assess the importance of identified adhesion receptors, we undertook long-term in vivo inhibition studies using monoclonal antibodies to inhibit the contribution of MAdCAM-1 in leukocyte trafficking to the decidua or to deplete alpha4beta7(+) leukocytes. In addition, implantation sites of mouse strains genetically deficient in specific adhesion receptors were investigated. Our results underline the importance of predicted adhesion pathways in the recruitment of monocyte-like cells, especially those expressing alpha4beta7. Interestingly, maternal/fetal units with inhibited recruitment of alpha4beta7(+) leukocytes or the absence of these cells are characterized by reduced size and frequency of uterine NK cells.

Functional involvement of P-selectin and MAdCAM-1 in the recruitment of alpha4beta7-integrin-expressing monocyte-like cells to the pregnant mouse uterus.

Interleukin-13 (IL-13) is a cytokine with a crucial role in the development of allergic asthma. The IL-13 receptor shares the IL-4Ralpha subunit with the IL-4R system, but contains as a specific component the IL-13Ralpha1 chain. Blocking signal release by IL-13 without affecting IL-4 function is a potentially interesting therapeutical option for the treatment of asthma. Employing genetic immunization, we generated a set of novel monoclonal antibodies to the IL-13Ralpha1 receptor that proved very specific and efficient inhibitors of human IL-13 activity. Receptor binding antibodies were identified by their specific reactivity with both human monocytes and a murine pro-B cell line overexpressing human IL-13Ralpha1 by flow cytometry and cell ELISA. A luciferase reporter cell system based on STAT6-mediated promoter activation in murine Ba/F3 cells was employed to screen the antibodies for IL-13 antagonistic properties. Inhibitory antibody effects were quantified by interference with IL-13-dependent proliferation of TF-1 cells. The capability of blocking IL-13-driven responses of primary, inflammation-relevant cells was tested by Western blot analysis of STAT6 tyrosine phosphorylation and expression of 15-lipoxygenase in monocytes from fresh blood. The most potent inhibitory antibody identified, GM1E7, inhibited IL-13-driven gene activation and cell proliferation in immune cell lines with IC(50) values in the low nanomolar range. Both short-term (STAT6 activation) and long-term (15-LO induction) responses of primary human blood cells to IL-13 were almost entirely blocked, whereas IL-4 effects remained virtually unaffected. GM1E7 is superior to available agents interfering with IL-13 activity in terms of specificity and efficiency and offers potential novel therapeutic perspectives for the treatment of allergic asthma.

Blockade of interleukin-13-mediated cell activation by a novel inhibitory antibody to human IL-13 receptor alpha1.

Leukocyte recruitment to the pregnant mouse uterus is associated with highly regulated patterns of expression of vascular adhesion receptors. One striking observation is the localized expression of mucosal vascular addressin cell adhesion molecule (MADCAM1) and selectin, platelet (SELP, formerly P-selectin) by maternal vessels in the vascular zone (VZ) during the first half of pregnancy. From midgestation onwards, endothelial cells lining the maternal vessels of the VZ in addition express vascular cell adhesion molecule-1 (VCAM1). The predominant cell population within these vessels is monocyte-like cells. Granulocytes and low numbers of lymphocytes are also present. Murine fetal trophoblast cells are almost devoid of adhesion molecules, including SELP. In contrast, spontaneous abortions of allogeneic pregnancies are characterized by dramatic upregulation of SELP on maternal VZ vessels and on fetal trophoblast cells. Upregulation of SELP is associated with a dramatic influx of highly activated granulocytes, which infiltrate the vessels and tissue of the VZ and the trophoblast. The majority of the activated granulocytes within the trophoblast undergo nuclear fragmentation, which can be detected by TUNEL staining. To demonstrate that SELP is involved in the recruitment of granulocytes to the pregnant uterus, we undertook long-term in vivo inhibition studies using a monoclonal antibody to inhibit the contribution of SELP to leukocyte trafficking to the decidua. In addition, the pregnant uteri of syngeneic Selp(-/-) x Selp(-/-) mice were investigated and compared to the controls. Our results clearly demonstrate the importance of SELP for granulocyte access to the pregnant mouse uterus under physiological and pathological conditions.

Selectin, platelet plays a critical role in granulocyte access to the pregnant mouse uterus under physiological and pathological conditions.

Rekrutierung von Leukozyten in den Schwangere Maus-Uterus ist stark regulierten Muster des Ausdrucks der vaskul&auml;ren Adh&auml;sion Rezeptoren zugeordnet. Eine auffallende Beobachtung ist die lokalisierte Ausdruck der Schleimhaut vaskul&auml;re Adressein Zelladh&auml;sionsmolek&uuml;l (MADCAM1) und selectin, Thrombozyten (SELP, ehemals P-selectin) durch m&uuml;tterliche Schiffe in der vaskul&auml;ren Zone (VZ) in der ersten H&auml;lfte der Schwangerschaft. Von Midgestation ab schnelles Endothelzellen die m&uuml;tterlichen Gef&auml;&szlig;e von der VZ zus&auml;tzlich Futter vaskul&auml;re Zelle Adh&auml;sion Molek&uuml;l-1 (VCAM1). Die vorherrschende Zellpopulation innerhalb dieser Schiffe ist Monozyte-&auml;hnlichen Zellen. Granulozyten und geringe Zahl von Lymphozyten sind auch vorhanden. Murine fetalen Trophoblast Zellen entbehren fast Adh&auml;sionsmolek&uuml;le, einschlie&szlig;lich SELP. Im Gegensatz dazu sind spontane Abtreibungen allogene Schwangerschaften durch dramatische Hochregulation der SELP auf m&uuml;tterliche VZ-Schiffe und fetalen Trophoblast Zellen gekennzeichnet. Hochregulation der SELP ist eine dramatische Zustrom von hoch aktivierten Granulozyten, die die Schiffe und Gewebe von der VZ und Trophoblastzellen infiltrieren zugeordnet. Die meisten der aktivierten Granulozyten innerhalb der Trophoblast unterziehen nukleare Zersplitterung, die nicht entdeckt werden kann von TUNEL zu beflecken. Um zu demonstrieren, dass die Einstellung von Granulozyten in den schwangeren Uterus SELP beteiligt ist, unternahmen wir in-vivo Hemmung Langzeitstudien mit einen monoklonaler Antik&ouml;rper um den Beitrag der SELP hemmen Leukozyte Menschenhandel an der Decidua. Dar&uuml;ber hinaus wurden die schwangere Uteri syngenen Selp(-/-) x Selp(-/-) M&auml;use untersucht und im Vergleich zu den Kontrollen. Unsere Ergebnisse zeigen deutlich die Bedeutung der SELP f&uuml;r Granulozyten Zugriff auf die schwangere Maus Geb&auml;rmutter unter physiologischen und pathologischen Bedingungen.

Selectin, spielt Thrombozyten eine wichtige Rolle in Granulozyten Zugang in den Uterus Schwangere Maus unter physiologischen und pathologischen Bedingungen.

Several leukocyte populations have been described within the pregnant mouse uterus, some of which express the integrin beta 7 (ITGB7). Here we demonstrate that the majority of the ITGB7(+) decidual leukocytes belong to the dendritic cell (DC) lineage. By multiparameter flow cytometric analysis we demonstrated the existence of three distinct DC subsets, characterized by differential expression of ITGA4/ITGB7 (formerly alpha4beta7-integrin) and ITGAE/ITGB7 (formerly alphaEbeta7-integrin). Importantly, the predominant DC subsets reside in distinct microdomains of the Day 9 pregnant mouse uterus. ITGAX(+) ITGAM(med) ITGA4/ITGB7(+) ITGAE(-) (formerly CD11c(+) CD11b(med) alpha4beta7(+) alphaE(-)) cells represent the majority of DCs in the vascular zone (VZ), whereas ITGAX(+) ITGAM(-) ITGAE/ITGB7(+) (formerly CD11c(+) CD11b(-) alphaEbeta7(+)) DCs are mainly located in the lower central decidua basalis (cDB) and the underlying myometrium. A population of ITGAX(+) ITGAM(low) DCs lacking ITGB7 are restricted to the cDB. Confocal microscopy studies show direct contact of VZ DCs with uterine natural killer (uNK) cells, suggesting a functional relationship between both cell populations. Collectively, our data identify three phenotypically distinct DC subsets residing in distinct microdomains of the uterus. The differential expression of ITGA4/ITGB7 and ITGAE/ITGB7 suggests distinct functional roles of the different DC subsets during early pregnancy.

Identification of ITGA4/ITGB7 and ITGAE/ITGB7 expressing subsets of decidual dendritic-like cells within distinct microdomains of the pregnant mouse uterus.

Mehrere Leukozyten-Populationen sind innerhalb der Geb&auml;rmutter Schwangere Maus beschrieben worden, von die einige die Integrin-Beta 7 (ITGB7) schnelles. Hier zeigen wir, dass die Mehrheit der ITGB7(+) decidual Leukozyten die dendritischen Zellen (DC)-Linie geh&ouml;ren. Multiparameter Flow Cytometric Analyse zeigen wir die Existenz von drei unterschiedlichen DC-Teilmengen, gekennzeichnet durch differenzielle Expression von ITGA4/ITGB7 (ehemals alpha4beta7-Integrin) und ITGAE/ITGB7 (ehemals alphaEbeta7-Integrin). Wichtig ist, befinden sich die vorherrschenden DC-Teilmengen in unterschiedlichen Microdomains der Geb&auml;rmutter Schwangere Maus Tag 9. ITGAX(+) ITGAM(med) ITGA4/ITGB7(+) ITGAE(-) (fr&uuml;her CD11c(+) CD11b(med) alpha4beta7(+) alphaE(-)) Zellen stellen die Mehrheit der DCs in der vaskul&auml;ren Zone (VZ), w&auml;hrend ITGAX(+) ITGAM(-) ITGAE/ITGB7(+) (fr&uuml;her CD11c(+) CD11b(-) alphaEbeta7(+)) DCs sich haupts&auml;chlich in den unteren zentralen Decidua Basalis (cDB) und die zugrunde liegende Geb&auml;rmutter befinden. Eine Bev&ouml;lkerung von ITGAX(+) ITGAM(low) DCs fehlt ITGB7 beschr&auml;nken sich auf die cDB. Konfokale Mikroskopie Studien zeigen Directo VZ DCS mit uterine nat&uuml;rliche (uNK) Killerzellen, eine funktionale Beziehung zwischen beiden Zelle Bev&ouml;lkerungen vorgeschlagen. Unsere Daten ermitteln gemeinsam drei ph&auml;notypisch unterschiedliche DC Teilmengen in unterschiedlichen Microdomains der Geb&auml;rmutter wohnten. Differentielle Ausdruck der ITGA4/ITGB7 und ITGAE/ITGB7 schl&auml;gt verschiedene Funktionsrollen der verschiedenen DC-Teilmengen w&auml;hrend der fr&uuml;hen Schwangerschaft.

Identifikation von ITGA4/ITGB7 und ITGAE/ITGB7 Teilmengen decidual dendritischen-wie die Zellen unterschiedliche Microdomains der Geb&auml;rmutter Schwangere Maus zum Ausdruck zu bringen.

The vascular addressins mucosal addressin cell adhesion molecule-1, P-selectin and ICAM-1 permit alpha(4)beta(7)-integrin-expressing DC, especially those of the myeloid lineage (CD11c(+)CD11b(+) DC), to access the pregnant mouse uterus. Injection of blocking monoclonal antibodies against mucosal addressin cell adhesion molecule-1 in P-selectin(-/-) mice or experimental approaches with beta7-integrin(-/-) or ICAM-1(-/-) mice revealed that limited access or absence of CD11c(+)CD11b(+) DC at the maternal/fetal interface negatively affects the frequency, size and functional properties of uterine NK (uNK) cells. Adoptive transfer of DC obtained from WT mice into beta7-integrin(-/-) mice abrogates these effects and emphasizes the importance of DC in uNK cell differentiation. Interestingly, those implantation sites lacking CD11c(+)CD11b(+) DC are characterized by decreased IL-15 and IL-12 mRNA and/or protein levels. Chronic administration of IL-15 in these mice gives rise to uNK cell numbers and size comparable to those of WT mice, whereas additional injection of IL-12 positively affects the IFN-gamma expression of uNK cells. Real-time RT-PCR and protein arrays performed with isolated uterine DC underline the role of DC as a source of IL-15 and IL-12 in the pregnant mouse uterus.

DC within the pregnant mouse uterus influence growth and functional properties of uterine NK cells.

Die vaskul&auml;re Addressins Schleimhaut Adressein Zelle Adh&auml;sion Molek&uuml;l-1, P-selectin und ICAM-1 erlauben Alpha (4) Beta 7-Integrin-Ausdruck DC, insbesondere der myeloid Abstammung (CD11c(+)CD11b(+) DC), die schwangere Maus Geb&auml;rmutter Zugriff auf. Injektion von blockierenden monoklonalen Antik&ouml;rpern gegen Schleimhaut-Adressein Zelle Adh&auml;sion Molek&uuml;l-1 bei P-selectin(-/-) M&auml;usen oder experimentelle Ans&auml;tze mit beta7-integrin(-/-) oder ICAM-1(-/-) M&auml;usen ergab, dass beschr&auml;nkter Zugriff oder Fehlen von CD11c(+)CD11b(+) DC an der m&uuml;tterlichen und fetalen Schnittstelle die Frequenz, die Gr&ouml;&szlig;e und die funktionellen Eigenschaften der uterine NK (uNK) Zellen negativ beeinflusst. Adoptiven Transfer DC gewonnenen WT M&auml;use in beta7-integrin(-/-) M&auml;use hebt diese Effekte und unterstreicht die Bedeutung der DC in uNK Zelldifferenzierung. Interessanterweise sind Netzwerktraffic Einnistung fehlt CD11c(+)CD11b(+) DC charakterisiert durch verminderte IL-15 und IL-12-mRNA und Protein-Ebene. Chronische Verwaltung des IL-15 bei diesen M&auml;usen verursacht uNK-Zell-Zahlen und Gr&ouml;&szlig;e vergleichbar mit denen der WT-M&auml;use, w&auml;hrend zus&auml;tzliche Injektion von IL-12 den IFN-Gamma-Ausdruck von uNK Zellen positiv beeinflusst. Real-Time RT-PCR und Protein Arrays durchgef&uuml;hrt mit isolierten uterine DC unterstreichen die Rolle der DC als Quelle von IL-15 und IL-12 in der Geb&auml;rmutter der schwangeren Maus.

DC innerhalb der Geb&auml;rmutter Schwangere Maus beeinflussen Wachstum und funktionale Eigenschaften von uterine NK-Zellen.

In vitro cultured mesenchymal stromal cells (MSC) are characterized by a short proliferative lifespan, an increasing loss of proliferation capacity and progressive reduction of differentiation potential. Laminin-1, laminin-5, collagen IV and fibronectin are important constituents of the basement membrane extracellular matrix (ECM) that are involved in a variety of cellular activities, including cell attachment and motility.

Improved proliferation and differentiation capacity of human mesenchymal stromal cells cultured with basement-membrane extracellular matrix proteins.

gp130 is a common receptor chain for cytokines such as interleukin (IL)-27 and IL-6. During experimental tuberculosis (TB), IL-27 prevents optimal antimycobacterial protection and limits the pathological sequelae of chronic inflammation. The anti-inflammatory properties of IL-27 have been attributed mainly to its suppressive effect on T helper (TH) cells. However, because gp130 cytokines also suppress the inflammatory immune response of macrophages, IL-27 may also regulate inflammation by limiting the secretion of pro-inflammatory cytokines. To specifically address the role of gp130 cytokines on macrophages, the outcome of experimental TB was analysed in macrophage/neutrophil-specific gp130-deficient (LysM(cre) gp130(loxP/loxP)) mice. In these mice, the enhanced induction of inflammatory cytokines and increased expression of the inducible nitric oxide synthase (NOS2) and LRG47 was linked to a greatly augmented TH17 immune response and matrix metalloproteinase (MMP)-9 expression. However, this amplified inflammatory immune response in Mtb-infected LysM(cre) gp130(loxP/loxP) mice was not associated with reduced bacterial loads and/or accelerated pathology. Our study revealed an immunoregulatory function of gp130 cytokines on macrophages/granulocytes, which is, however, not critical for modulating the outcome of TB.

gp130 on macrophages/granulocytes modulates inflammation during experimental tuberculosis.

Human populations are rarely exposed to one pathogen alone. Particularly in high incidence regions such as sub-Saharan Africa, concurrent infections with more than one pathogen represent a widely underappreciated public health problem. Two of the world's most notorious killers, malaria and tuberculosis, are co-endemic in impoverished populations in the tropics. However, interactions between both infections in a co-infected individual have not been studied in detail. Both pathogens have a major impact on the lung as the prime target organ for aerogenic Mycobacterium tuberculosis and the site for one of the main complications in severe malaria, malaria-associated acute respiratory distress syndrome (MA-ARDS). In order to study the ramifications caused by both infections within the same host we established an experimental mouse model of co-infection between Mycobacterium tuberculosis and Plasmodium berghei NK65, a recently described model for MA-ARDS. Our study provides evidence that malaria-induced immune responses impair host resistance to Mycobacterium tuberculosis. Using the natural routes of infection, we observed that co-infection exacerbated chronic tuberculosis while rendering mice less refractory to Plasmodium. Co-infected animals presented with enhanced inflammatory immune responses as reflected by exacerbated leukocyte infiltrates, tissue pathology and hypercytokinemia accompanied by altered T-cell responses. Our results--demonstrating striking changes in the immune regulation by co-infection with Plasmodium and Mycobacterium--are highly relevant for the medical management of both infections in humans.

Natural transmission of Plasmodium berghei exacerbates chronic tuberculosis in an experimental co-infection model.

Anti-inflammatory treatment of autoimmune diseases is associated with an increased risk of reactivation tuberculosis (TB). Besides interleukin (IL-17)A, IL-22 represents a classical T helper (TH)17 cytokine and shares similar pathological effects in inflammatory diseases such as psoriasis or arthritis. Whereas IL-17A supports protective immune responses during mycobacterial infections, the role of IL-22 after infection with Mycobacterium tuberculosis (Mtb) is yet poorly characterized. Therefore, we here characterize the cell types producing IL-22 and the protective function of this cytokine during experimental TB in mice. Like IL-17A, IL-22 is expressed early after infection with Mtb in an IL-23-dependent manner. Surprisingly, the majority of IL-22-producing cells are not positive for IL-17A but have rather functional characteristics of interferon-gamma-producing TH1 cells. Although we found minor differences in the number of naive and central memory T cells as well as in the frequency of TH1 and polyfunctional T cells in mice deficient for IL-22, the absence of IL-22 does not affect the outcome of Mtb infection. Our study revealed that although produced by TH1 cells, IL-22 is dispensable for protective immune responses during TB. Therefore, targeting of IL-22 in inflammatory disease may represent a therapeutic approach that does not incur the danger of reactivation TB.

Article	Year
Functional involvement of P-selectin and MAdCAM-1 in the recruitment of alpha4beta7-integrin-expressing monocyte-like cells to the pregnant mouse uterus. European journal of immunology\| PubMed ID: 15484189	2004
Blockade of interleukin-13-mediated cell activation by a novel inhibitory antibody to human IL-13 receptor alpha1. Molecular immunology\| PubMed ID: 16364441	2006
Selectin, platelet plays a critical role in granulocyte access to the pregnant mouse uterus under physiological and pathological conditions. Biology of reproduction\| PubMed ID: 17151351	2007
Identification of ITGA4/ITGB7 and ITGAE/ITGB7 expressing subsets of decidual dendritic-like cells within distinct microdomains of the pregnant mouse uterus. Biology of reproduction\| PubMed ID: 18562709	2008
DC within the pregnant mouse uterus influence growth and functional properties of uterine NK cells. European journal of immunology\| PubMed ID: 19593769	2009
Improved proliferation and differentiation capacity of human mesenchymal stromal cells cultured with basement-membrane extracellular matrix proteins. Cytotherapy\| PubMed ID: 20807021	2010
gp130 on macrophages/granulocytes modulates inflammation during experimental tuberculosis. European journal of cell biology\| PubMed ID: 21144615	2011
Natural transmission of Plasmodium berghei exacerbates chronic tuberculosis in an experimental co-infection model. PloS one\| PubMed ID: 23110184	2012
IL-22 is mainly produced by IFNγ-secreting cells but is dispensable for host protection against Mycobacterium tuberculosis infection. PloS one\| PubMed ID: 23460846	2013

Jochen Behrends

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