The role of Triggering Receptor expressed on myeloid Cells-2 in bacterial lung infections

Pneumonie, meistens verursacht durch Streptococcus pneumoniae, ist ein weltweites Gesundheitsproblem und betrifft alle Altersgruppen sowie immunsupprimierte Patienten. Alveolarmacrophagen (AMs) sind die wichtigsten Zellen des angeborenen Immunsystems der Lunge in frühen Phasen einer Infektion und außerdem notwendig für die Erhaltung des homöostatischen Gleichgewichts. Triggering receptor expressed on myeloid cells-2 (TREM-2) ist ein Oberflächenezeptor der von AMs und anderen myeloischen Zellen exprimiert wird. Neben seiner Rolle in Knochenbildung und Gehirnfunktion, ist TREM-2 ein wichtiger Negativregulator von Immunantworten, im Gegensatz zu dem nahe verwandten Rezeptor TREM-1, einem Verstärker von Entzündungsreaktionen. Dieser Kontrast ist interessant, da beide Rezeptoren mit dem gleichen Adaptorprotein, DAP12, assoziieren. Da TLRs sowie TREM-2 konstitutiv von mononuklearen Phagozyten der Lunge exprimiert werden und essentielle Faktoren für die Modulation von Entzündungsreaktionen sind, gehen wir davon aus, dass TREM-2 eine wichtige Rolle in der Pathogenese einer bakteriellen Pneumonie spielt. In dieser Arbeit verifiziere ich, dass TREM-2-/- AMs eine verstärkte phagozytische Aktivität haben, wenn sie mit S. pneumoniae. stimuliert werden. Die Zellen zeigen außerdem eine erhöhte PI3-Kinase Aktivität sowie erhöhte ERK Aktivität. Weiters gibt meine Arbeit Hinweise auf eine regulatorische Funktion von TREM-2 für die Organisation des Aktin-Zytoskeletts sowie für die Cytokinproduktion. Der Fokus dieser Arbeit lag auf der Herstellung eines TREM-2/DAP12 Fusionskonstrukts und der Generation von stabil transfizierten Zelllinien, die N-terminal getaggte TREM-1, TREM-2 oder das Fusionskonstrukte exprimieren. Das Ziel war ein Pulldown Experiment um mögliche intrazelluläre Interaktionspartner von TREM-2 zu identifizieren.Alveolar macrophages (AMs) are the first line of defense, as they protect the lung from various pathogens and are crucial for the maintenance of lung homeostasis. The triggering receptor expressed on myeloid cells-2 (TREM-2) is a surface receptor, which is expressed on AMs and many other myeloid cells. Next to the role in bone formation and brain function, TREM-2 seems to be a negative regulator of immune responses. Considering that Toll-like receptors (TLRs) as well as TREM-2 are constitutively expressed on lung mononuclear phagocytes and that these cells are crucial in the modulation of the inflammatory response upon infection, we hypothesize that TREM-2 plays an important role during bacterial pneumonia. In my work I show, that TREM-2 deficient AMs show enhanced phagocytosis of S. pneumoniae compared to wildtype cells. One possible explanation for this might be elevated Phosphatidylinositol 3 kinase (PI3-Kinase) activity, as well as increased ERK activity in the TREM-2 knockout cells. Furthermore I show evidence for the role of TREM-2 in the regulation of the actin cytoskeleton as well as for the regulatory function of TREM-2 in cytokine production. Another aspect of my diploma project was the identification of interactors downstream of TREM-2 or TREM-2/DAP12. We for this purpose decided to perform pulldown experiments and it was my task to make a TREM-2/DAP12 fusion construct and to generate stably transfected cell lines, expressing N-terminal tagged TREM-1, TREM-2 or the chimeric construct. The respective data are presented in this thesis

ACE2 is the critical in vivo receptor for SARS-CoV-2 in a novel COVID-19 mouse model with TNF-and IFN?-driven immunopathology

Despite tremendous progress in the understanding of COVID-19, mechanistic insight into immunological, disease-driving factors remains limited. We generated maVie16, a mouse-adapted SARS-CoV-2, by serial passaging of a human isolate. In silico modeling revealed how only three Spike mutations of maVie16 enhanced interaction with murine ACE2. maVie16 induced profound pathology in BALB/c and C57BL/6 mice, and the resulting mouse COVID-19 (mCOVID-19) replicated critical aspects of human disease, including early lymphopenia, pulmonary immune cell infiltration, pneumonia, and specific adaptive immunity. Inhibition of the proinflammatory cyto-kines IFN? and TNF substantially reduced immunopathology. Importantly, genetic ACE2-deficiency completely prevented mCOVID-19 development. Finally, inhalation therapy with recombinant ACE2 fully protected mice from mCOVID-19, revealing a novel and efficient treatment. Thus, we here present maVie16 as a new tool to model COVID-19 for the discovery of new therapies and show that disease severity is determined by cytokine-driven immunopathology and critically dependent on ACE2 in vivo. © Gawish et al

Myeloid Cells Restrict MCMV and Drive Stress-Induced Extramedullary Hematopoiesis through STAT1

Cytomegalovirus (CMV) has a high prevalence worldwide, is often fatal for immunocompromised patients, and causes bone marrow suppression. Deficiency of signal transducer and activator of transcription 1 (STAT1) results in severely impaired antiviral immunity. We have used cell-type restricted deletion of Stat1 to determine the importance of myeloid cell activity for the defense against murine CMV (MCMV). We show that myeloid STAT1 limits MCMV burden and infection-associated pathology in the spleen but does not affect ultimate clearance of infection. Unexpectedly, we found an essential role of myeloid STAT1 in the induction of extramedullary hematopoiesis (EMH). The EMH-promoting function of STAT1 was not restricted to MCMV infection but was also observed during CpG oligodeoxynucleotide-induced sterile inflammation. Collectively, we provide genetic evidence that signaling through STAT1 in myeloid cells is required to restrict MCMV at early time points post-infection and to induce compensatory hematopoiesis in the spleen

The Triggering Receptor Expressed on Myeloid Cells 2 Inhibits Complement Component 1q Effector Mechanisms and Exerts Detrimental Effects during Pneumococcal Pneumonia

Phagocytosis and inflammation within the lungs is crucial for host defense during bacterial pneumonia. Triggering receptor expressed on myeloid cells (TREM)-2 was proposed to negatively regulate TLR-mediated responses and enhance phagocytosis by macrophages, but the role of TREM-2 in respiratory tract infections is unknown. Here, we established the presence of TREM-2 on alveolar macrophages (AM) and explored the function of TREM-2 in the innate immune response to pneumococcal infection in vivo. Unexpectedly, we found Trem-2(-/-) AM to display augmented bacterial phagocytosis in vitro and in vivo compared to WT AM. Mechanistically, we detected that in the absence of TREM-2, pulmonary macrophages selectively produced elevated complement component 1q (C1q) levels. We found that these increased C1q levels depended on peroxisome proliferator-activated receptor-δ (PPAR-δ) activity and were responsible for the enhanced phagocytosis of bacteria. Upon infection with S. pneumoniae, Trem-2(-/-) mice exhibited an augmented bacterial clearance from lungs, decreased bacteremia and improved survival compared to their WT counterparts. This work is the first to disclose a role for TREM-2 in clinically relevant respiratory tract infections and demonstrates a previously unknown link between TREM-2 and opsonin production within the lungs

The role of triggering receptor expressed on myeloid cells-2 in inflammation

Die Feinabstimmung von TLR-mediierten Immunreaktionen ist ausschlaggebend für die Induktion von Entzündung und die Beseitigung von Bakterien unter gleichzeitiger Vermeidung von entzündungsinduzierten Kollateralschäden. Die Rekrutierung von Immunzellen und darauf folgende Phagozytose ist dabei von höchster Bedeutung für die effiziente Elimination von Pathogenen. Der "Triggering receptor expressed on myeloid cells (TREM)-2" unterdrückt einerseits die TLR-mediierte Zytokin Produktion und wurde andererseits als Phagozytoserezeptor für Bakterien beschrieben. In Anbetracht des Einflusses von TREM-2 auf diese essentiellen Mechanismen der Immunantwort wurde die Grundhypothese für diese Arbeit formuliert, nämlich dass TREM-2 eine wichtige Rolle während bakterieller Infektionen sowie während steriler Entzündung spielen würde. Unter Verwendung von Wildtyp (WT) - und TREM-2 defizienten (TREM-2-/-) Mäusen, wurde die Funktion von TREM-2 in verschiedenen Makrophagenpopulationen untersucht, sowie in vivo, während Gram-negativer Sepsis, Endotoxemie und Pneumokokken Pneumonie. Diese Arbeit zeigt, dass der Einfluss von TREM-2 auf die TLR- induzierte Entzündung sowie auf die Phagozytose von Makrophagen stark zelltyp-spezifisch ist. Während TREM-2 in vitro in Peritonealmakrophagen (PM) und "bone marrow-derived" Makrophagen (BMM) die Produktion von inflammatorischen Faktoren inhibiert und die Phagozytose verstärkt, zeigt er in Alveolarmakrophagen (AM) genau den gegenteiligen Effekt, nämlich eine Verstärkung der Zytokinantwort und eine Verringerung der Phagozytosekapazität. Die unerwarteten Effekte von TREM-2 in AMs konnten auf eine zelltyp-spezifische Unterdrückung der Expression von Komplement Komponente 1q (C1q) zurückgeführt werden. In vivo, war die Rolle von TREM-2 komplex, da TREM-2 die Immunantwort auf mehreren Ebenen beeinflusst. Während TREM-2 in allen drei Fällen die frühe Entzündungsantwort unterdrückte, war sein Einfluss auf den späteren Verlauf stark abhängig vom jeweiligen Modell. Im Fall der sterilen Entzündung durch Lipopolysaccharid (LPS) war die TREM-2 vermittelte Verzögerung der frühen Entzündung von Nachteil für das Überleben von WT Mäusen, da damit auch eine verspätete Expression des protektiven Negativregulators A20 einherging. Im Vergleich zur Endotoxemie, war während Bakteriellen Infektionen der Einfluss von TREM-2 auf die Phagozytose von größerer Bedeutung und überschattete die Wirkung von TREM-2 auf die frühe Entzündung. Demnach verbesserte TREM-2 die Phagozytose von E. coli, was der Verzögerung der frühen Entzündungsantwort entgegenwirkte. Im Gegensatz dazu, war die Phagozytose von Pneumokokken, durch die TREM-2-mediierte Unterdrückung der C1q Expression verschlechtert. Zusammengefasst zeigen diese Daten, dass TREM-2 ein wichtiger Modulator der Immunantwort ist und für ein besseres Verständnis seiner Funktionen in einem Krankheits- und Zelltyp-spezifischen Kontext untersucht werden muss.Toll-like receptor (TLR) mediated responses have to be tightly controlled in order to first promote the induction of inflammation and efficient clearance of bacteria, while at the same time limiting collateral damage. The recruitment of immune cells and subsequent phagocytosis of bacteria are crucial effector mechanisms during this process. The triggering receptor expressed on myeloid cells (TREM)-2 was shown to negatively regulate the induction of inflammatory cytokines upon TLR activation and has been identified as a phagocytic receptor for bacteria. Considering TREM-2's impact on these important host defense mechanisms, the hypothesis of this thesis was that TREM-2 would play an important role during bacterial infection, as well as during sterile inflammation. Using wild-type (WT) and TREM-2 knockout (TREM-2-/-) mice the role of TREM-2 was investigated in primary macrophage populations, and in vivo during Gram-negative sepsis, endotoxemia and pneumococcal pneumonia. This work shows that the contribution of TREM-2 to the regulation of inflammatory responses and the phagocytic capacity of macrophages strongly depends on the cell-type investigated. While in vitro TREM-2 suppressed TLR-induced cytokine responses and promoted phagocytosis by peritoneal macrophages (PM) and bone marrow-derived macrophages (BMM), it enhanced TLR responses and suppressed phagocytosis by alveolar macrophages (AM). This difference was attributed to the suppression of complement component 1q (C1q) by TREM-2 in AM. In vivo, TREM-2 consistently dampened early inflammation, whereas the effects on outcome strongly depended on the type of injury. During endotoxemia TREM-2 impaired survival by delaying the induction of the otherwise protective negative regulator A20. However, upon infections with viable bacteria, TREM-2's impact on bacterial phagocytosis overshadowed its effects on early inflammation. As such, TREM-2 improved the uptake of E. coli, which antagonized the otherwise harmful effect of a delayed early inflammatory response. Conversely, TREM-2 exhibited detrimental effects during pneumococcal pneumonia, by affecting local C1q production and phagocytosis by AM. In summary, TREM-2 exerts cell-type specific effects, illustrating the importance of studying the role of TREM-2 in a context and disease specific manner.submitted by Riem GawishAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersWien, Med. Univ., Diss., 2015OeBB(VLID)171531

The lipid-sensor TREM2 aggravates disease in a model of LCMV-induced hepatitis

textabstractLipid metabolism is increasingly being appreciated to affect immunoregulation, inflammation and pathology. In this study we found that mice infected with lymphocytic choriomeningitis virus (LCMV) exhibit global perturbations of circulating serum lipids. Mice lacking the lipid-sensing surface receptor triggering receptor expressed on myeloid cells 2 (Trem2 -/-) were protected from LCMV-induced hepatitis and showed improved virus control despite comparable virus-specific T cell responses. Non-hematopoietic expression of TREM2 was found to be responsible for aggravated hepatitis, indicating a novel role for TREM2 in the non-myeloid compartment. These results suggest a link between virus-perturbed lipids and TREM2 that modulates liver pathogenesis upon viral infection. Targeted interventions of this immunoregulatory axis may ameliorate tissue pathology in hepatitis