Clearance Of Apoptotic Debris Research Articles

AXL Receptor in Cancer Metastasis and Drug Resistance: When Normal Functions Go Askew.

Simple SummaryAXL is a member of the TAM (TYRO3, AXL, MER) family of receptor tyrosine kinases. In normal physiological conditions, AXL is involved in removing dead cells and their remains, and limiting the duration of immune responses. Both functions are utilized by cancers in the course of tumour progression. Cancer cells use the AXL pathway to detect toxic environments and to activate molecular mechanisms, thereby ensuring their survival or escape from the toxic zone. AXL is instrumental in controlling genetic programs of epithelial-mesenchymal and mesenchymal-epithelial transitions, enabling cancer cells to metastasize. Additionally, AXL signaling suppresses immune responses in tumour microenvironment and thereby helps cancer cells to evade immune surveillance. The broad role of AXL in tumour biology is the reason why its inhibition sensitizes tumours to a broad spectrum of anti-cancer drugs. In this review, we outline molecular mechanisms underlying AXL function in normal tissues, and discuss how these mechanisms are adopted by cancers to become metastatic and drug-resistant.The TAM proteins TYRO3, AXL, and MER are receptor tyrosine kinases implicated in the clearance of apoptotic debris and negative regulation of innate immune responses. AXL contributes to immunosuppression by terminating the Toll-like receptor signaling in dendritic cells, and suppressing natural killer cell activity. In recent years, AXL has been intensively studied in the context of cancer. Both molecules, the receptor, and its ligand GAS6, are commonly expressed in cancer cells, as well as stromal and infiltrating immune cells. In cancer cells, the activation of AXL signaling stimulates cell survival and increases migratory and invasive potential. In cells of the tumour microenvironment, AXL pathway potentiates immune evasion. AXL has been broadly implicated in the epithelial-mesenchymal plasticity of cancer cells, a key factor in drug resistance and metastasis. Several antibody-based and small molecule AXL inhibitors have been developed and used in preclinical studies. AXL inhibition in various mouse cancer models reduced metastatic spread and improved the survival of the animals. AXL inhibitors are currently being tested in several clinical trials as monotherapy or in combination with other drugs. Here, we give a brief overview of AXL structure and regulation and discuss the normal physiological functions of TAM receptors, focusing on AXL. We present a theory of how epithelial cancers exploit AXL signaling to resist cytotoxic insults, in order to disseminate and relapse.

Defects in Germinal Center Selection in SLE.

Germinal centers (GCs) are the primary site at which clonal expansion and affinity maturation of B cells occur. B cells encounter antigen and receive T cell help in the GC light zone (LZ) and then migrate to the dark zone where they proliferate and undergo somatic mutation before cycling back to the LZ for further rounds of selection. Tolerance to autoantigens is frequently lost de novo as GC B cells undergo class switching and somatic mutation. This loss of tolerance is regulated by a variety of mechanisms including cell death, failure to compete for T cell help, and failure to differentiate into effector cells. Systemic lupus erythematosus (SLE) is characterized by loss of tolerance to nucleic acid antigens. While defects in tolerance occur in the naïve repertoire of SLE patients, pathogenic autoantibodies also arise in the GC by somatic mutation from non-autoreactive precursors. Several B cell defects contribute to the loss of GC tolerance in SLE, including polymorphisms of genes encoded by the Sle1 locus, excess TLR7 signaling, defects in FcRIIB expression, or defects of B cell apoptosis. Extrinsic soluble factors, such as Type-1 IFN and B cell-activating factor, or an increased number of T follicular helper cells in the GC also alter B cell-negative selection. Finally, defects in clearance of apoptotic debris within the GC result in BCR-mediated internalization of nucleic acid containing material and stimulation of autoantibody production by endosomal TLR-driven mechanisms.

The Role of Macrophages in Promoting and Maintaining Homeostasis at the Fetal-Maternal Interface.

A successful pregnancy requires that the maternal immune system adapts properly to avoid rejection of the semi-allogeneic fetus without compromising the ability to protect the mother and the fetus against infections. In this review, we describe the role of decidual macrophages in creating a homeostatic environment at the fetal-maternal interface. We also discuss their role in pregnancy complications as well as future possibilities to modulate macrophage function therapeutically. Decidual macrophages are enriched at the fetal-maternal interface and play a major role in the regulation of inflammatory responses and the maintenance of a tolerant environment. Their function is, however, not restricted to immune tolerance, but extends to include functions such as the recognition and clearance of infections, the clearance of apoptotic debris, and tissue remodeling. Decidual macrophages seem to largely function as tissue-resident macrophages that are crucial for maintaining homeostasis and reproductive success.

Long-term persistence of anti-β2 glycoprotein I in treated leprosy patients.

β2 glycoprotein I (β2GPI) is a phospholipid binding protein that plays an important role in endothelial stability, blood coagulation, clearance of apoptotic debris and other physiologic processes. Anti-β2GPI antibodies occur in normal individuals and transiently during the course of infections, but are also associated with thrombotic events in autoimmune disease: the antiphospholipid syndrome (APS). A total of 31 out of 37 treated leprosy patients previously found to present high titers of IgM anti-β2GPI and/or anticardiolipin antibodies (aCL) remained positive for IgM antiphospholipid antibodies (aPL), and exhibited high titers of anti-β2GPI. The 37 patients were part of the 77 aPL-positive patients from a previous study that evaluated 158 leprosy patients. The median time elapsed between the first and second sample was 66 months. None of the 37 patients had any thrombotic event and 24 had a reactional state and were still requiring the use of prednisone, thalidomide or both. None of them fulfilled World Health Organization criteria for leprosy recurrence.

AB0480 Towards A Novel Tolerogenic Dendritic Cell Based Immunotherapy for Systemic Lupus Erythematosus

BackgroundTolerogenic dendritic cells (tolDCs) can be considered as an attractive strategy to treat autoimmune diseases. The tolerogenic potential of autologous tolDCs has not yet been evaluated to treat systemic lupus...

Abstract 63: MerTK and Mfge8-Dependent Efferocytosis Orchestrates Postischemic Cardiac Healing Through VEGF-A and Il-10 Regulation

Inflammation orchestrates tissue remodeling after myocardial infarction (MI). Efferocytosis is mainly carried out by the membrane protein tyrosin kinase mertk (MerTK) and the milk fat globule epidermal growth factor (Mfge8) and may balance inflammation resolution after MI. We reasoned that efficient MerTK and Mfge8-dependent clearance of apoptotic debris by infiltrated myeloid cells is critical to the angiogenic and regenerative fine-tuning of the ischemic cardiac tissue. We first demonstrated that the left ventricle ejection fraction assessed by echocardiography and the number of capillaries were decreased by around 1.5-fold in MerTK -/-, Mfge8 -/- and double deficient mice when compared to wild-type animals (WT), 14 days after the onset of ischemia (p<0.01, n=10 per group). These effects were associated with an increased infarct size, fibrotic area as well as apoptotic cells number and were also observed in irradiated WT mice transplanted with bone marrow-derived cells isolated from MerTK -/-, Mfge8 -/- and double deficient animals. In cardiac tissue of WT CD45.1 recipient mice, the number of infiltrated CD45.2 MerTK -/-/Mfge8 -/- bone marrow-derived cells was similar to that of CD45.2 WT cells. In addition, the amount of Ly6Chigh and Ly6Clow monocytes as well as that of macrophages was unaffected in infarcted heart of MerTK -/-/Mfge8 -/- bone-marrow transplanted compared to WT mice. In contrast, deficient efferocytosis was related to an increased number of cardiac annexinV-positive microparticles and a downregulation of the pro-angiogenic growth factor VEGF-A and the anti-inflammatory cytokine IL-10 protein levels. Notably, FACS-sorted Ly6Clow monocytes from ischemic cardiac tissue of MerTK -/-/Mfge8 -/- mice expressed lower levels of VEGF-A when compared to WT cells. Hence, inhibition of efferocytosis leads to an adverse cardiac remodeling after MI through phenotypic alterations of phagocytic cells.

Increased type I interferon disrupts marginal zone barrier integrity via dissociation of lymphotoxin-expressing B cells and marginal zone macrophages in systemic autoimmunity (P4160)

Abstract In the spleen, membrane expressed Lymphotoxin (LT) on B cells maintains the integrity of marginal zone macrophages (MZM) which acts as the final barrier to prevent follicular entry of autoantigens derived from apoptotic debris. In this study, we found that there was an age-dependent decrease in the number of MZMs and clearance of apoptotic debris in the spleens of autoimmune BXD2 mice. Selective depletion of MZMs by repeated injection of recombinant LTR fusion protein to BXD2 mice accelerated autoimmune phenotypes. Interestingly, in BXD2 spleen, the major LT producing MZ and MZ precursor B cells are primarily localized in the follicles due to type I IFN-induced upregulation of CD69. Adoptive transfer of B cells from BXD2-Ifnar-/- but not BXD2 to BXD2-Rag2-/- mice resulted in retention of B cells in the MZ, leading to repopulation of MZMs. Consistent with this, there was no age-related loss of MZMs in the spleens of BXD2-Ifnar-/- mice. Reconstitution of BXD2-Rag2-/- with a mixed bone marrow of BXD2-GFP+Ifnar+ and BXD2-GFP-Ifnar- (1:1) resulted in a 50:50 repopulation of donor MZMs in recipient mice, thereby excluding the direct effect of type I IFNs on MZM maintenance. Our present study suggests that one novel mechanism associated with type I IFN-induced autoimmunity is related to the effects on promoting follicular migration of LT expressing B cells. The dissociation of LT+ B cells and MZMs disrupts MZ barrier and enables follicular entry of apoptotic antigens.

Attenuated Inflammatory Response in Triggering Receptor Expressed on Myeloid Cells 2 (TREM2) Knock-Out Mice following Stroke

BackgroundTriggering receptor expressed on myeloid cells-2 (TREM2) is a microglial surface receptor involved in phagocytosis. Clearance of apoptotic debris after stroke represents an important mechanism to re-attain tissue homeostasis and thereby ensure functional recovery. The role of TREM2 following stroke is currently unclear.Methods and ResultsAs an experimental stroke model, the middle cerebral artery of mice was occluded for 30 minutes with a range of reperfusion times (duration of reperfusion: 6 h/12 h/24 h/2 d/7 d/28 d). Quantitative PCR (qPCR) revealed a greatly increased transcription of TREM2 after stroke. We subsequently analyzed the expression of pro-inflammatory cytokines, chemokines and their receptors in TREM2-knockout (TREM2-KO) mice via qPCR. Microglial activation (CD68, Iba1) and CD3-positive T-cell invasion were analyzed via qPCR and immunohistochemistry. Functional consequences of TREM2 knockout were assessed by infarct volumetry. The acute inflammatory response (12 h reperfusion) was very similar between TREM2-KO mice and their littermate controls. However, in the sub-acute phase (7 d reperfusion) following stroke, TREM2-KO mice showed a decreased transcription of pro-inflammatory cytokines TNFα, IL-1α and IL-1β, associated with a reduced microglial activity (CD68, Iba1). Furthermore, TREM2-KO mice showed a reduced transcription of chemokines CCL2 (MCP1), CCL3 (MIP1α) and the chemokine receptor CX3CR1, followed by a diminished invasion of CD3-positive T-cells. No effect on the lesion size was observed.ConclusionsAlthough we initially expected an exaggerated pro-inflammatory response following ablation of TREM2, our data support a contradictory scenario that the sub-acute inflammatory reaction after stroke is attenuated in TREM2-KO mice. We therefore conclude that TREM2 appears to sustain a distinct inflammatory response after stroke.

Macrophage scavenger receptor 1 (Msr1) promotes autoantibody-mediated arthritis by impairing B cell tolerance (171.18)

Abstract Peripheral tolerance is achieved through several mechanisms, including induction of the hyporesponsive state of anergy in potentially autoreactive lymphocytes. The class A macrophage scavenger receptor, Msr1, is reported to participate in the maintenance of immunological tolerance by performing functions such as the clearance of apoptotic debris and the tonic sampling of self-antigens. We evaluated the role of Msr1 in a mouse model of autoantibody-dependent arthritis. Genetic deficiency of Msr1 in the K/BxN TCR transgenic model (Msr1-/- K/BxN) decreased the incidence and severity of arthritis. However, arthritis induced by transfer of arthritogenic autoantibodies was unimpaired in naïve mice lacking Msr1, suggesting that Msr1 deficiency in K/BxN mice affected the pathogenic events preceding autoantibody synthesis. Indeed, antibodies against the key autoantigen in this model, glucose 6-phosphate isomerase (GPI), were undetectable in Msr1-/- K/BxN mice, despite the presence of an expanded population of GPI-specific B cells. We did not observe any defect in T cell numbers or function. We hypothesized that Msr1 might control the availability of autoantigen and found that Msr1-/- macrophages displayed impaired uptake of GPI. Taken together, these data suggest a model in which deficiency of Msr1 can lead to a higher concentration of soluble autoantigens, resulting in B cell anergy and protection from autoimmune disease.

OP04 Circulating levels of the long pentraxin PTX3, but not hepatocyte derived C-reactive protein, correlate with severity following human acute liver injury

IntroductionThe innate immune response may underpin the development of multiorgan failure following acute liver injury, particularly following paracetamol overdose (POD). Pentraxin 3 (PTX3) is a long pentraxin induced by interleukin...

The liaison between apoptotic cells and macrophages – the end programs the beginning

The efficient execution of apoptotic cell death with the clearance of apoptotic debris by phagocytes is a key regulatory mechanism ensuring tissue homeostasis. Failure in this execution program contributes to the pathogenesis of many human diseases. In this review, we describe the current knowledge regarding the interaction of apoptotic cells with their professional 'captors', the macrophages, with special emphasis on the immunological outcome. Removal of apoptotic cells must be considered as a process that actively delivers signals to polarize macrophages, which are fundamental for the resolution of inflammation. However, the sculpting of macrophage responses by apoptotic cells can be misused under certain inflammatory disease conditions, including tumor development.

Macrophages Produce TGF-β-Induced (β-ig-h3) following Ingestion of Apoptotic Cells and Regulate MMP14 Levels and Collagen Turnover in Fibroblasts

Phagocytic clearance of apoptotic cells by macrophages is an essential part in the resolution of inflammation. It coincides with activation of repair mechanisms, including accumulation of extracellular matrix. A possible link between clearance of apoptotic debris and accumulation of extracellular matrix has not been investigated. Production of collagen was measured in primary fibroblasts cocultured with macrophages. Ingestion of apoptotic cells by monocyte-derived macrophages led to up-regulation of collagen. Direct contact between macrophages and fibroblasts was not required for collagen up-regulation. Macrophages produced TGF-beta following ingestion of apoptotic cells, but the levels of this cytokine were lower than those required for a significant up-regulation of collagen. Simultaneously, the levels of TGF-beta-induced (TGFBI), or keratoepithelin/BIGH3, mRNA and protein were increased. In contrast, primary alveolar macrophages stimulated collagen production without exposure to apoptotic cells; there was no further increase in the levels of TGFBI, mRNA or protein, or collagen after ingestion of apoptotic cells. Stimulation of fibroblasts with TGFBI down-regulated MMP14 levels, decreased DNA binding by p53, increased DNA binding by PU.1, and up-regulated collagen protein but not mRNA levels. Overexpression of MMP14 or p53, or small interfering RNA-mediated inhibition of PU.1 led to an increase in MMP14 and a decline in collagen levels, whereas small interfering RNA-mediated inhibition of MMP14 led to elevation of collagen levels. In conclusion, monocyte-derived but not alveolar macrophages produce TGFBI following ingestion of apoptotic cells, leading to the down-regulation of MMP14 levels in fibroblasts through a mechanism involving p53 and PU.1, and to subsequent accumulation of collagen.

Tolling for Autoimmunity—Prime Time for 7

The reason certain self-antigens are consistently targeted by autoantibodies may be because they are self-adjuvants. Two papers in this issue of Immunity provide important insights into the contribution of Toll-like receptors in systemic autoimmune disease (Berland et al., 2006; Christensen et al., 2006).

The origin and pathogenic consequences of anti-dsDNA antibodies in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a multisystem autoimmune rheumatic disease characterized by the presence of autoantibodies, many of which are directed against antigens of nuclear origin. In the clinical setting, these antibodies are important in the management of patients with SLE, providing information about disease activity, subtype and prognosis. The clearance of apoptotic debris is defective in patients with SLE. This causes the inappropriate persistence of nuclear antigens and thus a breakdown in peripheral tolerance with ensuing autoantibody generation. Patients have a variety of self-directed antibodies, and it is increasingly clear that some, but not all, of these antibodies are directly pathogenic. This review discusses the current hypotheses regarding the features by which antibody pathogenicity may be determined and therefore predicted, as well as the mechanisms by which these autoantibodies cause tissue damage in SLE and in lupus nephritis in particular. Identifying these pathogenic antibodies may well hold the key to the development of new, targeted therapies.

Macrophages, Multinucleated Giant Cells, and Apoptosis in HIV+Patients and Normal Blood Donors

Clearance of apoptotic debris is carried out by cells of the monocyte/macrophage lineage and, as other macrophage functions, it can be altered in AIDS, leading to the accumulation of apoptotic cells observed in this disease. In this study we evaluated the ability of macrophages from human immunodeficiency virus (HIV)-infected patients to differentiate and to clear apoptotic debris in prolongedin vitrocultures. Peripheral blood mononuclear cells (PBMC) from infected hemophilia patients were cultured in the absence of exogenously added stimulators and the organization and morphological characteristics of the cultures were analyzed and correlated with clinical staging of the patients. Cell aggregates of different sizes involving macrophages and lymphocytes were formed in cultures from asymptomatic HIV+patients (CDC groups II–III) and controls and in 4/7 group IV C2 HIV+patients. In order to obtain viable and organized cultures, cells had to be handled carefully, allowing contact and undisturbed sedimentation in round-bottom tubes. Multinucleated giant cells (MGC) were formed through macrophage fusion after 5 days of culture in HIV−controls, group II and III patients, and some of the group IV C2 patients, while scarce formation of MGC was observed in AIDS patients or patients with advanced HIV disease. This paucity was correlated with impaired dead cell removal and accumulation of apoptotic debris. Viability of macrophages and MGC was reduced after 15 days. MGC and the macrophages (either free or in cell aggregates) were able to remove dead cells, clearing the cultures of cell debris. Furthermore, in group II and III HIV+hemophilic patients, increased macrophage–MGC phagocytic activity, suggestingin vivoactivation, was frequently observed. In HIV+patients with AIDS or advanced HIV disease (CDC groups IV A, IV C1, and IV D) dead cell removal was impaired and apoptotic debris accumulated. Long-term cultures of unstimulated PBMC are an interesting model for studying the role of macrophages and/or MGC in the removal of dead cells as well as examining the cellular milieu in which HIV replicates in an individual host.