Amanda J. Lanser

Identification of a unique TGF-β–dependent molecular and functional signature in microglia

Microglia are myeloid cells of the CNS that participate both in normal CNS function and in disease. We investigated the molecular signature of microglia and identified 239 genes and 8 microRNAs that were uniquely or highly expressed in microglia versus myeloid and other immune cells. Of the 239 genes, 106 were enriched in microglia as compared with astrocytes, oligodendrocytes and neurons. This microglia signature was not observed in microglial lines or in monocytes recruited to the CNS, and was also observed in human microglia. We found that TGF-β was required for the in vitro development of microglia that express the microglial molecular signature characteristic of adult microglia and that microglia were absent in the CNS of TGF-β1–deficient mice. Our results identify a unique microglial signature that is dependent on TGF-β signaling and provide insights into microglial biology and the possibility of targeting microglia for the treatment of CNS disease.

Modulating inflammatory monocytes with a unique microRNA gene signature ameliorates murine ALS

Amyotrophic lateral sclerosis (ALS) is a progressive disease associated with neuronal cell death that is thought to involve aberrant immune responses. Here we investigated the role of innate immunity in a mouse model of ALS. We found that inflammatory monocytes were activated and that their progressive recruitment to the spinal cord, but not brain, correlated with neuronal loss. We also found a decrease in resident microglia in the spinal cord with disease progression. Prior to disease onset, splenic Ly6Chi monocytes expressed a polarized macrophage phenotype (M1 signature), which included increased levels of chemokine receptor CCR2. As disease onset neared, microglia expressed increased CCL2 and other chemotaxis-associated molecules, which led to the recruitment of monocytes to the CNS by spinal cord-derived microglia. Treatment with anti-Ly6C mAb modulated the Ly6Chi monocyte cytokine profile, reduced monocyte recruitment to the spinal cord, diminished neuronal loss, and extended survival. In humans with ALS, the analogous monocytes (CD14+CD16-) exhibited an ALS-specific microRNA inflammatory signature similar to that observed in the ALS mouse model, linking the animal model and the human disease. Thus, the profile of monocytes in ALS patients may serve as a biomarker for disease stage or progression. Our results suggest that recruitment of inflammatory monocytes plays an important role in disease progression and that modulation of these cells is a potential therapeutic approach.

Dampening of death pathways by schnurri-2 is essential for T-cell development

Generation of a diverse and self-tolerant T-cell repertoire requires appropriate interpretation of T-cell antigen receptor (TCR) signals by CD4+ CD8+ double-positive thymocytes. Thymocyte cell fate is dictated by the nature of TCR–major-histocompatibility-complex (MHC)–peptide interactions, with signals of higher strength leading to death (negative selection) and signals of intermediate strength leading to differentiation (positive selection). Molecules that regulate T-cell development by modulating TCR signal strength have been described but components that specifically define the boundaries between positive and negative selection remain unknown. Here we show in mice that repression of TCR-induced death pathways is critical for proper interpretation of positive selecting signals in vivo, and identify schnurri-2 (Shn2; also known as Hivep2) as a crucial death dampener. Our results indicate that Shn2−/− double-positive thymocytes inappropriately undergo negative selection in response to positive selecting signals, thus leading to disrupted T-cell development. Shn2−/− double-positive thymocytes are more sensitive to TCR-induced death in vitro and die in response to positive selection interactions in vivo. However, Shn2-deficient thymocytes can be positively selected when TCR-induced death is genetically ablated. Shn2 levels increase after TCR stimulation, indicating that integration of multiple TCR–MHC–peptide interactions may fine-tune the death threshold. Mechanistically, Shn2 functions downstream of TCR proximal signalling compenents to dampen Bax activation and the mitochondrial death pathway. Our findings uncover a critical regulator of T-cell development that controls the balance between death and differentiation.

Targeting latency-associated peptide promotes antitumor immunity

Anti-LAP antibody induces antitumor immunity by affecting both adaptive and innate immune mechanisms. LAPping up tumor immunoregulation Tumors dodge the immune system in part by promoting immune regulatory cells. Gabriely et al. now report that antibodies to latency-associated peptide (LAP), which forms a complex with transforming growth factor–β (TGF-β), reduced tumor growth in multiple cancer models in mice. The authors found that antibodies to LAP decreased numbers of LAP+ regulatory T cells and tolerogenic dendritic cells within the tumor and TGF-β secretion in vitro. Moreover, anti-LAP antibodies decreased numbers of CD103+ CD8+ T cells in lymphoid organs; these cells were then shown to promote tumor growth. Furthermore, combining LAP antibodies with antigen-specific vaccination enhanced both antitumor immune response and immunological memory. Together, these data suggest that targeting LAP may enhance tumor immunotherapy. Regulatory T cells (Tregs) promote cancer by suppressing antitumor immune responses. We found that anti-LAP antibody, which targets the latency-associated peptide (LAP)/transforming growth factor–β (TGF-β) complex on Tregs and other cells, enhances antitumor immune responses and reduces tumor growth in models of melanoma, colorectal carcinoma, and glioblastoma. Anti-LAP decreases LAP+ Tregs, tolerogenic dendritic cells, and TGF-β secretion and is associated with CD8+ T cell activation. Anti-LAP increases infiltration of tumors by cytotoxic CD8+ T cells and reduces CD103+ CD8 T cells in draining lymph nodes and the spleen. We identified a role for CD103+ CD8 T cells in cancer. Tumor-associated CD103+ CD8 T cells have a tolerogenic phenotype with increased expression of CTLA-4 and interleukin-10 and decreased expression of interferon-γ, tumor necrosis factor–α, and granzymes. Adoptive transfer of CD103+ CD8 T cells promotes tumor growth, whereas CD103 blockade limits tumorigenesis. Thus, anti-LAP targets multiple immunoregulatory pathways and represents a potential approach for cancer immunotherapy.

Disruption of the ATP/adenosine balance in CD39−/− mice is associated with handling-induced seizures

Seizures are due to excessive, synchronous neuronal firing in the brain and are characteristic of epilepsy, the fourth most prevalent neurological disease. We report handling‐induced and spontaneous seizures in mice deficient for CD39, a cell‐surface ATPase highly expressed on microglial cells. CD39−/− mice with handling‐induced seizures had normal input–output curves and paired‐pulse ratio measured from hippocampal slices and lacked microgliosis, astrogliosis or overt cell loss in the hippocampus and cortex. As expected, however, the cerebrospinal fluid of CD39−/− mice contained increased levels of ATP and decreased levels of adenosine. To determine if immune activation was involved in seizure progression, we challenged mice with lipopolysaccharide (LPS) and measured the effect on microglia activation and seizure severity. Systemic LPS challenge resulted in increased cortical staining of Iba1/CD68 and gene array data from purified microglia predicted increased expression of interleukin‐8, triggering receptor expressed on myeloid cells 1, p38, pattern recognition receptors, death receptor, nuclear factor‐κB , complement, acute phase, and interleukin‐6 signalling pathways in CD39−/− versus CD39+/+ mice. However, LPS treatment did not affect handling‐induced seizures. In addition, microglia‐specific CD39 deletion in adult mice was not sufficient to cause seizures, suggesting instead that altered expression of CD39 during development or on non‐microglial cells such as vascular endothelial cells may promote the seizure phenotype. In summary, we show a correlation between altered extracellular ATP/adenosine ratio and a previously unreported seizure phenotype in CD39−/− mice. This work provides groundwork for further elucidation of the underlying mechanisms of epilepsy.

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Corrigendum: Identification of a unique TGF-β–dependent molecular and functional signature in microglia

Corrigendum: Identification of a unique TGF-β–dependent molecular and functional signature in microglia

Acute microglia ablation induces neurodegeneration in the somatosensory system

Previous studies have reported that microglia depletion leads to impairment of synapse formation and these cells rapidly repopulate from CNS progenitors. However, the impact of microglia depletion and repopulation in the long-term state of the CNS environment has not been characterized. Here, we report that acute and synchronous microglia depletion and subsequent repopulation induces gray matter microgliosis, neuronal death in the somatosensory cortex and ataxia-like behavior. We find a type 1 interferon inflammatory signature in degenerating somatosensory cortex from microglia-depleted mice. Transcriptomic and mass cytometry analysis of repopulated microglia demonstrates an interferon regulatory factor 7-driven activation state. Minocycline and anti-IFNAR1 antibody treatment attenuate the CNS type 1 interferon-driven inflammation, restore microglia homeostasis and reduce ataxic behavior. Neither microglia depletion nor repopulation impact neuropathology or T-cell responses during experimental autoimmune encephalomyelitis. Together, we found that acute microglia ablation induces a type 1 interferon activation state of gray matter microglia associated with acute neurodegeneration.Previous studies have shown that depletion of microglia at early developmental stages leads to neuronal death. Here the authors use an inducible system to ablate microglia in adulthood, showing that such depletion leads to ataxia-like behavior and neuronal loss, and identifying the inflammatory components that may contribute.

γδ T cells control humoral immune response by inducing T follicular helper cell differentiation

Abstractγδ T cells have many known functions, including the regulation of antibody responses. However, how γδ T cells control humoral immunity remains elusive. Here we show that complete Freund’s adjuvant (CFA), but not alum, immunization induces a subpopulation of CXCR5-expressing γδ T cells in the draining lymph nodes. TCRγδ+CXCR5+ cells present antigens to, and induce CXCR5 on, CD4 T cells by releasing Wnt ligands to initiate the T follicular helper (Tfh) cell program. Accordingly, TCRδ−/− mice have impaired germinal center formation, inefficient Tfh cell differentiation, and reduced serum levels of chicken ovalbumin (OVA)-specific antibodies after CFA/OVA immunization. In a mouse model of lupus, TCRδ−/− mice develop milder glomerulonephritis, consistent with decreased serum levels of lupus-related autoantibodies, when compared with wild type mice. Thus, modulation of the γδ T cell-dependent humoral immune response may provide a novel therapy approach for the treatment of antibody-mediated autoimmunity.Many immune functions have been reported for γδ T cells, including the regulation of antibody responses. Here the authors show that CXCR5+ γδ T cells release Wnt ligands to initiate the T follicular helper cell differentiation program and promote antibody production.

Erratum: Identification of a unique TGF-β-dependent molecular and functional signature in microglia (Nature Neuroscience (2014) 17 (131-143))

Corrigendum: Identification of a unique TGF-β–dependent molecular and functional signature in microglia