USP18 lack in microglia causes destructive interferonopathy of the mouse brain.

Microglia are tissue macrophages of the central nervous system (CNS) that control tissue homeostasis. Microglia dysregulation is thought to be causal for a group of neuropsychiatric, neurodegenerative and neuroinflammatory diseases, called "microgliopathies". However, how the intracellular stimulation machinery in microglia is controlled is poorly understood. Here, we identified the ubiquitin-specific protease (Usp) 18 in white matter microglia that essentially contributes to microglial quiescence. We further found that microglial Usp18 negatively regulates the activation of Stat1 and concomitant induction of interferon-induced genes, thereby terminating IFN signaling. The Usp18-mediated control was independent from its catalytic activity but instead required the interaction with Ifnar2. Additionally, the absence of Ifnar1 restored microglial activation, indicating a tonic IFN signal which needs to be negatively controlled by Usp18 under non-diseased conditions. These results identify Usp18 as a critical negative regulator of microglia activation and demonstrate a protective role of Usp18 for microglia function by regulating the Ifnar pathway. The findings establish Usp18 as a new molecule preventing destructive microgliopathy.

Loss of Trex1 in Dendritic Cells Is Sufficient To Trigger Systemic Autoimmunity.

Defects of the intracellular enzyme 3' repair exonuclease 1 (Trex1) cause the rare autoimmune condition Aicardi-Goutières syndrome and are associated with systemic lupus erythematosus. Trex1(-/-) mice develop type I IFN-driven autoimmunity, resulting from activation of the cytoplasmic DNA sensor cyclic GMP-AMP synthase by a nucleic acid substrate of Trex1 that remains unknown. To identify cell types responsible for initiation of autoimmunity, we generated conditional Trex1 knockout mice. Loss of Trex1 in dendritic cells was sufficient to cause IFN release and autoimmunity, whereas Trex1-deficient keratinocytes and microglia produced IFN but did not induce inflammation. In contrast, B cells, cardiomyocytes, neurons, and astrocytes did not show any detectable response to the inactivation of Trex1. Thus, individual cell types differentially respond to the loss of Trex1, and Trex1 expression in dendritic cells is essential to prevent breakdown of self-tolerance ensuing from aberrant detection of endogenous DNA.

Lymphotoxin-dependent prion replication in inflammatory stromal cells of granulomas.

Prior to invading the nervous system, prions frequently colonize lymphoid organs and sites of inflammatory lymphoneogenesis, where they colocalize with Mfge8+ follicular dendritic cells (FDCs). Here, we report that soft-tissue granulomas, a frequent feature of chronic inflammation, expressed the cellular prion protein (PrPC, encoded by Prnp) and the lymphotoxin receptor (LTbetaR), even though they lacked FDCs and did not display lymphoneogenesis. After intraperitoneal prion inoculation, granulomas of Prnp(+/+) mice, but not Prnp(-/-) granulomas or unaffected Prnp(+/+) skin, accumulated prion infectivity and disease-associated prion protein. Bone-marrow transfers between Prnp(+/+) and Prnp(-/-) mice and administration of lymphotoxin signaling antagonists indicated that prion replication required radioresistant PrPC-expressing cells and LTbetaR signaling. Granulomatous PrPC was mainly expressed by stromal LTbetaR+ mesenchymal cells that were absent from unaffected subcutis. Hence, granulomas can act as clinically silent reservoirs of prion infectivity. Furthermore, lymphotoxin-dependent prion replication can occur in inflammatory stromal cells that are distinct from FDCs.

Irregular Warfare Must Combine Good Medicine, with Both Good Tactics and Good Strategies: Position Paper by the French Special Operations Forces Medical Command.

INTRODUCTION: Military operations are no longer limited to the application of counterterrorism and counterinsurgency strategies; they are now characterized by hybrid, irregular, and unconventional features. While some authors have indicated the need for medical support to adapt to these new modes of military operations, they have focused mainly on the tactical level of care on the battlefield. As Sun Tzu states, "Strategy without tactics is the slowest route to victory. Tactics without strategy is the noise before defeat," further proposals are still needed on the application of both medical tactics and medical strategies in irregular warfare. METHODS: Medical experts from the French Special Operations Forces (SOF) Medical Command have identified specific medical challenges that special operations face in the context of the current transformation of armed confrontations into irregular warfare. RESULTS: This position paper presents original tactical medical proposals for improving medical support in irregular warfare, ranging from the definition of a Primary-Alternate-Contingency-Emergency medical plan to the promotion of telemedical support. Original strategic medical proposals have highlighted the importance of recognizing medical issues in irregular warfare, including the medical actions carried out through and with local partners and the multiple approaches to countering medical threats. CONCLUSIONS: The SOF medical community must be closely involved with and facilitate the responses to the shift to irregular warfare. International collaboration and interoperability are more necessary than ever, as they will enable a more effective combination of good medicine with both good tactics and good strategies. These perspectives can also be extended to improve medical care in the conventional armed forces and austere civilian settings. LEVEL OF EVIDENCE: N/A. STUDY TYPE: Original research.

IkappaB kinase 2 determines oligodendrocyte loss by non-cell-autonomous activation of NF-kappaB in the central nervous system.

The IκB kinase complex induces nuclear factor kappa B activation and has recently been recognized as a key player of autoimmunity in the central nervous system. Notably, IκB kinase/nuclear factor kappa B signalling regulates peripheral myelin formation by Schwann cells, however, its role in myelin formation in the central nervous system during health and disease is largely unknown. Surprisingly, we found that brain-specific IκB kinase 2 expression is dispensable for proper myelin assembly and repair in the central nervous system, but instead plays a fundamental role for the loss of myelin in the cuprizone model. During toxic demyelination, inhibition of nuclear factor kappa B activation by conditional ablation of IκB kinase 2 resulted in strong preservation of central nervous system myelin, reduced expression of proinflammatory mediators and a significantly attenuated glial response. Importantly, IκB kinase 2 depletion in astrocytes, but not in oligodendrocytes, was sufficient to protect mice from myelin loss. Our results reveal a crucial role of glial cell-specific IκB kinase 2/nuclear factor kappa B signalling for oligodendrocyte damage during toxic demyelination. Thus, therapies targeting IκB kinase 2 function in non-neuronal cells may represent a promising strategy for the treatment of distinct demyelinating central nervous system diseases.

Overexpression of lymphotoxin in T cells induces fulminant thymic involution.

Activated lymphocytes and lymphoid-tissue inducer cells express lymphotoxins (LTs), which are essential for the organogenesis and maintenance of lymphoreticular microenvironments. Here we describe that T-cell-restricted overexpression of LT induces fulminant thymic involution. This phenotype was prevented by ablation of the LT receptors tumor necrosis factor receptor (TNFR) 1 or LT beta receptor (LTbetaR), representing two non-redundant pathways. Multiple lines of transgenic Ltalphabeta and Ltalpha mice show such a phenotype, which was not observed on overexpression of LTbeta alone. Reciprocal bone marrow transfers between LT-overexpressing and receptor-ablated mice show that involution was not due to a T cell-autonomous defect but was triggered by TNFR1 and LTbetaR signaling to radioresistant stromal cells. Thymic involution was partially prevented by the removal of one allele of LTbetaR but not of TNFR1, establishing a hierarchy in these signaling events. Infection with the lymphocytic choriomeningitis virus triggered a similar thymic pathology in wt, but not in Tnfr1(-/-) mice. These mice displayed elevated TNFalpha in both thymus and plasma, as well as increased LTs on both CD8(+) and CD4(-)CD8(-) thymocytes. These findings suggest that enhanced T cell-derived LT expression helps to control the physiological size of the thymic stroma and accelerates its involution via TNFR1/LTbetaR signaling in pathological conditions and possibly also in normal aging.

Early and rapid engraftment of bone marrow-derived microglia in scrapie.

Prion neuroinvasion is accompanied by maximal activation of microglia, the significance of which for pathogenesis is unknown. Here, we used bone marrow (BM) cells expressing GFP (green fluorescent protein) to study the turnover of microglia in mouse scrapie. We found that >or=50% of all brain microglia were replaced by BM-derived cells before clinical disease onset. In terminally sick mice, microglia density increased threefold to fourfold. Hence BM-derived microglia rapidly and efficaciously colonize the brain in scrapie. Whereas reconstitution of wild-type mice with prion protein-deficient (Prnp(o/o)) BM did not alter scrapie pathogenesis, Prnp(o/o) mice transplanted with wild-type BM cells were resistant to peripherally administered prions despite high levels of infectivity in the spleen. Cerebellar homogenates from prion-inoculated Prnp(o/o) mice reconstituted with >10% of wild-type microglia failed to infect transgenic mice overexpressing the cellular prion protein. Hence, in contrast to previous reports, microglia are not competent for efficient prion transport and replication in vivo.

Germinal center B cells are dispensable in prion transport and neuroinvasion.

Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative diseases of animals and humans. Many TSEs are initiated by prion replication in the lymphoreticular system (LRS). The cellular and molecular prerequisites for prion trafficking within the LRS are not fully understood. Here we have manipulated CD40 and its ligand to investigate whether genetic or pharmacological ablation of germinal center B cells (GCBs), which migrate into and out of germinal centers, influences prion pathogenesis. In contrast to previous reports, no alteration of prion pathogenesis was detected in mice lacking CD40L and in mice treated with anti-CD40L antibodies. These results suggest that GCBs alone do not impact peripheral splenic prion transport, replication efficiency, or neuroinvasion, and point to other mechanisms affecting prion transport from lymphoreticular sites of replication to the nervous system.

The parallel universe: microRNAs and their role in chronic hepatitis, liver tissue damage and hepatocarcinogenesis.

In recent years, enormous progress has been made in identifying microRNAs (miRNAs) as important regulators of gene expression and their association with or control of various liver diseases such as fibrosis, hepatitis and hepatocellular carcinoma (HCC). Indeed, many genes encoding miRNAs as well as their targets have been described and their direct or indirect link to the respective liver diseases has been investigated in various experimental systems as well as in human tissue. Here we discuss current knowledge of miRNAs and their involvement in liver diseases, elaborating in particular on the contribution of miRNAs to hepatitis, fibrosis and HCC formation. We also debate possible prognostic, predictive and therapeutic values of respective miRNAs in liver diseases. The discovery of liver disease related miRNAs has constituted a major breakthrough in liver research and will most likely be of high relevance for future therapeutic strategies, especially when dealing with hepatitis, fibrosis and HCC.

TAK1 suppresses a NEMO-dependent but NF-kappaB-independent pathway to liver cancer.

The MAP3-kinase TGF-beta-activated kinase 1 (TAK1) critically modulates innate and adaptive immune responses and connects cytokine stimulation with activation of inflammatory signaling pathways. Here, we report that conditional ablation of TAK1 in liver parenchymal cells (hepatocytes and cholangiocytes) causes hepatocyte dysplasia and early-onset hepatocarcinogenesis, coinciding with biliary ductopenia and cholestasis. TAK1-mediated cancer suppression is exerted through activating NF-kappaB in response to tumor necrosis factor (TNF) and through preventing Caspase-3-dependent hepatocyte and cholangiocyte apoptosis. Moreover, TAK1 suppresses a procarcinogenic and pronecrotic pathway, which depends on NF-kappaB-independent functions of the I kappaB-kinase (IKK)-subunit NF-kappaB essential modulator (NEMO). Therefore, TAK1 serves as a gatekeeper for a protumorigenic, NF-kappaB-independent function of NEMO in parenchymal liver cells.

A lymphotoxin-driven pathway to hepatocellular carcinoma.

Hepatitis B and C viruses (HBV and HCV) cause chronic hepatitis and hepatocellular carcinoma (HCC) by poorly understood mechanisms. We show that cytokines lymphotoxin (LT) alpha and beta and their receptor (LTbetaR) are upregulated in HBV- or HCV-induced hepatitis and HCC. Liver-specific LTalphabeta expression in mice induces liver inflammation and HCC, causally linking hepatic LT overexpression to hepatitis and HCC. Development of HCC, composed in part of A6(+) oval cells, depends on lymphocytes and IKappa B kinase beta expressed by hepatocytes but is independent of TNFR1. In vivo LTbetaR stimulation implicates hepatocytes as the major LT-responsive liver cells, and LTbetaR inhibition in LTalphabeta-transgenic mice with hepatitis suppresses HCC formation. Thus, sustained LT signaling represents a pathway involved in hepatitis-induced HCC.

Stromal complement receptor CD21/35 facilitates lymphoid prion colonization and pathogenesis.

We have studied the role of CD21/35, which bind derivatives of complement factors C3 and C4, in extraneural prion replication and neuroinvasion. Upon administration of small prion inocula, CD21/35(-/-) mice experienced lower attack rates and delayed disease over both wild-type (WT) mice and mice with combined C3 and C4 deficiencies. Early after inoculation, CD21/35(-/-) spleens were devoid of infectivity. Reciprocal adoptive bone marrow transfers between WT and CD21/35(-/-) mice revealed that protection from prion infection resulted from ablation of stromal, but not hemopoietic, CD21/35. Further adoptive transfer experiments between WT mice and mice devoid of both the cellular prion protein PrP(C) and CD21/35 showed that splenic retention of inoculum depended on stromal CD21/35 expression. Because both PrP(C) and CD21/35 are highly expressed on follicular dendritic cells, CD21/35 appears to be involved in targeting prions to follicular dendritic cells and expediting neuroinvasion following peripheral exposure to prions.

Expression of lymphotoxin beta governs immunity at two distinct levels.

Interaction of lymphotoxin alpha(1)beta(2) (LTalpha(1)beta(2)) with its receptor is key for the generation and maintenance of secondary lymphoid organ microstructure. We used mice conditionally deficient for LTbeta on different lymphocyte subsets to determine how the LTbeta-dependent lymphoid structure influences immune reactivity. All conditionally LTbeta-deficient mice mounted normal immune responses against vesicular stomatitis virus (VSV), and were protected against lymphocytic choriomeningitis virus (LCMV). In contrast, they exhibited reduced immune responses against non-replicating antigens. Completely LTbeta-deficient mice failed to retain VSV in the marginal zone and died from VSV infections, and they became virus carriers following infection with the non-cytopathic LCMV, which was correlated with defective virus replication in dendritic cells. It was ruled out that LTbeta expression on lymphocytes influenced their activation, homing capacity, or maturation. We therefore conclude that LTbeta expression influences immune reactivity at two distinct levels: (i) Expression of LTbeta on lymphocytes enhances the induction of immune responses against limiting amounts of antigen. (ii) Expression of LTbeta on non-lymphocytes governs antiviral immunity by enhancing antigen presentation on antigen-presenting cells. This prevents cytotoxic T lymphocytes exhaustion or death of the host by uncontrolled virus spread.

Coincident scrapie infection and nephritis lead to urinary prion excretion.

Prion infectivity is typically restricted to the central nervous and lymphatic systems of infected hosts, but chronic inflammation can expand the distribution of prions. We tested whether chronic inflammatory kidney disorders would trigger excretion of prion infectivity into urine. Urinary proteins from scrapie-infected mice with lymphocytic nephritis induced scrapie upon inoculation into noninfected indicator mice. Prionuria was found in presymptomatic scrapie-infected and in sick mice, whereas neither prionuria nor urinary PrP(Sc) was detectable in prion-infected wild-type or PrP(C)-overexpressing mice, or in nephritic mice inoculated with noninfectious brain. Thus, urine may provide a vector for horizontal prion transmission, and inflammation of excretory organs may influence prion spread.

Chronic lymphocytic inflammation specifies the organ tropism of prions.

Prions typically accumulate in nervous and lymphoid tissues. Because proinflammatory cytokines and immune cells are required for lymphoid prion replication, we tested whether inflammatory conditions affect prion pathogenesis. We administered prions to mice with five inflammatory diseases of the kidney, pancreas, or liver. In all cases, chronic lymphocytic inflammation enabled prion accumulation in otherwise prion-free organs. Inflammatory foci consistently correlated with lymphotoxin up-regulation and ectopic induction of FDC-M1+ cells expressing the normal cellular prion protein PrPC. By contrast, inflamed organs of mice lacking lymphotoxin-alpha or its receptor did not accumulate the abnormal isoform PrPSc, nor did they display infectivity upon prion inoculation. By expanding the tissue distribution of prions, chronic inflammatory conditions may act as modifiers of natural and iatrogenic prion transmission.

NF kappa B controls the balance between Fas and tumor necrosis factor cell death pathways during T cell receptor-induced apoptosis via the expression of its target gene A20.

Activation-induced cell death (AICD), a term originally coined for the anti-CD3-induced apoptosis of T cell hybridomas and thymocytes, is predominantly driven by death receptors and has been involved in the control of autoreactive T cells in the periphery. In the Do-11.10 T cell hybridoma model of AICD, activation of the T cell receptor (TCR) results in Fas-dependent apoptosis. Here, we show that inhibition of the transcription factor nuclear factor kappa B (NF kappa B) in Do-11.10 cells resulted in increased sensitivity to TCR-mediated apoptosis, correlating with defective induction of the anti-apoptotic NF kappa B target gene A20. Stable expression of the zinc finger protein A20 in NF kappa B-negative Do-11.10 cells rescued the phenotype. TCR activation in NF kappa B-deficient Do-11.10 cells resulted predominantly in tumor necrosis factor (TNF) receptor 2 (TNFR2)-dependent bystander cell death rather than classical Fas-dependent AICD. Strikingly, A20 blocked TNF-mediated apoptosis and simultaneously restored TCR-induced Fas-dependent AICD. In addition, NF kappa B downstream of TNFR was required for up-regulation of Fas expression by endogenous TNF secreted in response to TCR stimulation. Together, these results suggest that NF kappa B can play both pro- and anti-apoptotic roles during AICD. We propose that NF kappa B controls the balance between Fas and TNF cell death pathways during AICD via the expression of the zinc finger protein A20.