Stat5b/Ezh2 axis governs high PD-L1 expressing tolerogenic dendritic cell subset in autoimmune diabetes.

Dendritic cells (DCs) are specialized antigen-presenting cells that play an important role in inducing and maintaining immune tolerance. The altered distribution and/or function of DCs contributes to defective tolerance in autoimmune diseases such as type 1 diabetes (T1D). In human T1D and in NOD mouse models, DCs share some defects and are often described as less tolerogenic and excessively immunogenic. In the NOD mouse model, the autoimmune response is associated with a defect in the Stat5b signaling pathway. We have reported that expressing a constitutively active form of Stat5b in DCs of transgenic NOD mice (NOD.Stat5b-CA), re-established their tolerogenic function, restored autoimmune tolerance and conferred protection from diabetes. However, the role and molecular mechanisms of Stat5b signaling in regulating splenic conventional DCs tolerogenic signature remained unclear. In this study, we reported that, compared to immunogenic splenic DCs of NOD, splenic DCs of NOD.Stat5b-CA mice exhibited a tolerogenic profile marked by elevated PD-L1 and PD-L2 expression, reduced pro-inflammatory cytokine production, increased frequency of the cDC2 subset and decreased frequency of the cDC1 subset. This tolerogenic profile was associated with increased Ezh2 and IRF4 but decreased IRF8 expression. We also found an upregulation of PD-L1 in the cDC1 subset and high PD-L1 and PD-L2 expression in cDC2 of NOD.Stat5b-CA mice. Mechanistically, we demonstrated that Ezh2 plays an important role in the maintenance of high PD-L1 expression in cDC1 and cDC2 subsets and that Ezh2 inhibition resulted in PD-L1 but not PD-L2 downregulation which was more drastic in the cDC2 subset. Additionally, Ezh2 inhibition severely reduced the cDC2 subset and increased the cDC1 subset and Stat5b-CA.DC pro-inflammatory cytokine production. Together our data suggest that the Stat5b-Ezh2 axis is critical for the maintenance of tolerogenic high PD-L1-expressing cDC2 and autoimmune tolerance in NOD.Stat5b-CA mice.

Inhibition of PI3K/C/EBPß axis in tolerogenic bone marrow-derived dendritic cells of NOD mice promotes Th17 differentiation and diabetes development.

Dendritic cells (DCs) are key regulators of the adaptive immune response. Tolerogenic dendritic cells play a crucial role in inducing and maintaining immune tolerance in autoimmune diseases such as type 1 diabetes in humans as well as in the NOD mouse model. We previously reported that bone marrow-derived DCs (BM.DCs) from NOD mice, generated with a low dose of GM-CSF (GM/DCs), induce Treg differentiation and are able to protect NOD mice from diabetes. We had also found that the p38 MAPK/C/EBPß axis is involved in regulating the phenotype, as well as the production of IL-10 and IL-12p70, by tolerogenic GM/DCs. Here, we report that the inhibition of the PI3K signaling switched the cytokine profile of GM/DCs toward Th17-promoting cytokines without affecting their phenotype. PI3K inhibition abrogated the production of IL-10 by GM/DCs, whereas it enhanced their production of IL-23 and TGFß. Inhibition of PI3K signaling in tolerogenic GM/DCs also induced naive CD4+ T cells differentiation toward Th17 cells. Mechanistically, PI3K inhibition increased the DNA-binding activity of C/EBPß through a GSK3-dependent pathway, which is important to maintain the semimature phenotype of tolerogenic GM/DCs. Furthermore, analysis of C/EBPß-/- GM/DCs demonstrated that C/EBPß is required for IL-23 production. Of physiological relevance, the level of protection from diabetes following transfusion of GM/DCs into young NOD mice was significantly reduced when NOD mice were transfused with GM/DCs pretreated with a PI3K inhibitor. Our data suggest that PI3K/C/EBPß signaling is important in controlling tolerogenic function of GM/DCs by limiting their Th17-promoting cytokines.

Dendritic Cells and Their Immunotherapeutic Potential for Treating Type 1 Diabetes.

Type 1 diabetes (T1D) results from the destruction of pancreatic beta cells through a process that is primarily mediated by T cells. Emerging evidence suggests that dendritic cells (DCs) play a crucial role in initiating and developing this debilitating disease. DCs are professional antigen-presenting cells with the ability to integrate signals arising from tissue infection or injury that present processed antigens from these sites to naïve T cells in secondary lymphoid organs, thereby triggering naïve T cells to differentiate and modulate adaptive immune responses. Recent advancements in our knowledge of the various subsets of DCs and their cellular structures and methods of orchestration over time have resulted in a better understanding of how the T cell response is shaped. DCs employ various arsenal to maintain their tolerance, including the induction of effector T cell deletion or unresponsiveness and the generation and expansion of regulatory T cell populations. Therapies that suppress the immunogenic effects of dendritic cells by blocking T cell costimulatory pathways and proinflammatory cytokine production are currently being sought. Moreover, new strategies are being developed that can regulate DC differentiation and development and harness the tolerogenic capacity of these cells. Here, in this report, we focus on recent advances in the field of DC immunology and evaluate the prospects of DC-based therapeutic strategies to treat T1D.

Machine Learning in Modeling of Mouse Behavior.

Mouse behavior is a primary outcome in evaluations of therapeutic efficacy. Exhaustive, continuous, multiparametric behavioral phenotyping is a valuable tool for understanding the pathophysiological status of mouse brain diseases. Automated home cage behavior analysis produces highly granulated data both in terms of number of features and sampling frequency. Previously, we demonstrated several ways to reduce feature dimensionality. In this study, we propose novel approaches for analyzing 33-Hz data generated by CleverSys software. We hypothesized that behavioral patterns within short time windows are reflective of physiological state, and that computer modeling of mouse behavioral routines can serve as a predictive tool in classification tasks. To remove bias due to researcher decisions, our data flow is indifferent to the quality, value, and importance of any given feature in isolation. To classify day and night behavior, as an example application, we developed a data preprocessing flow and utilized logistic regression (LG), support vector machines (SVM), random forest (RF), and one-dimensional convolutional neural networks paired with long short-term memory deep neural networks (1DConvBiLSTM). We determined that a 5-min video clip is sufficient to classify mouse behavior with high accuracy. LG, SVM, and RF performed similarly, predicting mouse behavior with 85% accuracy, and combining the three algorithms in an ensemble procedure increased accuracy to 90%. The best performance was achieved by combining the 1DConv and BiLSTM algorithms yielding 96% accuracy. Our findings demonstrate that computer modeling of the home-cage ethome can clearly define mouse physiological state. Furthermore, we showed that continuous behavioral data can be analyzed using approaches similar to natural language processing. These data provide proof of concept for future research in diagnostics of complex pathophysiological changes that are accompanied by changes in behavioral profile.

Volumetric MRI demonstrates atrophy of the olfactory cortex in AD.

OBJECTIVE: Alzheimer disease (AD) is a chronic neurodegenerative disorder that affects millions of individuals worldwide. Symptoms include memory dysfunction and deficits in attention, planning, language, and overall cognitive function. Olfactory dysfunction is a common symptom of AD and evidence supports that it is an early marker. Furthermore, olfactory bulb and entorhinal cortex atrophy are well described in AD. However, in AD, no studies have assessed the olfactory cortex as a whole and if sex effects are observed. METHODS: Magnetic Resonance Imaging was used to scan 39 participants with an average age of 72 years and included men and women. AAL Single-Subject Atlas (implemented in PNEURO tool - PMOD 3.8) was used to determine the volume of the olfactory cortex and the hippocampus. Olfactory cortex volume was lower in both men and women AD cases compared with controls. This decrease was more apparent in the left olfactory cortex and was influenced by age. As expected, hippocampal volume was also significantly reduced in AD. However, this was only observed in the male cohort. A significant correlation was observed between levels of education and hippocampal volume in controls that were not detected in the AD participants. Asymmetry was observed in the olfactory cortex volume when comparing left and right volumes in both the control and AD participants, which was not observed in the hippocampus. RESULTS: These data highlight the importance of the role of olfactory cortical atrophy in the pathogenesis of AD and the interplay between the olfactory deficits and degeneration of olfactory regions in the brain.

Elucidating the Role of Ezh2 in Tolerogenic Function of NOD Bone Marrow-Derived Dendritic Cells Expressing Constitutively Active Stat5b.

Tolerogenic dendritic cells (toDCs) are crucial to controlling the development of autoreactive T cell responses and the prevention of autoimmunity. We have reported that NOD.CD11cStat5b-CA transgenic mice expressing a constitutively active (CA) form of Stat5b under the control of a CD11c promoter are protected from diabetes and that Stat5b-CA-expressing DCs are tolerogenic and halt ongoing diabetes in NOD mice. However, the molecular mechanisms by which Stat5b-CA modulates DC tolerogenic function are not fully understood. Here, we used bone marrow-derived DCs (BMDCs) from NOD.CD11cStat5b-CA transgenic mice (Stat5b-CA.BMDCs) and found that Stat5b-CA.BMDCs displayed high levels of MHC class II, CD80, CD86, PD-L1, and PD-L2 and produced elevated amounts of TGFß but low amounts of TNFα and IL-23. Stat5b-CA.BMDCs upregulated Irf4 and downregulated Irf8 genes and protein expression and promoted CD11c+CD11b+ DC2 subset differentiation. Interestingly, we found that the histone methyltransferase Ezh2 and Stat5b-CA bound gamma-interferon activated site (GAS) sequences in the Irf8 enhancer IRF8 transcription, whereas Stat5b but not Ezh2 bound GAS sequences in the Irf4 promoter to enhance IRF4 transcription. Injection of Stat5b-CA.BMDCs into prediabetic NOD mice halted progression of islet inflammation and protected against diabetes. Importantly, inhibition of Ezh2 in tolerogenic Stat5b-CA.BMDCs reduced their ability to prevent diabetes development in NOD recipient mice. Taken together, our data suggest that the active form of Stat5b induces tolerogenic DC function by modulating IRF4 and IRF8 expression through recruitment of Ezh2 and highlight the fundamental role of Ezh2 in Stat5b-mediated induction of tolerogenic DC function.

NLRX1 inhibits the early stages of CNS inflammation and prevents the onset of spontaneous autoimmunity.

Nucleotide-binding, leucine-rich repeat containing X1 (NLRX1) is a mitochondria-located innate immune sensor that inhibits major pro-inflammatory pathways such as type I interferon and nuclear factor-κB signaling. We generated a novel, spontaneous, and rapidly progressing mouse model of multiple sclerosis (MS) by crossing myelin-specific T-cell receptor (TCR) transgenic mice with Nlrx1-/- mice. About half of the resulting progeny developed spontaneous experimental autoimmune encephalomyelitis (spEAE), which was associated with severe demyelination and inflammation in the central nervous system (CNS). Using lymphocyte-deficient mice and a series of adoptive transfer experiments, we demonstrate that genetic susceptibility to EAE lies within the innate immune compartment. We show that NLRX1 inhibits the subclinical stages of microglial activation and prevents the generation of neurotoxic astrocytes that induce neuronal and oligodendrocyte death in vitro. Moreover, we discovered several mutations within NLRX1 that run in MS-affected families. In summary, our findings highlight the importance of NLRX1 in controlling the early stages of CNS inflammation and preventing the onset of spontaneous autoimmunity.

Exome sequencing in multiple sclerosis families identifies 12 candidate genes and nominates biological pathways for the genesis of disease.

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system characterized by myelin loss and neuronal dysfunction. Although the majority of patients do not present familial aggregation, Mendelian forms have been described. We performed whole-exome sequencing analysis in 132 patients from 34 multi-incident families, which nominated likely pathogenic variants for MS in 12 genes of the innate immune system that regulate the transcription and activation of inflammatory mediators. Rare missense or nonsense variants were identified in genes of the fibrinolysis and complement pathways (PLAU, MASP1, C2), inflammasome assembly (NLRP12), Wnt signaling (UBR2, CTNNA3, NFATC2, RNF213), nuclear receptor complexes (NCOA3), and cation channels and exchangers (KCNG4, SLC24A6, SLC8B1). These genes suggest a disruption of interconnected immunological and pro-inflammatory pathways as the initial event in the pathophysiology of familial MS, and provide the molecular and biological rationale for the chronic inflammation, demyelination and neurodegeneration observed in MS patients.

Experimental Autoimmune Encephalomyelitis Potentiates Mouse Mast Cell Protease 4-Dependent Pressor Responses to Centrally or Systemically Administered Big Endothelin-1.

Multiple sclerosis is a neurodegenerative disease affecting predominantly female patients between 20 and 45 years of age. We previously reported the significant contribution of mouse mast cell protease 4 (mMCP-4) in the synthesis of endothelin-1 (ET-1) in healthy mice and in a murine model of experimental autoimmune encephalomyelitis (EAE). In the current study, the cardiovascular effects of ET-1 and big endothelin-1 (big-ET-1) administered systemically or intrathecally were assessed in the early preclinical phase of EAE in telemetry instrumented/conscious mice. Chymase-specific enzymatic activity was also measured in the lung, brain, and mast cell extracts in vitro. Finally, the impact of EAE immunization was studied on the pulmonary and brain mRNA expression of different genes of the endothelin pathway, interleukin-33 (IL-33), and monitoring of immunoreactive tumor necrosis factor-α (TNF-α). Systemically or intrathecally administered big-ET-1 triggered increases in blood pressure in conscious mice. One week post-EAE, the pressor responses to big-ET-1 were potentiated in wild-type (WT) mice but not in mMCP-4 knockout (KO) mice. EAE triggered mMCP-4-specific activity in cerebral homogenates and peritoneal mast cells. Enhanced pulmonary, but not cerebral preproendothelin-1 and IL-33 mRNA were found in KO mice and further increased 1 week post-EAE immunization, but not in WT animals. Finally, TNF-α levels were also increased in serum from mMCP-4 KO mice, but not WT, 1 week post-EAE. Our study suggests that mMCP-4 activity is enhanced both centrally and systemically in a mouse model of EAE.

NLRX1 Enhances Glutamate Uptake and Inhibits Glutamate Release by Astrocytes.

Uptake of glutamate from the extracellular space and glutamate release to neurons are two major processes conducted by astrocytes in the central nervous system (CNS) that protect against glutamate excitotoxicity and strengthen neuronal firing, respectively. During inflammatory conditions in the CNS, astrocytes may lose one or both of these functions, resulting in accumulation of the extracellular glutamate, which eventually leads to excitotoxic neuronal death, which in turn worsens the CNS inflammation. NLRX1 is an innate immune NOD-like receptor that inhibits the major inflammatory pathways. It is localized in the mitochondria and was shown to inhibit cell death, enhance ATP production, and dampen oxidative stress. In the current work, using primary murine astrocyte cultures from WT and Nlrx1-/- mice, we demonstrate that NLRX1 potentiates astrocytic glutamate uptake by enhancing mitochondrial functions and the functional activity of glutamate transporters. Also, we report that NLRX1 inhibits glutamate release from astrocytes by repressing Ca2+-mediated glutamate exocytosis. Our study, for the first time, identified NLRX1 as a potential regulator of glutamate homeostasis in the CNS.

Astrocytes Maintain Glutamate Homeostasis in the CNS by Controlling the Balance between Glutamate Uptake and Release.

Glutamate is one of the most prevalent neurotransmitters released by excitatory neurons in the central nervous system (CNS); however, residual glutamate in the extracellular space is, potentially, neurotoxic. It is now well-established that one of the fundamental functions of astrocytes is to uptake most of the synaptically-released glutamate, which optimizes neuronal functions and prevents glutamate excitotoxicity. In the CNS, glutamate clearance is mediated by glutamate uptake transporters expressed, principally, by astrocytes. Interestingly, recent studies demonstrate that extracellular glutamate stimulates Ca2+ release from the astrocytes' intracellular stores, which triggers glutamate release from astrocytes to the adjacent neurons, mostly by an exocytotic mechanism. This released glutamate is believed to coordinate neuronal firing and mediate their excitatory or inhibitory activity. Therefore, astrocytes contribute to glutamate homeostasis in the CNS, by maintaining the balance between their opposing functions of glutamate uptake and release. This dual function of astrocytes represents a potential therapeutic target for CNS diseases associated with glutamate excitotoxicity. In this regard, we summarize the molecular mechanisms of glutamate uptake and release, their regulation, and the significance of both processes in the CNS. Also, we review the main features of glutamate metabolism and glutamate excitotoxicity and its implication in CNS diseases.

Role of the p38 MAPK/C/EBPß Pathway in the Regulation of Phenotype and IL-10 and IL-12 Production by Tolerogenic Bone Marrow-Derived Dendritic Cells.

Dendritic cells (DCs) play a major role in innate and adaptive immunity and self-immune tolerance. Immunogenic versus tolerogenic DC functions are dictated by their levels of costimulatory molecules and their cytokine expression profile. The transcription factor C/EBPß regulates the expression of several inflammatory genes in many cell types including macrophages. However, little is known regarding the role of C/EBPß in tolerogenic versus immunogenic DCs functions. We have previously reported that bone marrow-derived DCs generated with GM-CSF (GM/DCs) acquire the signature of semi-mature tolerogenic IL-10-producing DCs as opposed to immunogenic DCs generated with GM-CSF and IL-4 (IL-4/DCs). Here, we show that tolerogenic GM/DCs exhibit higher levels of phosphorylation and enhanced DNA binding activity of C/EBPß and CREB than immunogenic IL-4/DCs. We also show that the p38 MAPK/CREB axis and GSK3 play an important role in regulating C/EBPß phosphorylation and DNA binding activity. Inhibition of p38 MAPK in GM/DCs resulted in a drastic decrease of C/EBPß and CREB DNA binding activities, a reduction of their IL-10 production and an increase of their IL-12p70 production, a characteristic of immunogenic IL-4/DCs. We also present evidence that GSK3 inhibition in GM/DCs reduced C/EBPß DNA binding activity and increased expression of costimulatory molecules in GM/DCs and their production of IL-10. Analysis of GM/DCs of C/EBPß-/- mice showed that C/EBPß was essential to maintain the semimature phenotype and the production of IL-10 as well as low CD4⁺ T cell proliferation. Our results highlight the importance of the p38MAPK-C/EBPß pathway in regulating phenotype and function of tolerogenic GM/DCs.

Exhaustive Multi-Parametric Assessment of the Behavioral Array of Daily Activities of Mice Using Cluster and Factor Analysis.

Using automated supervised behavioral assessment software, we recorded and analyzed 24 h non-interrupted recordings of mice for a duration of 11 days. With the assistance of free R programming, we used correlation matrix-based hierarchical clustering and factor analysis to separate the 33 activities into meaningful clusters and groups without losing the exhaustive nature of the findings. These groups represent novel meaningful behavioral patterns exhibited by mice in home cage. Thirty-three activities were separated into 5 clusters based on dissimilarity between activities and 6 factors based on statistical modeling. Using these two methods, we describe and compare behavioral arrays of two groups of animals: 1. Continuously recorded for 11 days in social isolation and 2. Intermittently socially isolated for recording on days 1, 3, 5, 8, and 10, while socializing on the other days. This is the first work to our knowledge that interprets mouse home cage activities throughout a 24 h period and proposes a base line of a daily routine of a healthy C57Bl/6J mouse that can be used for various experimental paradigms, including disease, neuroinflammation, or drug testing to trace behavioral changes that follow intervention. In this work, we defined the necessary acclimatization period for the 24 h recording paradigm of home cage behavior. We demonstrated the behavioral changes that are associated with the effect of social isolation, intermittent socialization, and re-introduction to a familiar home cage. We provide the full description of the codes used in R.

The Dual Immunoregulatory function of Nlrp12 in T Cell-Mediated Immune Response: Lessons from Experimental Autoimmune Encephalomyelitis.

Although the etiology of multiple sclerosis (MS) remains enigmatic, the role of T cells is unquestionably central in this pathology. Immune cells respond to pathogens and danger signals via pattern-recognition receptors (PRR). Several reports implicate Nlrp12, an intracellular PRR, in the development of a mouse MS-like disease, called Experimental Autoimmune Encephalomyelitis (EAE). In this study, we used induced and spontaneous models of EAE, as well as in vitro T cell assays, to test the hypothesis that Nlrp12 inhibits Th1 response and prevents T-cell mediated autoimmunity. We found that Nlrp12 plays a protective role in induced EAE by reducing IFNγ/IL-4 ratio in lymph nodes, whereas it potentiates the development of spontaneous EAE (spEAE) in 2D2 T cell receptor (TCR) transgenic mice. Looking into the mechanism of Nlrp12 activity in T cell response, we found that it inhibits T cell proliferation and suppresses Th1 response by reducing IFNγ and IL-2 production. Following TCR activation, Nlrp12 inhibits Akt and NF-κB phosphorylation, while it has no effect on S6 phosphorylation in the mTOR pathway. In conclusion, we propose a model that can explain the dual immunoregulatory function of Nlrp12 in EAE. We also propose a model explaining the molecular mechanism of Nlrp12-dependent regulation of T cell response.

Loss of NLRX1 Exacerbates Neural Tissue Damage and NF-κB Signaling following Brain Injury.

Traumatic and nontraumatic brain injury results from severe disruptions in the cellular microenvironment leading to massive loss of neuronal populations and increased neuroinflammation. The progressive cascade of secondary events, including ischemia, inflammation, excitotoxicity, and free-radical release, contribute to neural tissue damage. NLRX1 is a member of the NLR family of pattern recognition receptors and is a potent negative regulator of several pathways that significantly modulate many of these events. Thus, we hypothesized that NLRX1 limits immune system signaling in the brain following trauma. To evaluate this hypothesis, we used Nlrx1-/- mice in a controlled cortical impact (CCI) injury murine model of traumatic brain injury (TBI). In this article, we show that Nlrx1-/- mice exhibited significantly larger brain lesions and increased motor deficits following CCI injury. Mechanistically, our data indicate that the NF-κB signaling cascade is significantly upregulated in Nlrx1-/- animals. This upregulation is associated with increased microglia and macrophage populations in the cortical lesion. Using a mouse neuroblastoma cell line (N2A), we also found that NLRX1 significantly reduced apoptosis under hypoxic conditions. In human patients, we identify 15 NLRs that are significantly dysregulated, including significant downregulation of NLRX1 in brain injury following aneurysm. We further demonstrate a concurrent increase in NF-κB signaling that is correlated with aneurysm severity in these human subjects. Together, our data extend the function of NLRX1 beyond its currently characterized role in host-pathogen defense and identify this highly novel NLR as a significant modulator of brain injury progression.

The type II transmembrane serine protease matriptase cleaves the amyloid precursor protein and reduces its processing to ß-amyloid peptide.

Recent studies have reported that many proteases, besides the canonical α-, ß-, and γ-secretases, cleave the amyloid precursor protein (APP) and modulate ß-amyloid (Aß) peptide production. Moreover, specific APP isoforms contain Kunitz protease-inhibitory domains, which regulate the proteolytic activity of serine proteases. This prompted us to investigate the role of matriptase, a member of the type II transmembrane serine protease family, in APP processing. Using quantitative RT-PCR, we detected matriptase mRNA in several regions of the human brain with an enrichment in neurons. RNA sequencing data of human dorsolateral prefrontal cortex revealed relatively high levels of matriptase RNA in young individuals, whereas lower levels were detected in older individuals. We further demonstrate that matriptase and APP directly interact with each other and that matriptase cleaves APP at a specific arginine residue (Arg-102) both in vitro and in cells. Site-directed (Arg-to-Ala) mutagenesis of this cleavage site abolished matriptase-mediated APP processing. Moreover, we observed that a soluble, shed matriptase form cleaves endogenous APP in SH-SY5Y cells and that this cleavage significantly reduces APP processing to Aß40. In summary, this study identifies matriptase as an APP-cleaving enzyme, an activity that could have important consequences for the abundance of Aß and in Alzheimer's disease pathology.

Supervised and Unsupervised Learning Technology in the Study of Rodent Behavior.

Quantifying behavior is a challenge for scientists studying neuroscience, ethology, psychology, pathology, etc. Until now, behavior was mostly considered as qualitative descriptions of postures or labor intensive counting of bouts of individual movements. Many prominent behavioral scientists conducted studies describing postures of mice and rats, depicting step by step eating, grooming, courting, and other behaviors. Automated video assessment technologies permit scientists to quantify daily behavioral patterns/routines, social interactions, and postural changes in an unbiased manner. Here, we extensively reviewed published research on the topic of the structural blocks of behavior and proposed a structure of behavior based on the latest publications. We discuss the importance of defining a clear structure of behavior to allow professionals to write viable algorithms. We presented a discussion of technologies that are used in automated video assessment of behavior in mice and rats. We considered advantages and limitations of supervised and unsupervised learning. We presented the latest scientific discoveries that were made using automated video assessment. In conclusion, we proposed that the automated quantitative approach to evaluating animal behavior is the future of understanding the effect of brain signaling, pathologies, genetic content, and environment on behavior.

NLR-Dependent Regulation of Inflammation in Multiple Sclerosis.

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) associated with inappropriate activation of lymphocytes, hyperinflammatory responses, demyelination, and neuronal damage. In the past decade, a number of biological immunomodulators have been developed that suppress the peripheral immune responses and slow down the progression of the disease. However, once the inflammation of the CNS has commenced, it can cause serious permanent neuronal damage. Therefore, there is a need for developing novel therapeutic approaches that control and regulate inflammatory responses within the CNS. Nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) are intracellular regulators of inflammation expressed by many cell types within the CNS. They redirect multiple signaling pathways initiated by pathogens and molecules released by injured tissues. NLR family members include positive regulators of inflammation, such as NLRP3 and NLRC4 and anti-inflammatory NLRs, such as NLRX1 and NLRP12. They exert immunomodulatory effect at the level of peripheral immune responses, including antigen recognition and lymphocyte activation and differentiation. Also, NLRs regulate tissue inflammatory responses. Understanding the molecular mechanisms that are placed at the crossroad of innate and adaptive immune responses, such as NLR-dependent pathways, could lead to the discovery of new therapeutic targets. In this review, we provide a summary of the role of NLRs in the pathogenesis of MS. We also summarize how anti-inflammatory NLRs regulate the immune response within the CNS. Finally, we speculate the therapeutic potential of targeting NLRs in MS.

Constitutively active Stat5b signaling confers tolerogenic functions to dendritic cells of NOD mice and halts diabetes progression.

Defects in dendritic cells (DCs) development and function lead to autoimmune disorders. Autoimmune diabetes in humans and NOD mice results from a breakdown of self-tolerance, ending in T cell-mediated ß-cell destruction. DCs dysfunction in NOD mice results in part from a defect in the JAK-STAT5 signaling pathway associated with the idd4 susceptibility locus. The involvement of Stat5b in DCs tolerogenic functions remains unknown. We have generated transgenic mice (NOD.CD11cStat5b-CA) expressing a constitutively active form of the Stat5b gene (Stat5b-CA) under control of CD11c promoter. All NOD.CD11cStat5b-CA mice were protected against diabetes. Protection was associated with an increased in the pool and suppressive function of Tregs, a promotion of Th2 and Tc2 immune response and a decreased percentage of CD8+ T cells. Splenic DCs of NOD.CD11cStat5b-CA mice acquired a mature phenotype, promoted and induced better conversion of CD4+CD25-Foxp3- T cells into Tregs (CD4+CD25+Foxp3+ T cells) than DCs of NOD mice. Stat5b-CA.DC-educated CD4+CD25- T cells delayed diabetes onset whereas Stat5b-CA.DC-educated Tregs blocked ongoing diabetes in 8-10 weeks old NOD recipient mice. Importantly, injection of Stat5b.CA.DC to 8-10-week old NOD mice halted diabetes progression and educated their splenocytes to loose their diabetogenic potential when transferred to NOD.SCID mice. Our work is the first to report that an active form of Stat5b restored DCs tolerogenic functions that re-educated Tregs to re-establish and to sustain long-term protective immune response against diabetes in NOD mice.

Significant Contribution of Mouse Mast Cell Protease 4 in Early Phases of Experimental Autoimmune Encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a mouse model that reproduces cardinal signs of clinical, histopathological, and immunological features found in Multiple Sclerosis (MS). Mast cells are suggested to be involved in the main inflammatory phases occurring during EAE development, possibly by secreting several autacoids and proteases. Among the latter, the chymase mouse mast cell protease 4 (mMCP-4) can contribute to the inflammatory response by producing endothelin-1 (ET-1). The aim of this study was to determine the impact of mMCP-4 on acute inflammatory stages in EAE. C57BL/6 wild type (WT) or mMCP-4 knockout (KO) mice were immunized with MOG35-55 plus complete Freund's adjuvant followed by pertussis toxin. Immunized WT mice presented an initial acute phase characterized by progressive increases in clinical score, which were significantly reduced in mMCP-4 KO mice. In addition, higher levels of spinal myelin were found in mMCP-4 KO as compared with WT mice. Finally, whereas EAE triggered significant increases in brain levels of mMCP-4 mRNA and immunoreactive ET-1 in WT mice, the latter peptide was reduced to basal levels in mMCP-4 KO congeners. Together, the present study supports a role for mMCP-4 in the early inflammatory phases of the disease in a mouse model of MS.