reactivity

Specificity requirements and functional properties of microbiota-reactive peri-weaning Tregs

PROJECT SUMMARY This application seeks to define the specificity requirements and functional properties of regulatory T cells (Tregs) that maintain tolerance to the microbiota. RORgt+ Tregs generated during an early-life peri-weaning window (from approximately P14 to P28 in mice) are particularly critical for intestinal tolerance. Mice that first encounter their microbiota outside this window still generate Tregs, but these cells are functionally inferior to those induced during the peri-weaning period and fail to maintain tolerance. The features of peri-weaning Tregs that make them so essential for intestinal homeostasis are not well defined. Here we propose to test two non-mutually exclusive hypotheses: 1) that the unique functionality of peri-weaning Tregs requires a distinct functional state; and 2) that reactivity with specific members of the microbiota is required for peri-weaning Tregs to maintain intestinal tolerance to a complex SPF microbiota. We have developed a model of intestinal inflammation based on oral delivery of the non-steroidal anti- inflammatory drug (NSAID) piroxicam that reveals underlying immune dysregulation in mice with defects in peri-weaning Tregs. When we applied this model to gnotobiotic mice colonized with defined microbiota communities we found that one community (OMM12) induced Tregs capable of preventing inflammation while the other community (ASF) did not, despite similar induction of RORgt+ peri-weaning Tregs by both communities. This exciting result suggests a previously unappreciated specificity requirement for induction of peri-weaning Tregs and indicates that differences in the microbes encountered early in life can have lifelong ramifications for immune tolerance. To better understand the basis of this specificity requirement, we developed a pipeline to rapidly screen the reactivity of T cells and applied it to mice colonized with the protective OMM12 community. This analysis revealed that the antigen-specific Treg response is biased toward only a subset of the microbiota. Thus, by tracking and characterizing microbiota-reactive peri-weaning Tregs at unprecedented resolution, we uncovered an unexpected bias in the microbiota-reactivity of Tregs. We are now ideally positioned to examine how the specificities and functional properties of peri-weaning Tregs are linked to their unique role in intestinal tolerance. In Aim 1, we will define the specificity of microbiota- reactive peri-weaning Tregs at homeostasis, using new tools developed through our screening pipeline, and we will determine whether missing the weaning period alters Treg responses to the microbiota. In Aim 2, we will compare the transcriptional programs of peri-weaning and post-weaning Tregs to identify peri-weaning- specific features. We will also build on our analyses from Aim 1 to determine if functional differences are linked to reactivity with specific members of the microbiota. In Aim 3, we will explore why specific members of the microbiota are required for induction of protective peri-weaning Tregs. We will define communities of microbes that do or do not confer protection in our piroxicam model, and we will profile the Tregs in these communities, including microbiota-reactive Tregs with defined specificities, to test the hypothesis that a key aspect of peri- weaning Treg function is specificity for only certain gut microbes.

Dissecting the role for astrocytes in mediating adverse outcomes of maternal immune activation.

Prenatal infections cause maternal immune activation (MIA), a major risk factor for several neurodevelopmental disorders, including schizophrenia, autism spectrum disorders (ASD), and attention deficit hyperactivity disorder (ADHD). Consequently, elucidating the mechanisms by which MIA alters brain function is critical for understanding the pathophysiology of these disorders and developing effective treatments. While the effects of MIA on neurons and microglia have been extensively studied, the impact of MIA on astrocytes, key regulators of brain physiology and homeostasis, remain unknown that significantly impedes our understanding the mechanisms of MIA-induced neurobehavioral abnormalities. To address this major knowledge gap, we conducted pilot studies that suggest that MIA increases impulsivity-like behaviors and amphetamine-induced hyperactivity and enhances extracellular levels of glutamate (GLU) and dopamine (DA) in the dorsal striatum (DS). MIA also increased pro-inflammatory signatures of astrocytes, including up- regulation of the Nuclear Factor kappa B (NF-κB) pathway and increased GFAP immunoreactivity in DS astrocytes. Collectively, these novel findings support our overarching hypothesis that MIA increases astrocyte reactivity, leading to increased gliotransmission (e.g., GLU), which in turn enhances DS DA release and DA- dependent behaviors. To test this hypothesis, we will leverage the expertise of the research team in molecular, physiological and neurobehavioral approaches and conduct the following Specific Aims: In Aim 1, we will identify the MIA-induced cellular and physiological changes characteristic of astrocyte reactivity. In Aim 2, we will determine the circuit mechanisms by which MIA increases DA signaling. In Aim 3, we will identify the molecular mechanisms whereby reactive astrocytes contribute to MIA-induced cellular and behavioral abnormalities. These studies will enhance the current understanding of the effects of MIA on brain functions and generate new insight into potential treatment strategies for MIA-associated neurodevelopmental disorders.

Bridging Local and System-Wide Autoreactive, Extrafollicular B Cell Signatures in a TLR7-Driven Model

Project Summary A substantial body of literature has described the development of autoreactive humoral responses in the context of autoimmune disease and recently discerned an exciting new avenue for investigation. While early work focused on canonical mechanisms of activation through the germinal center (GC) response, recent studies have found GC infrastructure to be dispensable for the onset of chronic autoimmunity. It has become clear that an alternative pathway of B cell activation, the extrafollicular (EF) pathway, can drive the onset of new autoreactivity in multiple human disorders including rheumatoid arthritis and systemic lupus erythematosus (SLE). In comparison to the GC pathway, the EF pathway represents a less stringent method for B cell activation, leads to accelerated antibody-secreting cell (ASC) formation, and thus has a higher propensity for the production of autoreactive B cell effectors and ASCs. Recently, our group has identified a similar skew toward the EF response in the context of severe viral infection, tied to acute tolerance loss, increased disease severity, and complicated recovery from infection. These findings highlight how further study of the EF response is crucial to our understanding of autoimmune induction across multiple areas of disease. Toll-like receptor 7 (TLR7) stimulation has been identified as a key contributor to EF B cell development in SLE, and several studies have now linked TLR7 overstimulation to chronic autoimmune disease. While EF effector B cell populations have now been identified in both murine models and humans, substantial gaps in our knowledge remain to be answered concerning i) the origins of these cells and ii) the system-wide and microenvironmental signaling and organization that drive this differentiation pathway. We propose to address these gaps, here, by utilizing a TLR7 agonist (R848) in a murine model to characterize the autoreactive response within the blood and draining lymph node through innovative high-throughput analytical techniques. Systemic shifts in proteomic signatures and immune cell phenotype will be monitored in the blood throughout the induction of autoreactivity, using novel applications of machine-learning based classification. These signatures will then be connected to developing inflammatory microenvironments identified within the draining lymph node by applying a customized set of software tools to spatial transcriptomic data. This work will deepen our understanding of the immunologic mechanisms by which the EF pathway can lead to “run-away” autoreactive B cell development, with the added potential for identification of early blood-based biomarkers for this developing autoreactivity. The above proposed work will provide an ideal training opportunity for the candidate to develop experience with advanced immunologic laboratory techniques, rigorous bioinformatic analysis, a systems-level view of immunology, and scientific communication. The Woodruff and Sanz Labs are highly experienced within the autoimmune disease space with extensive experience with the required techniques and established routes for clinical collaboration to act on these findings.

Implementing a New Paradigm for Antifungal Drug Development

About 30% of the drugs currently in clinical use function through covalent modification of their target. Yet, until recently, none of these covalent drugs were specifically designed to utilize this irreversible mode of action. It is our hypothesis that the production of a new class of covalent inactivators, designed to selectively modify new drug targets, will lead to novel agents with efficacy against both native and drug-resistant pathogenic fungal species. Because of their novelty these agents will also offer a greater opportunity to bypass the existing mechanisms of drug resistance. Pathogenic fungal infections remain among the leading causes of human mortality, and this threat is rising due to the increasing prevalence of drug- resistance strains and the paucity of effective antifungal drugs against the more virulent fungal species. Our proposed new drug target is an enzyme that plays a critical role in a uniquely microbial pathway that is essential for the survival of fungal organisms. To test our hypothesis and achieve the goals of this project we plan to complete the following specific aims during the initial R21 phase of this project: (1) Optimization of the potency of novel enzyme inactivators. Our goals here are to use our strong preliminary results to address critical barriers that must be overcome to convert potent enzyme inactivators into advanced drug candidates, thereby achieving higher target selectivity and increasing compound reactivity once bound to the target; (2) Enhance the antifungal capability of these enzyme inactivators. Our strategy for this aim is focused on the incorporation of conjugate partners into this new class of covalent inactivators, enabling them to potentially utilize the existing nutrient uptake systems to achieve toxic levels in Candida species; (3) Examine the target selectivity of our new antifungal agents. Results from fungal growth inhibition and fungal killing assays will be used to evaluate and rank the efficacy of our compounds against both wild-type and drug-resistant Candida strains. Specific milestones are presented to evaluate our achievement of these initial aims. Once accomplished we will immediately proceed to the R33 phase of this project, with the aims of: (4) Pharmacological evaluation of lead candidates, though ranking the drug candidates based on their ADME, pharmacokinetic and toxicity properties; and then (5) Evaluate the efficacy of our candidates against pathogenic fungal infections. A systematic infection animal model will be utilized for candidate screening to identify the best agents against disseminated fungal infections, followed by further efficacy screening in an oral infection model. Completion of these aims will produce, refine and evaluate a new class of antifungal agents with a novel mode of action against an unexplored but essential fungal target. The agents with the most promising drug profiles will then be moved into advanced preclinical trials used to select the most effective new antifungal agents.

From the first spark to catching your breath: A love story in neuroscience

What are the things that draw us to a particular field of science and what is it that keeps us there? For Dr. Bahia, there was a particular attraction to sensory nerves; the monitors of the worlds inside and outside of our bodies. In this talk, Dr. Bahia will outline her career path as a neuroscientist resulting in the title of Research Associate. She will also talk about the longest project she has participated in, 'exploring the role of ion channels in sensory nerves' (rupress.org/jgp/article/147/6/451/43495/The-exceptionally-high-reactivity-of-Cys-621-is)

Role of Oxytocin in regulating microglia functions to prevent brain damage of the developing brain

Every year, 30 million infants worldwide are delivered after intra-uterine growth restriction (IUGR) and 15 million are born preterm. These two conditions are the leading causes of ante/perinatal stress and brain injury responsible for neurocognitive and behavioral disorders in more than 9 million children each year. Both prematurity and IUGR are associated with perinatal systemic inflammation, a key factor associated with neuroinflammation and identified to be the best predictor of subsequent neurological impairments. Most of pharmacological candidates have failed to demonstrate any beneficial effect to prevent perinatal brain damage. In contrast, environmental enrichment based on developmental care, skin-to-skin contact and vocal/music intervention appears to confer positive effects on brain structure and function. However, mechanisms underlying these effects remain unknown. There is strong evidence that an adverse environment during pregnancy and the perinatal period can influence hormonal responses of the newborn with long-lasting neurobehavioral consequences in infancy and adulthood. Excessive cortisol release in response to perinatal stress induces pro-inflammatory and brain-programming effects. These deleterious effects are known to be balanced by Oxytocin (OT), a neuropeptide playing a key role during the perinatal period and parturition, in social behavior and regulating the central inflammatory response to injury in the adult brain. Using a rodent model of IUGR associated with perinatal brain damage, we recently reported that Carbetocin, a brain permeable long-lasting OT receptor (OTR) agonist, was associated with a significant reduction of activated microglia, the primary immune cells of the brain. Moreover this reduced microglia reactivity was associated to a long-term neuroprotection. These findings make OT a promising candidate for neonatal neuroprotection through neuroinflammation regulation. However, the causality between the endogenous OT and central inflammation response to injury has not been established and will be further studied by the lab.

What about antibiotics for the treatment of the dyskinesia induced by L-DOPA?

L-DOPA-induced dyskinesia is a debilitating adverse effect of treating Parkinson’s disease with this drug. New therapeutic approaches that prevent or attenuate this side effect is clearly needed. Wistar adult male rats submitted to 6-hydroxydopamine-induced unilateral medial forebrain bundle lesions were treated with L-DOPA (oral or subcutaneous, 20 mg kg-1) once a day for 14 days. After this period, we tested if doxycycline (40 mg kg-1, intraperitoneal, a subantimicrobial dose) and COL-3 (50 and 100 nmol, intracerebroventricular) could reverse LID. In an additional experiment, doxycycline was also administered repeatedly with L-DOPA to verify if it would prevent LID development. A single injection of doxycycline or COL-3 together with L-DOPA attenuated the dyskinesia. Co-treatment with doxycycline from the first day of L-DOPA suppressed the onset of dyskinesia. The improved motor responses to L-DOPA remained intact in the presence of doxycycline or COL-3, indicating the preservation of L-DOPA-produced benefits. Doxycycline treatment was associated with decreased immunoreactivity of FosB, cyclooxygenase-2, the astroglial protein GFAP and the microglial protein OX-42 which are elevated in the basal ganglia of rats exhibiting dyskinesia. Doxycycline also decreased metalloproteinase-2/-9 activity, metalloproteinase-3 expression and reactive oxygen species production. Metalloproteinase-2/-9 activity and production of reactive oxygen species in the basal ganglia of dyskinetic rats showed a significant correlation with the intensity of dyskinesia. The present study demonstrates the anti-dyskinetic potential of doxycycline and its analog compound COL-3 in hemiparkinsonian rats. Given the long-established and safe clinical use of doxycycline, this study suggests that these drugs might be tested to reduce or to prevent L-DOPA-induced dyskinesia in Parkinson’s patients.

Biomarkers for Addiction Treatment Development: fMRI Drug Cue Reactivity as an Example

This webinar is mainly focused on “Biomarkers for Addiction Treatment Development: fMRI Drug Cue Reactivity as an Example”. Biomarkers and Biotypes of Drug Addiction: funding opportunities at NIDA, Tanya Ramey (NIDA, US) Neuroimaging-based Biomarker Development for Clinical Trials, Owen Carmicheal (Pennington Biomedical Research Center, USA) ENIGMA-Addiction Cue Reactivity Initiative (ACRI) and Checklist, Hamed Ekhtiari (Laureate Institute for Brain Research, USA) ENIGMA-ACRI Checklist: Participant Characteristics, General fMRI Information, General Task Information, Cue Information, Task-related Assessments, Pre-Post Scanning Consideration (James Prisciandaro, Medical University of South Carolina, USA; Marc Kaufman, McLean Hospital/Harvard Medical School, USA; Anna Zilverstand, University of Minnesota; Torsten Wüstenberg, Charité Medical University Berlin, Germany; Falk Kiefer, University of Heidelberg, Germany; Amy Janes, Harvard Medical School, USA) How to Add fMRI Drug Cue Reactivity to the ENIGMA Consortium: Road Ahead, Hugh Garavan, University of Vermont)

Assessing the hyperexcitability of the epileptic brain by burst-suppression EEG reactivity

Assessment of the rat ischemic brain by burst-suppression EEG reactivity

Cerebrovascular reactivity to hypercapnia and neurovascular coupling explore different vascular beds

Characterization of astrocyte reactivity in a model of encephalopathy of prematurity

A common mechanism for saliency detection and motor reactivity in humans and rhesus monkeys

Contribution of peripheral neuronal activity to spinal microglial reactivity in chronic pain

The default EEG macrostate reactivity to the subject’s own name depends on awareness

Drebrin controls scar formation and astrocyte reactivity upon traumatic brain injury by regulating membrane trafficking

Impact of astrocyte reactivity in cocaine addiction

Impact of calorie-restricted cafeteria feeding and treadmill exercise on sucrose intake, sensitivity and reactivity in diet-induced obese male and female rats

Investigating the effects of acute THC vapour exposure on stress reactivity and fear conditioning

Investigation of Microglial and Astrocytic Reactivity in Mouse Models of Down Syndrome

Macrophage-derived amphiregulin modulates astrocyte network reactivity

Reactivity to conditioned threat cues is distinct from exploratory drive in the elevated plus-maze

Repeated stress in the early adolescent period: Are CB1 receptors mediating stress-reactivity in a sex-specific manner?

STAT3-mediated astrocytic reactivity in glioblastoma multiforme

Ubiquinol supplementation improves gender-dependent cerebral vasoreactivity and ameliorates chronic inflammation and endothelial dysfunction associated with mild cognitive impairment

What is the contribution of cerebrovascular reactivity to measures of haemodynamic lags in patients with left hemisphere stroke?

Adenosine triggers astrocyte reactivity to drive the pathogenesis of sepsis-associated encephalopathy

FENS Forum 2024

Assessing the EEG reactivity to photic stimulation in the sedated epileptic brain

FENS Forum 2024

The default EEG reactivity to photic stimulation is impaired following global cerebral ischaemia and reperfusion

FENS Forum 2024

Epigenetic modulation of astrocyte reactivity to prevent neurodegeneration

FENS Forum 2024

Loss of cyclooxygenase-1 attenuates microglia reactivity after optic nerve injury

FENS Forum 2024

Modulating the astrocyte reactivity by blocking P2X7R and Panx1 in vitro – intercepting the neuroinflammation chronicity development

FENS Forum 2024

Newly synthesized fatty acid analogue (NKS-3) rescues microglial reactivity in a murine model of diet-induced obesity

FENS Forum 2024

Cue reactivity in electronic cigarette users is unrelated to sign-tracking propensity

FENS Forum 2024

Cue reactivity of non-dopamine neurons in the midbrain

FENS Forum 2024

Unraveling the neurogenic niche symphony: Hippocampal astrocytes mitigate microglial reactivity to inflammation

FENS Forum 2024