bluuebunny/crossref_metadata_2025_split huggingface dataset:
T-cell infiltrates and microglia adopt long-term gene signature changes leading to age-specific responses to traumatic brain injury in mice
Authors: Chen Zhangying , Islam Mecca , Davis Booker , Schwulst Steven 
Published: January 2023

Abstract
Aged traumatic brain injury (TBI) patients suffer increased mortality and long-term neurocognitive/neuropsychiatric morbidity than younger patients. Microglia, the resident macrophages of the brain, are complicit in both. We hypothesized that aged microglia would fail to return to a homeostatic state after TBI and adopt a long-term, injury-associated state within aged brains compared to young brains after TBI. Young and aged male C57BL/6 mice underwent TBI via controlled cortical impact vs. sham injury and were sacrificed four months post-TBI. We utilized single-cell RNA sequencing to examine age-associated cellular responses after TBI. Brains were harvested with CD45+ cells isolated via florescence-activated cell sorting. cDNA libraries were prepared via the 10x Genomics Chromium Single Cell 3’ Reagent Kit, followed by sequencing on a HiSeq 4000 instrument and computation analyses. Post-injury, aged mice demonstrated a proportional decrease in homeostatic microglia, and greater increased infiltrating T cells compared to young-adult mice. Of note, aged mice post-injury had a subpopulation of age-specific, immune-inflammatory microglia resembling gene profiles of neurodegenerative disease-associated microglia with enriched pathways involving in leukocyte recruitment. Contrastingly, post-injury, aged mice demonstrate a heterogenous T-cell infiltration with gene profiles corresponding to CD8 effector memory, CD8 native-like, CD4, and double-negative T cells and enriched pathways such as macromolecule synthesis. Taken together, our data showed that age-specific gene signature changes in the T-cell infiltrates and the microglial subpopulation contribute to increased vulnerability of the aged brain to TBI.
Supported by R01 GM130662
Neural Dysfunction Following Francisella tularensis Infection
Authors: Tena Anahis , Hightower Rebekah T , Sanchez Guillen Michelle A , Spencer Charles T 
Published: January 2023

Abstract
Francisella tularensis is a gram-negative intracellular coccobacillus that is the causative agent of tularemia. Local, acute inflammation is beneficial since it can clear bacterial infections, remove cellular damage and promote the healing of tissue. However, F. tularensis is capable of bypassing the immune system and causing an over-activation of inflammation, known as a “cytokine storm”. This cytokine storm increases host cell damage, nutrient availability and serves an immune distraction allowing the bacterium to replicate. Other infections that cause cytokine storms include Ebola, pandemic influenza, hantavirus, and HIV, all BSL3 pathogens.
The routes of transmission for F. tularensis are many and intertwined. Infection could be the result of inhaling air droplets containing infectious bacteria, coming into direct contact with an infected animal or being bitten by an infected arthropod such as a tick, flea or mosquito. Eating and drinking contaminated meat and water can also lead to infection and different disease manifestations occur depending on the route of transmission.
While extensive research has documented infection of the spleen, liver and lung, there hasn’t been any reports of F. tularensis productively infecting the brain, until now. This work focuses on characterizing the effects of F. tularensis infection of the brain. We hypothesize that damage caused by F. tularensis neural infection will have a negative effect on behavior. This work aims to identify these negative effects based on the anatomical location of the bacteria foci, regions containing numerous bacteria. With the use of brain mapping and behavioral assays, we seek to determine the impact of F. tularensis infection on behavior and cognition.
Outside-in IL1b-HMGB1 signaling axis may drive astrocytic response in drug resistant epilepsies.
Authors: Clarkson Ben D S , Lafrance-Corey Reghann , McCarthy Christina , Johnson Renee , Shrestha Ramila Barun , Howe Charles L 
Published: January 2023

Abstract
We recently reported normalization of CSF cytokine levels following successful treatment of drug-resistant epilepsy (DRE) with recombinant interleukin-1 receptor antagonist (rIL1RA). To further understand the mechanism of this therapeutic outcome, we interrogated the ability of serum and CSF factors that are elevated in drug resistant-seizure disorders to promote inflammatory responses in astrocytes and human brain microvessel endothelial cells (HBMVECs). We report that CCL2, TNFa, IL6, and CXCL8/1 were strongly induced in 1) astrocytes and HBMVECs stimulated with IL1β (blocked by rIL1RA) and 2) astrocytes but not HBMVECs in response to DRE patient serum but not control serum (partially blocked by rIL1RA). This suggested that other serum factors contribute to astrocyte activation in this DRE. HMGB1 and IL18 were also dysregulated in DRE patient serum; however, IL18 failed to stimulate cytokine secretion from astrocytes. In contrast, HMGB1 induced cytokine secretion from astrocytes, which was blocked by the TLR4 inhibitor TAK242. Remarkably, TAK242 also prevented cytokine production by astrocytes in response to serum from some patients with DRE. Finally, IL1β induced the polarized uptake and secretion of HMGB1 across transwell inserts harboring confluent monolayers of HBMVECs suggesting that IL1b signaling may cause endothelial cells to transport serum HMGB1 into the perivascular space—promoting TLR4-dependent cytokine release from astrocytes. This translation of an “outside” inflammatory signal into an “inside” inflammatory response may underlie seizure disorders that are resistant to anti-seizure drugs.
Trained Immunity in Neurodegenerative Diseases
Authors: Bossu’ Paola , Sireno Laura , Lembo Micaela , Toppi Elisa 
Published: January 2023

Abstract
Alzheimer’s disease (AD) and Parkinson’s disease (PD) are complex and progressive neurodegenerative disturbances leading to cognitive or motor dysfunctions and lacking effective therapy. A deregulation of innate immune response and inflammation appears to be deeply involved in both illnesses, though with still unclear mechanisms. Evidence in animals attributes a neurodegeneration-modulating role to mechanisms of innate memory, but human data on this topic are still scarce. In this study, we investigated whether -possibly due to concomitant factors including enhanced infectious burden, microbial dysbiosis and genetic risk factors linked to immunity-the innate immune cells of AD and PD patients show long-term functional reprogramming characterized by either potentiated or tolerant responses, which could ultimately contribute to disease development. Thus, following repeated in vitro stimulation with either heat-inactivated Candida albicans or LPS, we measured the cytokine response of CD14+ cells from AD and PD patients, both in comparison with those of age- and gender-matched healthy control subjects and in association with symptom severity. We observed different cytokine patterns associated with tolerant or potentiated responses among the three groups of subjects, pointing to different and disease-dependent trained immune status in patients. Our results would provide insights about how previous triggering of innate immunity can influence neuroinflammation and disease progression, offering new knowledge on trained immunity impact in neurodegenerative diseases.
Effect of BPA on cytokine expression and regeneration in larval zebrafish
Authors: Walser-Kuntz Debby , Wang Hongshen , Alvarez Luis , Intasin Pear 
Published: January 2023

Abstract
Bisphenol-A (BPA) binds to estrogen receptors ER-α and β and is used in the production of polycarbonate plastic and epoxy resins. BPA presents a potential environmental challenge to the immune system of both humans and wildlife due to its use in food containers and thermal receipts and its resulting presence in surface water and wastewater runoff. We observed that BPA exposure downregulates the expression of the chemokine that functions to recruit neutrophils in both mice (KC) and zebrafish (cxcl8) and that it decreases neutrophil recruitment to the injury site in MPO::GFP zebrafish. Neutrophils are essential for the removal of damaged tissue and invading microbes, thus facilitating tissue regeneration of the tail fin. To investigate BPA’s effects on larval zebrafish tail regeneration, we induced a tail injury at day 3 post fertilization (dpf) and measured tail fin growth at day 1 and 3 post injury (dpi). Fish exposed to 100 ng/ml BPA regenerate a smaller tail fin area at 3 dpi (p-value &lt; 0.01). We hypothesized that BPA exposure and altered neutrophil recruitment could affect regeneration by directly altering the expression of regeneration-related genes wnt10a and il-10 or their downstream genes and/or indirectly by regulating inflammatory genes including il-1 and il-6. The expression of il-1, il-6, il-10, and wnt10a were detected in zebrafish embryos from 2–6 dpf using qPCR. BPA exposure reduces both il-1 and il-6 expression 3 hours post-injury, suggesting that BPA reduces the inflammatory response. To determine whether BPA alters gene expression of wnt10a, il-10, or their downstream-regulated genes qPCR will be performed at 24- and 72-hours post-injury.
Supported by Carleton College.
The N-glycosyltransferase ST6GAL1 plays a key role in M2 macrophage anti-inflammatory functions
Authors: Barnes Rob , Dolatshahi Sepideh 
Published: January 2023

Abstract
Macrophages are effector cells of the innate immune system, and play distinct roles depending on their polarization state. Glycosylation is a ubiquitous post translational modification made to proteins that plays a large role in protein biophysical properties and function. Through computational analysis of a publicly available macrophage transcriptome data set, we demonstrated that glycosylation changes dramatically across different macrophage polarizations. Specifically, we identified the gene coding for the glycosyltransferase ST6GAL1, which terminally sialylates protein glycans, as significantly differentially expressed across macrophage polarities. ST6GAL1 was downregulated in macrophages polarized by M1 polarizing signals TNFa and TNFa+IFNg and upregulated by M2 polarizing signals IL-4 and IL-13. We hypothesized that changes in terminal sialic acid frequency during glycosylation play a role in M2 macrophage immunosuppressive actions. An analysis of SIGLEC signaling pathways revealed that increasing ST6GAL1 expression was concordant with increasing CD52 expression, a ligand for SIGLEC10. SIGLEC proteins contain an immunoreceptor tyrosine-based inhibitory motif (ITIM) which inhibits immune action upon activation. SIGLEC10 expression increased in both M1 and M2 polarized macrophages, suggesting a possible mode of immunosuppression regulated by ST6GAL1. The increased frequency of terminally sialylated glycans and CD52 on M2 surfaces may increase the frequency with which M2 macrophages bind to and activate SIGLEC proteins on other immune cells. To test this hypothesis, we will knockout and amplify M2 macrophage ST6GAL1 expression to determine its influence on M2 immunosuppressive capacity.
Notch2, Bcl6, and IRF2 govern differentiation and survival of murine nonclassical monocytes.
Authors: O’Connor Kevin W , Liu Tiantian , Kim Sunkyung , Murphy Theresa , Murphy Kenneth M 
Published: January 2023

Abstract
Nonclassical (patrolling) monocytes are a population of monocytes which monitor the vasculature and participate in tissue repair. Though nonclassical monocytes are thought to be developmentally derived from classical monocytes, the mechanisms that control the differentiation of classical monocytes into nonclassical monocytes are still not completely understood. Signaling through the Notch2 receptor appears to trigger development of nonclassical monocytes. Also, the transcription factors C/EBPβ and Nur77 (Nr4a1) are required for nonclassical monocyte development in vivo. We have now identified additional transcription factor requirements for the differentiation and/or survival of nonclassical monocytes in vivo. We find that mice with conditional deletion of Bcl6 in myeloid cells or germline IRF2 deficiency show a severe loss of nonclassical monocytes. Both Bcl6 and IRF2 are induced during the transition from classical to nonclassical monocyte. In vitro culture of myeloid progenitors on stromal cells expressing DLL1, a Notch2 ligand, recapitulates aspects of nonclassical monocyte development seen in vivo, confirming Notch signaling as a key initiating event in the development of nonclassical monocytes. This system allows for the identification of the downstream targets of Notch2 signaling that drive nonclassical monocyte development, as well as the analysis of the impact of other transcription factors on the differentiation and survival of these cells in vivo. We present a model of the steps required for development of murine nonclassical monocytes.
Supported by NIH (RO1 AI150297, RO1 CA248919)
Nuclear soluble cGAS senses DNA virus infection
Authors: Wu Yakun , Song Kun , Hao Wenzhuo , Wang Lingyan , Li Shitao 
Published: January 2023

Abstract
The DNA sensor cGAS detects cytosolic foreign DNA from pathogens or self-DNA from cellular damage and instigates type I interferon (IFN) expression. Recent studies find that cGAS also localizes in the nucleus and tethers to the chromatin. Despite how cGAS is inhibited in the nucleus is well elucidated, whether nuclear cGAS participates in DNA sensing is not clear. Here, we report that herpes simplex virus 1 (HSV-1) infection causes the release of cGAS from the chromatin into the nuclear soluble fraction, and the free nuclear cGAS senses viral DNA to induce innate immune response. We first found that the infection of several HSV-1 strains led to cGAS release from the chromatin in several types of cells, including in RAW264.7 macrophages, THP-1 monocytes, and bone marrow-derived macrophages. DNA damage agent cisplatin failed to induce nuclear soluble cGAS, suggesting that cGAS release from the chromatin is not due to DNA damage or apoptosis. Interestingly, inhibition of HSV-1 replication blocked the release of nuclear soluble cGAS. To determine the role of nuclear cGAS in DNA virus infection, we reconstituted a Dox-induced nuclear-only cGAS mutant into cGAS knockout RAW264.7 cells. After induction, only chromatin-bound cGAS was found in the reconstituted cells. Similarly, HSV-1 infected also induced nuclear soluble cGAS in this cell line. More importantly, the nuclear-only cGAS stable cells limited HSV-1 infection by producing cGAMP and inducing mRNA expression of type I IFN and interferon-stimulated genes. Taken all together, our study demonstrates that nuclear soluble cGAS is a DNA sensor inside the nucleus.
This research was funded by the National Institutes of Health (R21AI137750 and R01AI141399 to S.L.).
Lipopolysaccharide pre-treatment prevents viral induced death by priming macrophages for a robust anti-viral immune response
Authors: Resiliac Jenny , Rohlfing Michelle , Santoro Jennifer , Hussain Syed-Rehan A , Grayson Mitchell H 
Published: January 2023

Abstract

Rationale
Respiratory viruses are an increasing public health burden. Using Sendai virus (SeV, murine parainfluenza virus type 1), previous data indicate that pretreatment of C57BL6 wild-type (WT) mice with 0.1μg of lipopolysaccharide (LPS) prevented mortality from a normally lethal SeV infection. However, the mechanism by which LPS pre-treatment confers this survival advantage is unknown. LPS is a potent activator of macrophages. We hypothesized that LPS prevents viral induced death by stimulating the recruitment of macrophages, which could drive a more robust anti-viral immune response and subsequent survival. Therefore, we examined the change in lung macrophage subsets with LPS pretreatment of SeV infected mice.

Methods
WT mice were inoculated with 30 μL of 0.1 μg LPS or PBS i.n. 24 hours before SeV (2e6 pfu). After LPS sensitization, lungs were harvested at 6 hours, 24 hours, and days 1,3,5, and 7 post SeV infection (PI SeV); single lung cell suspensions isolated and stained for various innate immune cells by flow cytometry to assess the immune cells recruited by LPS.

Results
Mice pre-treated with LPS had a significant increase of alveolar macrophages (GR1- Siglec F+ CD11c+) at days 1 and 3 PI SeV (p=0.080 and p=0.0026, respectively; n≥6). Additionally, interstitial macrophages (GR1- Siglec F− CD11b+ CD11c+ F4/80+ MHCII+) were significantly increased at days 0 and 3 PI SeV (p=0.025 and p=0.019 respectively; n≥6). The LPS effect was no longer significant by day 5 PI SeV.

Conclusion
. LPS pretreatment of mice increases macrophages early during the SeV infection. Further studies will be needed to determine if the increase in these macrophages provides the protection from viral induced mortality seen in our model and the cytokines produced.
This work was supported by the Abigail Wexner Research Institute at Nationwide Children’s Hospital (MHG).
The impact of lipid nanoparticles (LNP) on anti-viral pathways and immune function in aged individuals
Authors: Connors Jennifer R , Cusimano Gina , Kim Kenneth , Alameh Mohamad-Gabriel , Weissman Drew , Kutzler Michele , Haddad Elias K 
Published: January 2023

Abstract
Aging leads to increased morbidity and mortality posing a big challenge to societies as the world population shifts to an increased proportion of aged individuals. Immunosenescence, a condition of chronic increased level of basal inflammation and immune system dysregulation, is suspected to decrease vaccine efficacy and enhance COVID-19 complications in older individuals. Current vaccination efforts like mRNA vaccines utilize lipid nanoparticle (LNP) platforms that elicit the immune response however, little is known about how LNP adjuvantation affects anti-viral pathways and immune function in aged individuals. We have previously shown that type I IFN production including the phosphorylation of IRF7 and TBK-1 in the TLR 7/8, RIG-I or cGAS-STING pathways and phagocytosis is dysregulated in innate immune cells (i.e., cDC1s, cDC2s, and CD14dimCD16+ monocytes) from healthy older adults (n=11). Our preliminary data utilizing LNP show that in response to LNP stimulus at 4 and 24 hours, older adults have decreased IFNy output. Thus, we further hypothesize that cDC1, cDC2, and CD14dim CD16+ monocytes from healthy, older adults (n=11) will have dysregulated anti-viral signaling and antigen presentation in response to LNP stimulation as measured by the defect in type I IFN production, phosphorylation of IRF7, TBK-1, and immune function like phagocytosis and antigen presentation via flow cytometry. These data support our previous reports of a delayed/decreased proinflammatory output following stimulation and demonstrate potential targets of antiviral pathways that can resolve dysfunctions of the aging immune system for an effective vaccination response.
Supported by NIH (U19 AI128910-04S1)
Astrocytic RIPK3 confers protection against deleterious neuroinflammation during Zika virus infection
Authors: Lindman Marissa , Angel Juan , Newman Kimberly , Atkins Colm , Daniels Brian 
Published: January 2023

Abstract
This study aims to identify the function(s) of RIPK3 signaling in astrocytes following Zika virus infection. Previous work found that RIPK3 signaling in Zika virus-infected neurons activates inflammatory transcription factors such as NFκB and IRF1, leading to the upregulation of inflammation-associated transcripts. We were thus interested in determining the role of RIPK3 signaling in astrocytes, which are critical regulators of neuroinflammation. Using mice with an astrocyte-specific conditional Ripk3 deletion, we found that intracranial Zika virus infection was significantly more lethal in mice deficient in astrocytic Ripk3 than in littermate controls. To identify mechanisms underlying this difference, we isolated and infected primary fore- and hind-brain astrocytes with Zika virus to determine the transcriptional consequences of genetic Ripk3 ablation. Surprisingly, we found increased expression of several chemokines, cytokines and ISGs in Ripk3−/− hindbrain astrocytes, in contrast to our previous findings in neurons. Subsequent leukocyte profiling from the brains of Zika virus-infected mice revealed increased numbers of CD4+ and CD8+ T cells, natural killer cells, and monocytes in mice deficient in astrocytic Ripk3 compared to those found in littermate controls. As previous work has demonstrated that astrocytic type I interferon signaling in the hindbrain is responsible for downregulating proinflammatory molecules to prevent lethal neuroinflammation, our data suggest that synergistic signaling between type I IFN and RIPK3 in hindbrain astrocytes suppresses deleterious neuroinflammation and promotes host survival in the setting of Zika virus encephalitis.
Supported by a grant from NIH (R01 NS120895).
Effector-mediated subversion of proteasome activator (PA)28αβ enhances lysosomal pathogen targeting within cytokine-activated macrophages
Authors: Shames Stephanie R 
Published: January 2023

Abstract
Macrophages activated by proinflammatory cytokines, such as tumor necrosis factor (TNF) and interferon (IFN)-γ, are potently and broadly microbicidal. Legionella pneumophila causes Legionnaires’ Disease via replication within host macrophages using an arsenal of hundreds of translocated virulence factors termed effector proteins. Effectors are critical for intracellular replication but can also enhance pathogen clearance in mammalian hosts via effector-triggered immunity. The effector LegC4 confers a fitness disadvantage on L. pneumophila in mouse models of Legionnaires’ Disease and uniquely potentiates the antimicrobial activity of macrophages activated with either TNF or IFN-γ. We found that LegC4 binds proteasome activator (PA)28α, a subunit of the PA28αβ (11S) proteasome regulator, and that the LegC4 restriction phenotype is abolished within PA28αβ-deficient macrophages. PA28αβ facilitates ubiquitin-independent proteasomal degradation of oxidant-damaged proteins. Impaired proteasome activity results in compensatory upregulation of lysosomal degradation pathways to relieve oxidative proteotoxic stress. We found that LegC4 impairs the resolution of oxidative proteotoxic stress and enhances phagolysosomal fusion with the Legionella-containing vacuole. PA28αβ has been traditionally associated with antigen presentation and adaptive immunity; however, our data support a model whereby suppression of PA28αβ by LegC4 impairs resolution of oxidative proteotoxic stress, which culminates in lysosomal killing of L. pneumophila within activated macrophages. This work provides a solid foundation on which to evaluate induced proteasome regulators as mediators of cell-autonomous immunity.
This work is supported by NIH grant P20GM130448.