bluuebunny/crossref_metadata_2025_split huggingface dataset:
Fam49b, the source of antigenic peptide for ERAAP surveillance, controls thymocyte development.
Authors: Park Chansu , Guan Jian , Rhee Peter , Sadegh-Nasseri Scheherazade , Shastri Nilabh 
Published: February 2023

Abstract
The aminopeptidase ERAAP, the ER aminopeptidase associated with antigen processing, is essential for trimming of peptides presented by classical and non-classical MHC class I molecules. The function of ERAAP is monitored by semi-invariant CD8+ T cells that recognize the QFL ligand. The QFL ligand consists of a 9-mer peptide encoded by the highly conserved Fam49a/b genes and is presented by non-classical Qa-1bmolecules expressed on surface of ERAAP-deficient cells only. Recently, Fam49b is identified as a novel negative regulator of TCR signaling, that functions by binding to active rac-1 and modulates reorganization of cytoskeleton. However, the function of Fam49b in T cells development has remained unclear. We show that Fam49b-deficient thymocytes receive higher TCR signaling and are defective in T cell development. Our studies reveal that Fam49b plays an important role in regulating thymocytes development and lineage commitment of conventional T cells as well as agonist selected T cells such as CD8αα αβ T cells. [This research was supported by NIH grants R01 AI130210 and R37 AI060040]
Unusual distribution of a non-classical MHC I restricted CD8+ T population
Authors: Guan Jian , Sadegh-Nasseri Scheherazade , Shastri Nilabh 
Published: February 2023

Abstract
A broad repertoire of peptides is constitutively presented through the MHC I molecules on the cell surface. ER aminopeptidase associated with Ag processing (ERAAP), a critical molecule in the antigen processing and presentation pathway, has profound effects on the generation of this peptide repertoire. Interestingly, the ERAAP deficient cells can be effectively detected by a unique population of the cytotoxic CD8+ T cell, named QFL-T cells which are specific for the ligand presented by the non-classical MHC Ib molecule, Qa-1b. The QFL-T cells showed similar characteristics with the other non-classical MHC Ib restricted unconventional T cells, such as the NKT or MAIT cells in their semi-invariant αβ TCRs, but their function(s) remains unknown. To study the potential function of the QFL-T cells, we assessed a variety of tissues for the ligand expression and the presence of the QFL-T cells. We observed high expression of this ERAAP dependent Qa-1b restricted ligand and relatively high frequency of the QFL-T cells in the small intestine. Remarkably, the number and the frequency of QFL-T cells among the intraepithelial lymphocytes (IELs) was tightly associated with the age and the gut microbiota of the mice. These findings suggest a role for gut microbiota in the homing/maintenance of the QFL-T cells in the small intestine.
Integration of affinity and abundance parameters for the identification of novel CD8+ T cell specificities in the CT26 tumor model
Authors: Arcia David , Boulanger Denise S. , James Edward N , Elliott Tim 
Published: February 2023

Abstract
The CT26 model is one of the most widely tested systems for cancer immunotherapy. Over 70% of the CD8+ CTL responses in CT26 tumors are directed against two peptides: SPSYVYHQF (AH1) presented by H-2Ld, or GGPESFYCASW (GSW11) presented by H-2Dd. Both peptides derive from the ecotropic murine leukaemia virus gp70 envelope glycoprotein, which is the highest expressed gene in CT26, and have relative low affinity values for their respective MHC-I. However, there is a lack of knowledge about other CTL specificities in CT26, the relationship between abundance and affinity of such peptides, and their roles in tumor rejection.
We identified 96 potential epitopes restricted by MHC-I molecules in BALB/c mice based on previously published immuno-transcriptomic data. We used a mathematical algorithm integrating the peptides relative affinity to MHC-I measured by NetMHC4.0, and their abundance in the proteome, which resulted in a prediction of the likelihood of peptide presentation.
Screening was performed in splenocytes from Treg-depleted CT26-challenged mice via IFNγ intracellular cytokine staining and T cell proliferation assays. Our approach led to the identification of a novel CTL specificity restricted by H-2Kd (Kd34), which shows moderate IFNγ responses in splenocytes from mice responding to the therapy. Moreover, using Kd34-specific dextramers, we were able to detect a small population of tumor-infiltrating lymphocytes (TILs) specific for this peptide, with similar phenotypic traits to AH1- and GSW1-specific TILs. The identification of such novel CD8+ T cell specificities will improve the understanding about the relationship between immunodominance and epitope abundance/affinity to better tailor vaccine strategies in cancer.
Dissociation of β2m from cell surface MHC class I triggers formation of noncovalent, transient heavy chain dimers
Authors: Springer Sebastian , Dirscherl Cindy , Löchte Sara , Hein Zeynep , Kopicki Janine-Denise , Harders Antonia , Linden Noemi , Weghuber Julian , Garcia-Alai Maria , Uetrecht Charlotte , Zacharias Martin , Lanzerstorfer Peter , Piehler Jacob 
Published: February 2023

Abstract
Major histocompatibility complex class I molecules (MHC I) that lose the antigenic peptide and the light chain beta-2 microglobulin (β2m) to become free heavy chains at the cell surface are known to associate, forming oligomers on the plasma membrane that are insufficiently understood. Here, we investigate the homotypic interaction of MHC I free heavy chains by combining a printed antibody micropattern assay with fluorescence recovery after photobleaching (FRAP) and with single molecule co-tracking in order to elucidate their molecular structure, abundance, and dynamics. We find that MHC I free heavy chain complexes are dimeric, transient, non-covalent, and mediated by the α3 domain. Free heavy chain interaction correlates with a decrease in the diffusion coefficient and an increase in the number of immobile molecules at the cell surface. Molecular docking and dynamics simulations suggest that in the complexes, the α3 domain of one FHC binds to another free heavy chain in a manner similar to the β2m light chain. We propose distinct functions of the MHC I free heavy chain dimers in signaling and in endocytic sorting.
Flow cytometry analysis of immune response to COVID-19 vaccine candidates using an in vitro stimulation model
Authors: Shi Xiaoshan , Bornheimer Scott J , Saksena Suraj , Widmann Stephanie , Tyznik Aaron 
Published: February 2023

Abstract
T cell immunity is vital for the control of viral infections. Cytotoxic CD8 T cells play a crucial role in eliminating virus-infected cells while CD4 helper T cells mediate host immune responses and promote B cells to secrete antibodies. For both, T cell recognition of viral antigens in the form of short peptides presented by HLA complexes is a prerequisite for T cell activation. Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has become a global pandemic that has disrupted economies and resulted in an enormous burden on health care systems. Despite the worldwide intensive efforts on development of COVID-19 therapeutics, a safe and effective COVID-19 treatment is not yet available. In this study, we established an in vitro stimulation system in which peripheral blood mononuclear cells were treated with putative SARS-CoV-2 immunogenic peptides. T cell immune responses were measured using different quantitative flow cytometry panels that were designed for evaluating cytokine production or identifying T cell subsets. Evaluation of host immune responses with a suite of flow cytometric panels may prove to be useful in the study of COVID-19.
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Male microbiota-associated metabolites restore macrophage efferocytosis in female lupus-prone mice via PPARγ and LXR signaling pathways
Authors: Harder James W , Ma Jing , Alard Pascale , Zhang Xiang , Yuan Fang , Kosiewicz Michele M 
Published: February 2023

Abstract
Sex and gut microbiota both influence systemic lupus erythematosus (SLE) development. We have found that male and lupus-prone female NZBxNZW F1 (BWF1) mice exhibit differences in gut microbiota and metabolomic profiles, and transfer of male microbiota suppresses disease in female mice. Here we determine how sex and microbiota may interact to affect SLE development. Transcriptomic analysis of female and male BWF1 spleens found phagocytosis-promoting genes were upregulated in males. Since defects in macrophage-mediated phagocytosis of apoptotic cells (efferocytosis) are found in SLE patients, we compared efferocytosis in vitro between male and lupus-prone female BWF1 mice. Macrophage-mediated efferocytosis was decreased in female compared to male BWF1 mice, but could be restored in female mice by transfer of male microbiota in vivo. Further transcriptomic analysis of spleens found that genes regulated by PPARγ and LXR receptor signaling were increased in male BWF1 mice. Our previous metabolomics analyses have found that two metabolites which can activate PPARγ and LXR signaling, phytol and its derivative phytanic acid, are higher in male BWF1 mice. We have also found that feeding phytanic acid to female mice delayed lupus onset. We show here that defective female BWF1 macrophage efferocytosis can be restored by phytanic acid treatment, and this requires both PPARγ and LXR signaling pathways. Furthermore, we found that phytanic acid may restore female BWF1 macrophage efferocytosis by upregulating the pro-efferocytic receptor, CD36. Taken together, our data indicate that BWF1 male microbiota produce metabolites that can enhance macrophage efferocytosis and suppress lupus.
The Contribution of Connexin 43 in Choroid Plexus to Experimental Autoimmune Encephalomyelitis
Authors: Reid Joshua , Manam Srikanth , Gadagkar Sudhindra R , Forsthuber Thomas , Nicholson Bruce J , Murthy Ashlesh K 
Published: February 2023

Abstract
Multiple Sclerosis (MS) is the most common progressive neurologic disease among young adults worldwide; yet the pathogenesis remains poorly understood. The myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalitis (EAE) in mice is a robust model and has been extensively used to understand the pathophysiology of MS. The expression of connexin 43 (CX43), a gap junction protein, has been shown to be enhanced in the choroid plexus during the development of acute EAE, with significantly increased CX43 expression at the peak of the disease. However, the contribution of such increased CX43 expression to the development of EAE has yet to be fully characterized. Using the Cre-Lox platform, we generated mice with conditional deletion of CX43 in the ciliated columnar epithelium of the choroid plexus (Foxj1Cre-CX43 KO mice; referred to as CX43 KO mice hereafter). We employed the MOG-induced EAE model in both male and female CX43 KO mice to understand the role of CX43 in the pathogenesis of EAE. Male CX43 and WT mice displayed comparable peak EAE clinical disease scores of 1.83±0.33 and 1.75±0.66, respectively. Female CX43 KO, however, displayed a significantly reduced peak EAE clinical disease score of 0.33±0.24 compared to 1.15±0.39 in female WT animals. These results suggest a role for CX43 in the ciliated columnar epithelium of choroid plexus to the development of EAE in female, not male, mice and underscore the need for further exploration of mechanism(s) underpinning the role of CX43 in the pathogenesis of EAE.
p38 MAP kinase signaling in microglia plays a sex-specific protective role in CNS autoimmunity and regulates microglial transcriptional states
Authors: Krementsov Dimitry N , McGill Mahalia , Richman Alyssa , Boyd Joseph , Frietze Seth 
Published: February 2023

Abstract
Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system. MS is three times more common in women, yet more severe in men. The mechanisms underlying these sex differences remain largely unknown. MS is initiated by autoreactive T cells, but CNS-infiltrating myeloid cells are key effector cells driving pathology. The role of CNS-resident microglial cells is less clear. We have previously shown that genetic ablation of p38α MAP kinase broadly in the myeloid lineage is protective in the autoimmune model of MS, experimental autoimmune encephalomyelitis (EAE), but only in females, and not males. Here, we sought to define the mechanisms responsible, using genetic approaches and bone marrow chimeras to ablate p38α in microglial cells, peripheral myeloid cells, or both. Deletion of p38α in both cell types recapitulated the previous sex difference, with reduced EAE severity in females. Unexpectedly, deletion of p38α in the periphery was protective in both sexes. In contrast, deletion of p38α in microglia exacerbated EAE in males only, revealing opposing roles of p38α in microglia vs. periphery. Single-cell RNAseq revealed a diversity of microglial states, connected by distinct convergent transcriptional trajectories. Microglial p38α deficiency resulted in enhanced transition from homeostatic to disease-associated microglia, with downregulation of regulatory genes such as Atf3, Rgs1, and Btg2, and upregulation of inflammatory genes such as Cd74, Trem2, and MHC class I and II. These results reveal a p38α-dependent male-specific molecular pathway in microglia that is protective in CNS autoimmunity, suggesting that autoimmunity in males and females may be driven by distinct cellular and molecular pathways.
Immune-neural transmission regulates gut sensation
Authors: Wu Chuan , Chen Zuojia , Luo Jialie , Li Jian , Kim Girak , Chesler Alexander , Urban Joseph , Trinchieri Giorgio , Li Wei 
Published: February 2023

Abstract
The gastrointestinal tract is known as the largest endocrine organ that encounters and integrates various immune stimulations and neuronal responses due to constant environmental challenges. Enterochromaffin (EC) cells, which function as chemosensors on the gut epithelium, are known to translate environmental cues into serotonin (5-HT) production, contributing to intestinal physiology. However, how immune signals participate in gut sensation and neuroendocrine response remains unclear. Interleukin-33 (IL-33) acts as an alarmin cytokine by alerting the system of potential environmental stresses. We here demonstrate that IL-33 induced instantaneous peristaltic movement and facilitated Trichuris muris expulsion. We found that IL-33 could be sensed by EC cells, inducing release of 5-HT. IL-33-mediated 5-HT release activated enteric neurons, subsequently promoting gut motility. Mechanistically, IL-33 triggered calcium influx via a non-canonical signaling pathway specifically in EC cells to induce 5-HT secretion. Our data establish an immune-neuroendocrine axis in calibrating rapid 5-HT release for intestinal homeostasis.
SARS-CoV-2 related cytokines induce barrier function changes in human aortic endothelial cell monolayers: Implications for cytokine storm and vascular dysfunction.
Authors: Campbell Carmen Elaine , Boisvert William 
Published: February 2023

Abstract
In 2019, a new strain of coronavirus, severe acute respiratory syndrome–coronavirus 2 (SARS-CoV-2), emerged in Wuhan, China. Nearly 25 million cases have since been reported in the US and almost 500,000 Americans have died. The aggressive inflammatory response of the host to SARS-CoV-2 infection plays a significant role in disease severity. Severe SARS-CoV-2 infection often leads to Acute Respiratory Distress Syndrome (ARDS), Severe Inflammatory Response Syndrome (SIRS), and Cytokine Release Syndrome (CRS). Observed in about a third of fatal COVID-19 patients, CRS is correlated to high mortality. Patients experiencing CRS also suffer from systemic vascular dysfunction. Thus, methods to assess endothelial cell function may prove essential to evaluate effectiveness of therapeutic interventions targeting cytokines. Our objective was to characterize dynamic responses of vascular endothelial cell monolayer barrier resistance (Rb) when treated with COVID-19 related cytokines. We quantified the effect of eight different cytokines to induce Rb changes in human aortic endothelial cell (HAoEC) monolayers using Electric Cell-substrate Impedance Sensing. We found that Rb of HAoEC monolayers was significantly reduced when treated with TNF-α, IL-1β, or IFN-γ compared to untreated controls. In the case of TNF-α, no significant difference in Rb was measured in cell monolayers treated with TNF-α and anti-TNF-α monoclonal antibody. On the other hand, anti-IL-1β antibody did not block the effect of IL-1β which induced a significant reduction in Rb compared to cytokine free controls. A significant reduction in Rb values were also noted in cell monolayers treated with TNF-α and IL-1β combinations suggesting a systemic synergistic effect.
Activation of Lyn kinase and translocation of its PKCδ target regulate phosphorylation in the IL-4-induced alternate pathway for BCR signaling
Authors: Khan Naeem , Hu Yongmei , Lowell Clifford A , Rothstein Thomas L 
Published: February 2023

Abstract
B cell signaling for activation via the BCR occurs as an isolated event only in vitro; in real life, BCR signaling takes place within a complex milieu that involves interactions with agents that trigger additional receptors. Chief among these agents is IL-4. We have shown that BCR signaling is reprogrammed by IL-4 receptor engagement, and that this reprogramming involves creation of a new, signalosome-independent, Lyn-dependent alternate signaling pathway out of pre-existing, otherwise noninteracting, elements. A unique aspect of alternate pathway BCR signaling is PKCδ phosphorylation. In dissecting this pathway, we unexpectedly found that IL-4 alone induces phosphorylation and activation of Lyn, and that Lyn immunoprecipitated from IL-4-treated B cells capably phosphorylates PKCδ in a cell-free system, although PKCδ phosphorylation does not occur in the absence of BCR triggering in vivo. This raised the question of why IL-4 alone failed to produce PKCδ phosphorylation. We considered the possibility that Lyn and PKCδ may be spatially separated. As expected, Lyn is located primarily in the membrane fraction, whereas PKCδ is located mainly in the cytosol fraction. However, when anti-Ig follows IL-4 treatment, PKCδ is found in the membrane fraction and phosphorylated. This translocation of PKCδ to the membrane fraction is not affected by loss of Lyn, although PKCδ is not phosphorylated without Lyn. Thus, PKCδ phosphorylation through the alternate pathway appears to consist of several discrete steps: 1.IL-4 induces Lyn phosphorylation and activation; 2. IL-4 produces a change such that PKCδ is translocated from the cytosol to the membrane after BCR engagement; 3. Activated Lyn in the membrane phosphorylates PKCδ in the membrane.
Perturbations of Marrow Stromal Cell function during Acute Inflammation
Authors: Matkins Victoria , Camacho Virginia , Hoang Ashley , Patel Sweta , Welner Robert S 
Published: February 2023

Abstract
Mesenchymal stromal cells (MSCs) reside in a complex network of blood cells and can differentiate into mature chondrocytes, osteoblasts, and adipocytes. This trilineage differentiation process is tightly regulated by intrinsic and extrinsic signals, leading to transcriptional programming of balanced lineage output. Inflammatory insults to this stromal homeostatic balance occur through the upregulation of activating cytokines. Pro-inflammatory cytokines’ impact on the hematopoietic lineage is well-defined, but similar impacts to stroma was lacking. To define these changes, we studied two well-characterized forms of inflammation: polyinosinic-polycytidylic acid (Poly(I:C)), that activates IFNα, and Lipopolysaccharide (LPS), that through TLR4 promotes IFNγ. Prior studies of inflammatory conditions show bone loss; therefore, we hypothesized that there will be an increase in the MSC pool to compensate for loss of bone maturation. Using flow cytometry and lineage tracing models for stroma, we phenotypically characterized the non-hematopoietic cells’ changes following perturbation. The lineage-specific Cre models used were stroma (Prx1), adipocytes (AdipoQ), and osteoblasts (OCN). Our data shows an increase in MSCs with decreases in osteoblasts and adipocytes with stimulation. Furthermore, we used reverse-phase protein array (RPPA) and cytokine arrays to map the signaling mechanisms and activated pathways. We found an increase in specifically MAPK pathway with an accompanying increase in cytokines linked to this pathways’ activation. Understanding changes in the bone marrow microenvironment during inflammation will allow for therapeutic intervention in mediating hematopoietic diseases such as leukemia.
The subunits of IL-12, originating from two distinct cells, can functionally synergize to protect against pathogen dissemination in vivo
Authors: Gerber Allison N , Singh Nevil J 
Published: February 2023

Abstract
Cytokines are typically single gene products that potently modify immune responses. The IL-12 family is an exception to this paradigm as these cytokines are heterodimers. In the case of IL-12, the subunits, IL-12p40 and IL-12p35 are encoded by two unlinked genes. However, it is canonical that activated myeloid cells co-express both these subunits simultaneously and secrete a fully active heterodimer of IL-12 which then drives IFNγ production by innate and adaptive cells. Using a chimeric mouse model, we identified a novel method of formation for generating functional IL-12 activity in vivo involving IL-12p40 monomers originating from hematopoietic cells which extracellularly associate with IL-12p35 from other non-hematopoietic cells. In the context of a parasitic challenge, we found that this two-cell mechanism of IL-12 formation was sufficient to drive T cell differentiation in local sites, distal to the initial infection, and thereby regulate systemic dissemination of the pathogen. Accordingly, we identify a new collaboration between hematopoietic and non-hematopoietic cells to produce functional IL-12 through the regulated release of individual subunits as a systemic alert system to prepare host tissues for potential pathogen arrival.