bluuebunny/crossref_metadata_2025_split huggingface dataset:
Investigating the role of CX3CR1 in a GITRL- dependent signal 4 checkpoint during LCMV clone 13 infection
Authors: Chang Yu-Han , Wang Kuan-Chung , Watts Tania H. 
Published: January 2023

Abstract
T cell-APC interactions early during chronic viral infection are critical for determining viral set point and disease outcome. Our recent work (Chang et al. Immunity. 2017) revealed a division of labour whereby monocyte-derived APCs (infAPCs), with low MHC II and CD80/86 expression compared to classical DCs (cDCs), preferentially up-regulate TNF receptor superfamily ligands, GITRL, 4-1BBL, OX40L and CD70, in response to type I interferon (signal 3) to provide a post-priming checkpoint (signal 4) for CD4 T cell responses. While cDCs contribute to signal 1 & 2 during priming in response to LCMV clone 13, GITRL on infAPCs binding to GITR on CD4 T cells results in increased surface expression of pro-survival receptors, including CD25, CD127 and OX40 to sustain Th1 helper cell accumulation, help for virus-specific CD8 T cell responses and viral control. Interestingly, CX3CR1, the receptor for fractalkine, was also upregulated on CD4 T cells by GITR signalling during clone 13 infection. CX3CR1 was recently shown to contribute to tumor-infiltration of CD4-helped CD8 CTLs (Ahrends et al. Immunity. 2017). In the chronic LCMV model, we observed that CX3CR1high virus-specific CD4 T cells in the spleen exhibit a markedly increased effector profile when compared to their CX3CR1int-low counterparts, with both higher frequency and per cell expression of Tbet and IFNg production. To investigate how CX3CR1 on CD4 T cells contributes to CD4 T cell help for the CD8 T cell response, we have crossed CX3CR1 deficient mice with TCR transgenic SMARTA mice, expressing an MHC II-restricted TCR specific for LCMV gp61–80. Overall, our data so far indicate that upregulation of CX3CR1 marks the most activated splenic CD4 Th1 effectors during chronic LCMV infection.
Aryl hydrocarbon Receptor Activation in DCs Regulates thymic Function and Immune Tolerance Induction.
Authors: Cole Shelby L. , Shepherd Celine Ann Beamer , Kreitinger Joanna , Shepherd David M. 
Published: January 2023

Abstract
The thymus is a primary immune organ that mediates central tolerance through proper education of developing thymocytes and contributes to peripheral tolerance via the production of thymic regulatory T cells (tTregs). Differentiation of thymocytes involves interactions with both thymic epithelial cells and antigen presenting cells including thymic dendritic cells (tDCs). The thymus is highly sensitive to dioxin, an environmental pollutant that is the prototypical ligand for the Aryl hydrocarbon receptor (AhR) and can induce severe thymic atrophy; however, the underlying mechanisms and ultimate consequences of AhR-mediated thymic atrophy are not currently defined. Moreover, determining how AhR activation in thymic DCs affects thymopoiesis and tTreg induction will further enhance our understanding of immune tolerance. We hypothesized that AhR activation in DCs blocks thymopoiesis and induces tTregs via increased CTLA-4 expression on tDCs. Using CD11ccre x AhRfx(AHRCD11c) conditional knockout mice, we observed that AhR deficiency in CD11c+ tDCs blocked dioxin-induced thymic atrophy and the generation of tTregs, an effect that occurred with other AhR ligands including ITE. Moreover, elevated CTLA-4 mRNA and protein expression was detected in CD11c+ tDCs suggesting a possible mechanism underlying these effects. Together, these findings support the premise that AhR activation in CD11c+ thymic DCs is a critical regulator of thymopoiesis, and the generation of central and peripheral immune tolerance. This research was supported by R01ES013784 (DMS) and NIH Diversity Supplement (SLC).
Complement 5a galvanizes immune inflammation in Hunter syndrome
Authors: Pandey Manoj K. , Prada Carlos Enrique , Magnusen Albert Frank , Nyamajenjere Tsitsi Carol , Mckay Mary Ashley , Rapien Jillian , Köhl Jörg 
Published: January 2023

Abstract
Hunter syndrome, is a lysosomal storage disease (LSD) caused by mutation in IDS that elicits deficiency of iduronate-2-sulfatase. IDS defect initiates excess storage of glycosaminoglycans (GAGs), i.e., heparan sulfate (HS) and dermatan sulfate (DS) in almost every cell of different organs, (e.g., liver, spleen, lung, heart, bones, skin, brain, and nervous system). Affected individuals exhibit abnormal appearance, mental development, organ function, physical skills and death at the age of 10–20 years. Mechanism by which such GAGs propagate disease in Hunter syndrome is wanted. We directly compared inflammatory responses to DS in different groups of BALB/c mice by injecting (i.p./day for 4 weeks) them separately with altered doses of DS, (0, 1, 2, 4, and 8 mg). Complement 5a (C5a) and their receptor (C5aR1) were measured in mice sera and monocytes. As compared to vehicle (PBS), DS treated mice sera and monocytes showed dose dependent increases of C5a and C5aR1. Human patients with Hunter syndrome also showed elevated serum level of C5a when compared to healthy controls. C5a-C5aR1 axis has been valued for increased staffing of immune cells and their venomous influence on tissue impairment in LSD and non LSDs. To explore this risk in Hunter syndrome, mice were injected in presence or absence of indicated amounts of DS and circulatory subsets of immune cells were measured. As compared to vehicle, DS treated mice showed dose dependent rises of monocytes, eosinophils, and neutrophils and their link to excess generation of pro-inflammatory cytokines. Our data showed that DS-induced C5a-C5aR1 axis wave the immune inflammation, which is fatal in Hunter syndrome. Targeting C5a/C5aR1 axis could be potential alternative therapy of Hunter syndrome.
Calcineurin positively regulates TCR-induced LFA-1-dependent cell adhesion via dephosphorylation of Lck Ser-59.
Authors: Otsuka Shizuka , Dutta Debjani , Mittelstadt Paul R. , Ashwell Jonathan D. 
Published: January 2023

Abstract
T cell adhesion via the interaction of the integrin leukocyte function-associated antigen-1 (LFA-1) with its ligand ICAM-1 is essential for the initiation of immune response. Upon ligation of T cell receptor (TCR), activation-induced LFA-1 conformational change occurs via phosphorylation of Thr-758 resulting in the enhancement of binding to ICAM-1. Recently, we reported that dephosphorylation of Lck Ser-59 (LckS59) by calcineurin results in enhanced TCR-proximal signaling, indicating that phosphorylated LckS59 negatively regulates T cell activation. Here we find that inhibition of calcineurin activity by immunosuppressive drugs such as cyclosporin A (CsA) rapidly reverses existing TCR-induced LFA-1-dependent cell adhesion as well as the phosphorylation of LFA-1 Thr-758 in Jurkat and primary human T cells. In addition, TCR-induced cell adhesion of T cells from LckS59A knock-in mice was stronger than T cells from LckWT mice. Consistent with enhanced cell adhesion, some of the signaling events downstream of TCR proximal signaling were also enhanced in LckS59A mouse T cells. Notably, LckS59A T cell adhesion was resistant to CsA inhibition. These observations provide new insights into a previously unrecognized role for calcineurin inhibitors in reversing ligand-dependent adhesion independently of its effects on NFAT activation.
AID phosphorylation regulates MMR during class switch recombination
Authors: Vuong Bao Q. , Choi Jee Eun , Matthews Allysia , Michel Genesis 
Published: January 2023

Abstract
Processing of AID (activation-induced cytidine deaminase)-dependent dU:dG mismatches by the base excision repair (BER) and mismatch repair (MMR) pathways generates untemplated mutations during somatic hypermuation (SHM) or DNA breaks during class switch recombination (CSR). Genetic deletion of UNG or MSH2, which are essential for BER or MMR, respectively, impairs CSR and alters the spectrum of mutations in SHM. Ablation of both UNG and MSH2 completely blocks CSR and limits SHM to GC transition mutations. Phosphorylation of AID at Ser38 (pS38-AID) is required for wild-type levels of CSR and SHM. B cells with a homozygous knock-in mutation of the AID phosphorylation site (AIDS38A/S38A) have intermediate levels of CSR and SHM as compared to wild-type and AID−/− B cells. AID phosphorylation is hypothesized to regulate BER through the interaction of pS38-AID with APE1, which functions downstream of UNG. Consistent with this hypothesis, AIDS38A/S38AMSH2−/− B cells have a complete block in CSR and significantly impaired SHM of Sμ and the JH4 intron. Surprisingly, CSR is also completely blocked in AIDS38A/S38AUNG−/− B cells; however, these cells generate significant mutations in Sμ and JH4 intron. These data demonstrate a critical role for pS38-AID in regulating BER and MMR and uncover a novel pS38-AID-dependent mechanism of mediating MMR-dependent DNA break formation and/or ligation during CSR.
The role of ERK2 in germinal center B cell selection
Authors: Negron Austin Joseph , Jeffreys Sean Alec , Forsthuber Thomas 
Published: January 2023

Abstract
The regulation of programmed cell death is a crucial control point for the maintenance of B cell tolerance in the germinal center (GC) and involves the coordination of several intracellular signaling pathways. Dysregulation of these pathways can lead to the survival of autoreactive B cells, resulting in autoimmune disease. The mitogen-activated protein kinase (MAPK) pathway is involved in the transduction of both survival and apoptotic signals received through the B cell receptor (BCR) and the Fas receptor (CD95), respectively. Extracellular signal regulated kinases 1 and 2 (ERK1/2) are important effectors of this pathway and play a major role in the B cell selection process. Though it is known that ERK2 specifically regulates the pro-apoptotic protein, Bmf, its function in GC B cells remains poorly understood. Our lab has developed a novel mouse model in which the deletion of Erk2 is accompanied by the expression of the fluorescent reporter protein, eYFP, allowing us to identify and isolate viable Erk2 Δ B cells using flow cytometry. Preliminary data using this model has shown that Erk2Δ B cells exhibit impaired survival ex vivo and express several activation-associated surface markers. This suggests that ERK2 plays a major role in facilitating GC B cell selection. Thus we hypothesize that Erk2 Δ B cells are impaired in their ability to participate in the GC reaction. We are currently investigating the ability of Erk2 Δ B cells to respond to activating and pro-apoptotic stimuli in vitro as well as to differentiate in to antibody-secreting plasma cells. Our results will contribute to a better understanding of the molecular programs involved in B cell selection, potentially offering new approaches to treating B cell-driven autoimmune diseases.
Low NK Cell ADCC as a Risk Factor in Chronic Fatigue Syndrome (CFS): Familial Risk for CFS or Differences between Human Populations?
Authors: Hudig Dorothy , Tang Jennifer , Sung Alexander P. , Guglielmo Michael J. , Smith-Gagen Julie , Bateman Lucinda , Redelman Douglas D. 
Published: January 2023

Abstract
ADCC (antibody-dependent cell-mediated cytotoxicity) requires the cytotoxic subset of NK cells that can also recognize antibody. In vivo function may depend on the number of these ADCC NK cells. We report on ADCC activity and the number of IgG FcR-positive NK cells/ul blood as risk factors for chronic fatigue syndrome (CFS). ADCC capacity was measured as CX1:1 (PMID 29113954), the % Daudi B tumor cells killed at a 1:1 ratio of CD16A FcR-pos CD56pos NK effector cells to Daudi targets with obinutuzumab anti-CD20. CFS patients met the 1994 Fukuda diagnostic guidelines. We examined 5 families with 2 or more patients per family, comparing patients with their non-CFS family members and with unrelated healthy controls. ADCC activity and effector cell counts were similar between CFS patients and their family members. Surprisingly, the CX1:1 of both the CFS patients and their non-CFS family members was low, 77% of the controls (P=0.03). The ADCC effector cell counts/ul blood were also lower for both (two-tailed P=0.09, insignificant). The potential for synergistic interaction of low CX1:1 and low effector cell counts was 24-fold greater for CFS family members than for unrelated healthy controls (P&lt;0.05). The ADCC similarity between patients and family members indicates that lower ADCC is unlikely to influence who is affected with CFS within families. The combined lower ADCC and lower effector cell counts of CFS family members could indicate a familial risk for CFS. However, caution is warranted by evidence that the CFS families are from a Utah population that is genetically skewed, while the controls have diversity representative of the USA. The ADCC differences could be between 2 human populations or between people more or less likely to suffer from CFS.
Different bactericidal and inflammatory activities of human and mouse blood
Authors: Lin Tian , Moorlag Simone J.C.F.M. , Liu Jialin , Ahmed Mohamed Yehia Hassan , Thundivalappil Sujatha R. , Riley Frank E. , Warren H. Shaw 
Published: January 2023

Abstract
Mouse models often serve as a selective gate for drugs developed to treat patients with inflammation and sepsis. However, mice are much more resistant than humans to intravenous challenge with LPS, a cell wall component of Gram-negative bacteria. It is common to detect up to 106 CFU/ml of live bacteria in the blood in many mouse models of bacterial challenge but less than 100 CFU/ml of bacteria in the blood of human adults with Gram-negative sepsis. These substantial differences between mice and humans during infections are unexplained, unstudied and usually ignored.
We studied and compared the growth of four encapsulated, clinically relevant strains of Escherichia coli (O4:K54:H5, O16:K1:H6, O18:K1:H7, O25:K5:H1) and Pseudomonas aeruginosa 12.4.4 in fresh human and mouse blood at 37°C for 30, 90 and 180 minutes. We also measured TNF and IL-6 levels in the supernatants, and performed a phagocytosis assay by using fluorescein isothiocyanate-labeled heat-killed bacteria incubated with human and mouse blood followed by flow cytometry. We found that E. coli and P. aeruginosa multiplied in mouse blood and plasma, but that the colony counts of each were decreased in human blood and plasma. More E. coli and P. aeruginosa were phagocytosed by leukocytes in human than those in mouse blood, but this difference reflected larger numbers of neutrophils in human blood than in mouse blood. Large amounts of pro-inflammatory cytokines TNF and IL-6 were detected in human but not in mouse blood after live bacterial inoculation. In conclusion, our results suggest that bacterial killing, phagocytosis, and cytokine induction in the blood compartment of patients with Gram-negative bacteremia may not be well mimicked in mouse models of bacterial challenge.
Histone h4 as a trigger for innate immune cells.
Authors: Hsieh I-Ni , White Mitchell , De Luna Xavier , Hartshorn Kevan L. 
Published: January 2023

Abstract
Extracellular histones have been considered to be responsible for several systemic inflammatory injuries, including lung, kidney and liver. We now show that histone H4 can directly activate neutrophil respiratory burst response, depolarization, and myeloperoxidase (MPO) release. In addition, histone H4 increased neutrophil adhesion activity and the expression of adhesion molecule. More interestingly, we found that histone H4 also induced a sustained rise in the intracellular calcium level in neutrophils, and that H4 increased caspase 3 activity in neutrophils over time, which can be inhibited by anti-H4 antibody treatment. Histone H4 induced IL-8 release from neutrophils and is also a strong inducer of IL-1 beta and TNF-alpha productions in monocytes and monocyte-derived macrophages. To counteract the effects of histone H4, we found that C reactive protein (CRP) and/or surfactant protein D (SP-D) could reduce histone H4-induced respiratory burst response, calcium flux, MPO release and caspase 3 activity in neutrophils. These findings suggest that extracellular histone H4 may play important roles in the innate immunity, and some endogenous peptides may have the ability to reduce H4-induced effects. This study will hopefully provide new treating targets for treating inflammatory diseases and novel strategies based on modification of CRP or SP-D with optimal histone binding site for treatment of severe infections.
Real-time visualisation and quantification of Neutrophil Extracellular Traps
Authors: Lovell Gillian F. , Bevan Nicola , Dale Tim , Trezise D. J. 
Published: January 2023

Abstract
The ability of neutrophils to release extracellular traps (NETs) is one of several mechanisms by which the body defends against infection. When neutrophils encounter invading pathogens, the cells release a mixture of antimicrobial proteins and chromatin to trap and degrade microbes. NETs have been implicated in a number of disorders including atherosclerosis, systemic lupus erythematosus (SLE) and thrombosis.
NETosis can be stimulated in vitro using a number of methods including chemical compounds, microbes and microcrystals. In order to better understand the signaling pathways involved, we developed a fully kinetic live-cell imaging assay for NETosis (96-well format, IncuCyte S3). Using a fluorescent cell impermeant DNA-binding reagent (IncuCyte CytoTox Green), NET release was visualised and quantified in real time. In both primary human neutrophils and differentiated ‘neutrophil-like’ dHL60 cells, PMA (100nM) induced rapid (2h onset, peak 4–6h), time-dependent increase in fluorescence and nuclear degradation. A concomitant increase in myeloperoxidase, elastase (immunofluorescence) and cell-free DNA (Picogreen) was observed, validating the NETosis signal. PMA-induced NETosis was ROS-dependent (CellRox Red) but did not cause externalisation of phosphatidylserine (PS, Annexin V). In contrast, ionomycin (5mM) induced NETosis more rapidly (onset time &lt;15′), did not induce ROS but did externalise PS. Neutrophils and dHL60 cells were both able to phagocytose bacterial bioparticles (IncuCyte pHrodo-E-coli). Taken together, these data illustrate how NETs and neutrophil signaling pathways can be robustly, meaningfully and efficiently analysed using automated live-cell imaging and compatible detection reagents.
Cardiac fibroblast pretreated by ANG II induced M1 phenotype macrophage selective apoptosis through TNF-TNFR1 and survival M1 conversion into M2 depended on Leptin-PI3K-Akt pathway
Authors: Su Zhaoliang , Lu Hongxiang , Shao Xiaoyi , Chen Rong , Tian Yu , Zong Gangjun , Xu Huaxi 
Published: January 2023

Abstract
Inflammatory monocytes and tissue-resident macrophage are key regulators of tissue injury, would healing, organ remodeling and regeneration. After tissue injury, most monocytes and macrophages infiltrate into injury sites, undergo phenotypic and functional changes: differentiation into proinflammatory M1 phenotype and anti-inflammatory M2 in milieu. The reprogramming M1 and M2 macrophages involve in inflammatory initiation, development, resolution, and organ function remodeling, respectively. Disturbances in macrophage reprogramming can lead to aberrant repair or uncontrolled inflammatory response. Timely modulation of the reprogramming of macrophages is essential for the treatment of inflammatory diseases. Therefore, it becomes fatal to clarify the M1 fate or the function shift of M1 and M2 during tissue injury development. In the present work, we demonstrated that cardiac fibroblasts (MCF) pretreated by angiotensin II (ANG II) (myofibroblast, MFB) could not only promote macrophage reprogramming toward M1 but also induced M1 macrophage selective apoptosis through TNF/TNFR1 axis independent on Fas/FasL; furthermore, in co-culturing system, leptin produced by MFB or M1/M2 cells could promote the survival M1 conversion into M2 macrophage via PI3K/Akt pathway. In vivo, TNF-alpha blockade significantly ameliorated experimental autoimmune myocarditis (EAM) development. Our data highlighted a novel mechanism about the fates of M1 macrophage in inflammatory diseases and our data also suggested a potential check point for inflammatory disease therapy target on macrophage.
Efficient Enrichment of Functional ILC Subsets from Human PBMC by Immunomagnetic Selection
Authors: Valdez Yanet , Kyei Stephen K. , Poon Grace F.T. , Kokaji Andy , Woodside Steven M. , Eaves Allen C. , Thomas Terry E. 
Published: January 2023

Abstract
Innate lymphoid cells (ILCs) are exceedingly rare but important regulators of homeostatic and disease-associated immune processes. The frequency of ILCs in peripheral blood of healthy humans is ~0.07% of CD45+ leukocytes. ILCs lack specific cell surface markers but can be divided into distinct subsets (ILC1, 2 and 3) based on their differential expression of effector cytokines and master transcription factors.
Currently, cell sorting is the most widely used method to isolate ILCs, but it is time consuming, expensive and often results in low purities and recoveries. Pre-enrichment of ILCs would allow for reduced sorting times and improved purities. Accordingly, we have developed a fast immunomagnetic negative selection method to pre-enrich all ILCs subsets from human leukapheresis samples. Briefly, unwanted cells are labelled with antibodies and magnetic particles and placed into an EasySep™ magnet. Unwanted cells are retained in the magnet and the enriched ILC fraction is simply poured off into a new tube. We find that total ILCs (defined as Lineage− CD45+ CD127+) are enriched from a frequency of 0.01 – 0.23% (n=28) to a final frequency of 17 – 86%, an enrichment of 200 – 1500 fold with virtually no loss of ILCs. ILC1 were enriched from 0.01 – 0.2% to 4.5 – 14%. ILC2 were enriched from 0.01 – 0.1% to 5.8 – 51% and ILC3 were enriched from 0.01 – 0.1% to 6 – 16%. This pre-enrichment drastically decreases sort time, allowing sorting over 3.7 × 105 ILCs from 2 × 109 PBMCs in only 12 minutes. Sorted cells maintained their functionality; when stimulated, ILC1s produced IFNγ, ILC2s secreted IL-13 and ILC3s produced IL-22. Our newly developed method of ILC pre-enrichment should aid human ILC research by enabling their rapid isolation when combined with cell sorting
Complement controls antiviral CD8+T cell memory development and function
Authors: Jhun Kevin S. , van der Heide Verena , Heeger Peter S. , Homann Dirk 
Published: January 2023

Abstract
Control of infectious disease is predicated on the combined efforts of both innate and adaptive immune defenses including the generation of pathogen-specific CD8+effector T cells (CD8+TE). Upon resolution of inflammation and pathogen clearance, the CD8+TE population contracts and the preferential survivors mature into CD8+TM, which provide further protection against recurrent infection. In this context, the precise properties of CD8+TM populations emerge as a consequence of the unique conditions during initial T cell priming and thus make the generation of T cell memory contingent upon a variety of co-stimulatory and -inhibitory interactions. Here, we demonstrate a non-redundant role for complement, an innate immune defense pathway, in the development of CD8+T cell memory generated in response to an acute lymphocytic choriomeningitis virus (LCMV) infection. Mice deficient for the complement component 3 (C3) mount a relatively normal LCMV-specific CD8+T cell response, yet the subsequent development of CD8+T cell memory is compromised by impaired phenotypic, metabolic and functional CD8+TM maturation. Interestingly, the contraction phase is also impaired which results in an increased abundance of CD8+TM. Despite this observation, C3-deficient CD8+TM exhibit a decreased capacity to undergo in vivo secondary expansion when adoptively transferred into naïve hosts for rechallenge. We propose that C3, but no other individual complement component evaluated (C3aR, C5aR, and DAF/CD55), exerts a unique co-stimulatory function that generates a balanced formation of CD8+T cell memory and recall responses.