bluuebunny/crossref_metadata_2025_split huggingface dataset:
Solutions for quantifying cytopathic effects after viral infection in vitro
Authors: Preyer Rosemarie , Kevin Strecker , Gladyschuk Vitaly , Klimovich Alexei , Ettischer-Schmid Nicole 
Published: November 2024

Abstract
Individual virological assays require a maximum of experience. For different experiments, we introduce an outstanding technology for evaluating them in a reliable way.
In viral infections in vitro, the cytopathic effect (CPE) occurs. In the beginning, only few cells show a CPE, which makes it different to estimate the success. Quanti-and qualification of CPE in stages 1-4 of cells can be helpful to assess the right timepoint for different approaches.
With the AID MultiSpot, an inverted microscope and a fluorescence EliSpot reader combined, the CPE can be monitored and quantified without cellular staining and w/o interrupting any experiment with liquid like medium.
Further assays like plaque assays are performed in platesfrom 6-384 wells, stained either with colorings like crystal violet or, mostly in 96-well format, with fluorescent dyes. The reader type for these (and many more) applications is the vSpot spectrum.
We treated cells with DMSO (Fig.1) or virus (Fig. 2), measured the CPE or counted the plaques. As the MultiSpot is automated, one well can be scanned that the whole well view is possible in magnifications from 4- to 20-fold. The vSpot is able to generate whole-well pictures from all plate formats due to its unique composition.
These readers enable to quantify easily the CPE to get an overview of the whole wells that no important event is missed. This can be done in order to relate different treated cells and to make results between different experiments comparable.
Immunophenotypic profiling reveals novel T cell subset associated with “tolerance” in pediatric transplant recipients
Authors: Zhang Wenming , Rao Mahil , Amouzgar Meelad , Reitsma Andrea , Pena Josselyn , Nova Patricia , Shah Anika , Bendall Sean , Esquivel Carlos , Martinez Olivia , Krams Sheri 
Published: November 2024

Abstract
Achieving allograft health remains a challenge in pediatric organ transplantation. Recently. we performed multilineage, single-cell analysis of the immune cell composition in pediatric solid-organ transplant recipients who received a heart, liver, or kidney allograft (n=52); 24 children had stable graft function while 28 children had a rejection episode. Hierarchical clustering of the mean cell type proportion across allograft types and health states demonstrated a distinct subpopulation of CD4 T cells (CD45RA-,CD25+,CD5+,CD38-) was significantly (p&lt;0.05) associated with stable graft function and were termed TOT cells. The cosine similarity analysis of proteomics data showed that TOT cells are phenotypically most like Tregs and least like CD4+ TEMRA. To characterize the TOT cells, we developed an 8-marker flow cytometry panel to specifically sort Treg and TOT populations and performed 10x single-cell RNA-seq and TCR-seq with subsequent analysis in R and Loupe Browser. Differentially expressed genes that were specifically associated with TOT cells included ANXA1, IL7R, TCF7, THEMIS, NELL2, KLRB1 and CD40LG while Treg expressed FOXP3, IKZF2, CTLA4, IL2RA, and RTKN2, consistent with their known function in immune suppression. TCR analysis indicates unique expansions of T cell clonotypes in TOT cells not seen in Treg. Further investigation into the immunoregulatory capacity of TOT cells could be instrumental in developing new strategies to prolong graft survival.
KCa3.1 channels regulate NK cell tumor infiltration in head and neck cancer
Authors: Alshwimi Abdulaziz , Chimote Ameet , Weatherford Layne , Wulff Heike , Khodoun Marat , Wise-Draper Trisha , Conforti Laura 
Published: November 2024

Abstract
NK cells are important mediators of antitumor immunity in head and neck squamous cell carcinoma (HNSCC) where they need to infiltrate the tumor and make contact with the cancer cells in order to kill them. However, low NK cell tumor infiltration has been associated with poor outcomes and immunotherapy resistance. Previously we have shown that KCa3.1 channels are important regulators of Ca2+ fluxes and chemotaxis in T cells (Chimote et al., Science Signaling 2018). However, little is known about the role of KCa3.1 in NK cell chemotaxis. In vitro studies with human NK cells activated with IL2/IL15 showed that the KCa3.1 selective blocker TRAM-34 inhibited chemotaxis and random migration, while pharmacological KCa3.1 activation increased chemotaxis. In vivo experiments with a humanized HNSCC mouse model generated by implantation of Cal27 cells (a human HNSCC cell line) in immune-deficient NSG mice, followed by injection of healthy donor PBMCs, showed that SKA-31, a selective KCa3.1 activator, significantly reduced tumor burden compared to vehicle. This effect was not due to an effect of SKA-31 on cancer cells. HNSCC cell lines and patient tumor biopsies showed low to null KCa3.1 expression, and KCa3.1 activation did not increase their proliferation. However, quantitation of immunohistochemistry images of tumors from the HNSCC mice revealed a 2-fold increase in infiltrated NK cells in the SKA-31 treated group, thus supporting the important role of KCa3.1 in anti-tumor immunity.
KRAS inhibition triggers coordinated neoplastic and immune remodeling in the tumor microenvironment
Authors: Lu Daniel , Osgood Tao , Ma Shining , Estrada Juan , Arias Vanessa , Yamawaki Tracy , Hoang Uyen , Rex Karen , DeVoss Jason , Canon Jude , Lipford Rusty , Wang Songli , Coxon Angela , Li Chi-Ming 
Published: November 2024

Abstract
KRAS inhibitors are a novel, recently approved class of small molecules that target cancers with KRAS mutations, and understanding molecular mechanisms by which these inhibitors stimulate anti-tumor immunity will improve patient outcomes. In this study, we utilized a multi-omic approach combining single-cell RNA-seq, spatial transcriptomics, and flow cytometry to define cell signatures and signaling events associated with response in a murine model expressing KRAS-G12C tumors. Treatment with KRAS inhibitors but not MEK inhibitors drove neoplastic differentiation toward cell states with increased oxidative stress and extracellular matrix remodeling capacity. These changes were accompanied by spatially-defined Notch and Integrin signaling patterns between neoplastic, stromal, and classical dendritic cell (cDC) niches, leading to accelerated cDC maturation and CD8 effector T-cell activation. Our multi-omic design distills the complex cellular consequences and intercellular communication patterns in the tumor microenvironment following KRAS inhibition and provides a framework for contextualizing and validating disease-relevant signatures to guide therapeutic strategy.
Host genetic mechanisms regulate gammaherpesvirus mediated exacerbation of CNS autoimmunity.
Authors: Sessions Katherine , Holt Emily , Waytashek Courtney , Usherwood Edward , Krementsov Dimitry 
Published: November 2024

Abstract
Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS). Chronic infection with the gammaherpesvirus Epstein Barr virus (EBV) is the leading environmental risk factor, and the mechanisms behind this risk, including the role of host genetics, are still unclear. We have previously shown that host genetics play an essential role in the infection outcomes of a rodent pathogen homologous to EBV, murine gammaherpesvirus 68 (MHV68), where poor control of MHV-68 replication was associated with an expansion of cytotoxic CX3CR1+KLRG1+ T helper cells (ThCTL). To explore how genetically-determined infection outcomes modulate CNS autoimmunity, C57BL/6J (B6) and 129S1/SvImJ (129S) mice were first chronically infected with MHV-68 and then immunized for induction of experimental autoimmune encephalomyelitis (EAE), a rodent model of MS. Viral load was higher in 129S mice, and EAE clinical scores were more exacerbated by MHV-68 infection compared with B6. MHV-68 infection induced higher levels of CD8 T cells and IFNg production in CNS in both strains. In contrast, MHV-68 infection only promoted ThCTL infiltration into the CNS of B6 mice. Ongoing experiments will address the functional role of CD8 T cells and ThCTL. Because poor control of EBV and enhanced anti-EBV responses are associated with greater MS risk, our findings suggest that this could result from genetically determined control of viral load and augmented ThCTL and/or CD8 T cell responses in the CNS.
Mouse models advancing immunology research at The Jackson Laboratory
Authors: Beckwith Jason , Merriam Jennifer , Valenzuela Alicia , Pomerleau Darcy , Davis Crystal , Kelmenson Jennifer 
Published: November 2024

Abstract
The Jackson Laboratory (JAX) provides genetically-defined mice to the scientific community. This poster will highlight recently added novel models with immunology research application.
The portfolio of strains useful for transplantation/PDX and “humanized” mouse research continues to expand and complement the NSG (NOD scid IL2rgc-/-) model as ideal tools for engraftment of human or mouse tissues.
Novel fluorescent strains include Traffic Light Reporters for tracking DNA repair pathways, Gene Editing Reporters for detecting homology directed repair (HDR) and non-homologous end joining (NHEJ) repair events, as well as several CRISPR Editing, Bxb1 Integrase Editing, Base Editor and Prime Editor tools.
Recently added CRISPR Array Repair Lineage Tracing (CARLIN and DARLIN) component strains allow researchers to achieve high barcode diversity in targeted cell populations.
In addition to cryopreserving each strain, our genetic quality control program confirms mutation identity and genetic background, and screens for unwanted alleles (such as stray GFP, Cre, neo, etc.).
Search the comprehensive collection using the improved JAX Mouse Search site (mice.jax.org).
This resource includes models created by many generous donating institutions. Researchers are encouraged to donate their mouse lines via a very short Strain Submission form (jax.org/donate-a-mouse).
JAX is supported by the NIH, The Howard Hughes Medical Institute and other private charitable foundations.
Generation of type I interferon autoantibody tests for at-risk populations
Authors: Premsrirut Prem , Garcia Reino Eduardo , Gao Nicholas , Ayoluwa Nana , Sosa Elie , Brereton Racquel , Patera Andrew , Desikan Shreya , Davis Evan , Cheng Hui-Ting , Mendoza Rachelle 
Published: November 2024

Abstract
A plethora of evidence has shown that 10-20% of cases of severe COVID-19 occur in individuals harboring autoantibodies that neutralize type I interferons (IFNs) and thus block their ability to initiate innate defenses and dampen viral replication. These autoantibodies are, therefore, currently the strongest and most common determinants of a growing range of severe viral diseases, especially in the elderly; yet, no routine test is available to the general population to gain knowledge of their immune insufficiency and potential risk from future infections. Here, we developed a set of assays to establish the levels of anti-IFN autoantibodies and their neutralizing activity and performed validation studies in a cohort of hospitalized COVID patients (N=388) to assess their sensitivity and specificity. Consistent with previous findings, our tests developed in a CLIA-certified laboratory were able to detect anti-IFN autoantibodies in a sizable fraction of patients with severe and critical COVID-19 pneumonia. Taken together, our results suggest that anti-IFN autoantibody testing should be considered for standard clinical screening.
VIP deficiency disrupts immune ovarian homeostasis
Authors: Gori Soledad , Castagnola Lara , Gallino Lucila , Vota Daiana , Grasso Esteban , Parborell Fernanda , Pérez Leirós Claudia , Hauk Vanesa , Ramhorst Rosanna 
Published: November 2024

Abstract
Ovarian aging is associated with inflammation, contributing to the decline in oocyte count and quality. Since the vasoactive intestinal peptide (VIP) is an immunopeptide found in the reproductive tract with anti inflammatory effects inducing a tolerogenic response, we focus on VIP contribution to ovarian immune homeostasis preventing premature aging. We observed that young VIP KO mice (3 months old) display an altered ovarian histology with an increase in atretic follicles and less mature follicles as in aged WT mice (8 months old). This microenvironment was accompanied by an increased fibrosis deposition, ROS and IL-1β production. Interestingly, we found a significant decrease in ovarian reserve and the presence of foamy macrophages in young VIP KO ovary associated with premature aging. Getting insight into the mechanism involved in foam cell generation, we pre-conditioned young WT or VIP KO macrophages with WT or VIP KO ovarian media for 48h to mimic the ovarian microenvironment. Macrophages from VIP KO mice display a differential phagocytic ability of apoptotic bodies after preconditioning with WT or VIP KO ovarian media. However, only in VIP KO macrophages conditioned with VIP KO ovarian media we found an increase in lipid droplet accumulation, consistent with the formation of foamy macrophages. These findings highlight the role of VIP in regulating ovarian immune microenvironment, thereby preserving ovarian health and preventing premature aging.
V-set immunoglobulin domain-containing protein 4 (VSIG4) as a potential non-invasive biomarker of lupus nephritis
Authors: Teymur Aygun , Zhang Shu , Saxena Ramesh , Cai Qi , Wu Tianfu 
Published: November 2024

Abstract
Objective: This study aimed to identify a non-invasive biomarker of lupus nephritis (LN) for diagnosis, disease monitoring or flare prediction of LN.
Methods: Both the VSIG4 free form and immune complex (ICx) bound form in the urine of LN patients (N = 97) were examined using ELISA, to compare with chronic kidney diseases (CKD) controls and healthy controls. Immunohistochemistry was used to confirm the elevated tissue expression of VSIG4 in the renal biopsy of LN. Immunofluorescence was used to examine the VSIG4-mediated immune complex deposition in the kidney of LN.
Results: The urinary levels of both VSIG4 free form (P = 0.01) and VSIG4-ICx (P = 6.3 x 10-14) and were significantly elevated in LN patients, particularly in renal active LN (P = 4.8 x 10-16) compared to healthy controls. VSIG4-ICx effectively discriminated HC from LN (AUC = 0.84, P&lt; 0.01) and active LN from inactive LN (AUC = 0.91, P&lt; 0.01). The urinary immune complex form of VSIG4 was positively correlated with renal pathology activity index (AI). Immunohistochemistry highlighted the significantly elevated VSIG4 expression in both glomeruli and tubules in LN patients, compared to healthy controls.
Conclusion: Our findings underscore the diagnostic potential of VSIG4 as a non-invasive urinary biomarker for LN. With its correlation to disease activity and renal pathology, VSIG4 presents a promising avenue for enhanced diagnosis and monitoring of lupus nephritis, paving the way for improved patient care.
Amyotrophic lateral sclerosis is associated with changes in peripheral neutrophil heterogeneity and function
Authors: Baird Lillia , Teener Samuel , Webber-Davis Ian , Feldman Eva , Goutman Stephen , Murdock Benjamin 
Published: November 2024

Abstract
There is overwhelming evidence that the immune system plays a significant role in amyotrophic lateral sclerosis (ALS), a devastating neurodegenerative disease characterized by motor neuron degeneration. Expected survival is 2-4 years from disease onset and treatment options are limited. There is significant clinical and pathogenic heterogeneity and an incomplete understanding of the underlying pathogenesis, including mechanisms of immune contribution. Neutrophils are consistently implicated in ALS, with increased count in the blood and infiltration into the spinal cord. Increased neutrophil count and activation are associated with shortened survival and neutrophil levels are the single best predictor of future ALS progression. Despite this, little is known about which neutrophil subpopulations are dysregulated or the underlying mechanisms contributing to disease progression. Primary neutrophils were assessed via flow and ex vivo stimulation. NET remnants (MPO:DNA, elastase, extracellular DNA) were quantitated in plasma. Distinct populations were observed in controls and ALS subjects, and ex vivo analysis of primary neutrophils and analysis of NET remnants demonstrates changes in effector functions that may contribute to BBB disruption and neuronal damage in ALS. These differences in neutrophil populations and function provide compelling evidence that neutrophils play a role in ALS progression, as well as provide novel targets for therapeutics.
Comparison of the immunological response in mice after bronchoscopic and surgical methods for delivery of Pseudomonas aeruginosa to murine airways
Authors: Schelde Karen , Sonneborn Claire , Buczkowski Natalia , Bobinsky Annemarie , Host Derek , Jafri Anjum , Barbato Elisha , Brumbaugh Sarah , Rietsch Arne , Hodges Craig , Bonfield Tracey , Drumm Mitchell 
Published: November 2024

Abstract
We have established improved methods to access the mouse airways and lungs for pulmonary research. We have applied a non-surgical, bronchoscopic system with a camera to bacterial infection studies and found it reduced mortality and weight change in infected mice compared to our previously used procedure, a tracheostomy involving surgery. The two procedures were compared with and without bacteria to identify the inflammatory responses to the surgery and to the bacteria involved. We found the mouse tolerated the bronchoscope much better than the tracheostomy which allowed delineation of the effects of the bacteria from those of the surgical wound. The bronchoscope appeared to reduce stress to the mice, compared to the tracheostomy, and provided a significant benefit of reducing morbidity and mortality, thus reducing the number of mice needed for experiments. We found that mice receiving sterile beads instilled in the lung by tracheostomy lost more weight compared to mice undergoing bronchoscopy. Adding bacteria gave rise to even greater weight loss displaying significantly higher in surgery compared to bronchoscopy groups. Furthermore for surgery, immune cells (especially neutrophils) were increased in broncho-alveolar lavage fluid, as were pro-inflammatory cytokines TNF-α, IL-1β, and IL-6. The effects from the surgical wound are significant and should be recognized and incorporated into experimental designs to best reflect the condition the mouse is intended to model.
γδ T cells are crucial effectors of antibacterial response in T2D complicated with diabetic foot
Authors: Lehman Natalia , Suchodolski Wojciech , Jakubik Karol , Huang Tsan-Shin , Zarobkiewicz Michał , Bojarska-Junak Agnieszka , Roliński Jacek 
Published: November 2024

Abstract
Nearly half a billion people worldwide have type 2 diabetes (T2D), which poorly controlled may lead to severe consequences, including diabetic foot. Growing evidence highlights the role of γδ T subsets in the antibacterial response. A total of 70 patients were divided into three groups: T2D, T2D with diabetic foot (DF), and individuals without T2D. Flow cytometry was used to assess the frequency of peripheral blood γδ T along with activation (CD69, CD56) and senescence (CD57) markers. Intracellular expression of cytokines was tested on peripheral γδ T and Vδ2 in vitro cultures. PBMCs were stimulated with S. aureus and P. aeruginosa lysates and the expression of CD25, HLA-DR, along with CD137, CD154 on Vδ1 and Vδ2 cells was determined. The predominance of the Vδ2 was observed in DF group. γδ T cells in DF patients were significantly activated, as evidenced by CD69 and CD56 expression. Moreover, upon phosphoantigen stimulation, both peripheral and zoledronate-derived Vδ2 exhibited increased levels of intracellular IL-17A and TNF compared to patients without T2D. The γδ T cell responses to bacterial lysates were more pronounced in Vδ2 than in Vδ1 cells within the DF group. Among Vδ1 cells more of them were CD137 positive than Vδ2. Overall, the data show differentiated antibacterial properties of Vδ1 and Vδ2 cells in T2D patients. Understanding of the role of γδ T cell subpopulations may improve the prevention of T2D complications.
IM156, a new AMPK activator, ameliorates polymicrobial sepsis by regulating inflammatory responses
Authors: Kang Ji Hyeon , Lee Sung Kyun 
Published: November 2024

Abstract
IM156, a biguanide with a higher potency of AMP-activated protein kinase (AMPK) activation than metformin, exhibits inhibitory activity against angiogenesis and cancer. In this study, we explored the anti-inflammatory effects of IM156 against polymicrobial sepsis. Administration of IM156 significantly improved the survival rate and tissue damage in cecal ligation and puncture (CLP)-induced sepsis. Mechanistically, IM156 markedly reduced the number of bacterial cells in the peritoneal fluid and peripheral blood in a CLP-induced sepsis model. IM156 also inhibited splenocyte apoptosis and the production of inflammatory cytokines, including IL-1β, IL-6, and IL-10, in CLP mice. Furthermore, IM156 strongly inhibited the reactive oxygen species (ROS) generation by enhancing antioxidative effects, which can lead to reduced neutrophil extracellular trap (NET) formation in response to lipopolysaccharide. In conclusion, these findings collectively indicate that IM156 can suppress the inflammatory response and provide protection against polymicrobial sepsis. These results suggest the potential of IM156 as a novel therapeutic agent for sepsis control.
Macrophage GPR18 signaling activation by Resolvin D2 mediates metabolic switch and cardiac repair after myocardial infarction.
Authors: Thakkar Abhishek , Thorp Edward , DeBerge Matthew , Fredman Gabrielle 
Published: November 2024

Abstract
Higher inflammatory response is a major cause of heart failure and mortality post myocardial infarction (MI), a major health problem worldwide. Resolution of inflammation, well established as an actively orchestrated programmed response, is mediated by specialized pro-resolving lipid mediators (SPM) and cellular processes like efferocytosis. Resolvin D2 (RvD2), one such potent SPM, is expressed in leukocytes and stimulates the resolution of inflammation through signaling by its receptor GPR18. RvD2 signaling attenuates inflammation in immune-related conditions through macrophage (Mφ) cell-intrinsic and extrinsic mechanisms. The therapeutic potential of SPM has been demonstrated in treatment of sepsis and arthritis. However, the role of Mφ RvD2-GPR18 signaling in heart post-MI has not been elucidated. Here, we demonstrate (i) induction of GPR18 receptor on cardiac Mφ, (ii) &gt;50% reduction in survival of myeloid- and Mφ-specific GPR18-/- mice, (iii) reduced heart function in GPR18-/- mice post-MI, (iv) a GPR18-dependent suppression of TLR4 signaling cytokines, and (v) suppression of TLR4-induced glycolytic metabolism by RvD2-GPR18 signaling in vitro in Mφ from human GPR18 chimeric adult mice. To summarize, our data suggest GPR18-dependent metabolic and phenotype switch to anti-inflammatory Mφ and cardiac repair post-MI. Further interrogation will delineate the intracellular pathways downstream of GPR18 and demonstrate the potential of RvD2 in reducing mortality post-MI.