bluuebunny/crossref_metadata_2025_split huggingface dataset:
Optimization of T Cell Activation Protocols to Enrich Stem Cell Memory Phenotypes: Metabolic and Health Assays as Predictive Tools
Authors: Haupt Kim , Sylvester Kayla , Lauer Anthony , Karassina Natasha , Vidugiriene Jolanta 
Published: November 2024

Abstract
The generation of T cells with a stem cell memory phenotype (T_SCM) holds significant promise for immunotherapy due to their longevity and potent antitumor activity. This study aimed to optimize T cell activation and expansion conditions to selectively enrich for the T_SCM population. Utilizing a suite of cell health and metabolism assays, we monitored T cell cultures to identify predictive markers of T_SCM enrichment. We observed that early activation steps correlated with a surge in ATP production and alterations in cell reducing potential, indicative of metabolic reprogramming. Subsequent activation phases were marked by an upregulation in both catabolic and anabolic pathways, as evidenced by the increased turnover of key metabolites including glucose, lactate, glutamine, glutamate, branched-chain amino acids (BCAA), and pyruvate. Notably, the rate of T cell activation influenced the composition of the resultant T cell populations, with the media composition further affecting these dynamics. Our findings suggest that metabolic assays, particularly those measuring changes in ATP levels and primary metabolite flux, can serve as effective early predictors of T_SCM enrichment. These insights may guide the development of more efficacious T cell-based immunotherapies by facilitating the targeted generation of T cells with a stem cell-like memory phenotype.
Excess airway iron during severe SARS-CoV-2 infection impairs neutrophil antifungal activity enabling 2° mold infection
Authors: Saitornuang Sawanan , Lam Cuong , Riegler Ashleigh , Meehan Will , Oh Joo-Yeun , Patel Rakesh , Haas Hubert , Saini Shefal , Lapi Suzanne , Leal Sixto 
Published: November 2024

Abstract
COVID-19-Associated Pulmonary Aspergillosis (CAPA) is an invasive 2° mold infection caused by Aspergillus species affecting ~5% of patients with severe SARS-CoV-2 (SARS-2) infection. In this study, we hypothesized that excess airway iron released during SARS-2 infection promotes 2° infection via enhancement of fungal growth and impairment of neutrophil antifungal activity. We show that SARS-2 bronchoaveolar lavage samples (n=48) have higher levels of total iron, ferric iron, and oxyhemoglobin relative to bacterial pneumonia (n=40). Likewise, FFPE lung tissues from SARS-2 infection exhibited increased extracellular ferric iron, ferrous iron, ferritin, and hemoglobin. We show that both iron and hemoglobin enhance the growth rate of Aspergillus fumigatus via siderophore and reductive iron assimilation dependent mechanism. Utilizing lung epithelial cell transmigration model, we show that iron enhances neutrophil intracellular ROS production resulting in enhanced conidiacidal activity. In contrast, transmigrated neutrophils exposed to both heme and hemoglobin exhibit decreased ROS production and NET formation with significant impairment in hyphal killing. When both neutrophils and fungi are co-cultured with iron, heme, or hemoglobin, fungi consistently win the battle characterized by uncontrolled hyphal proliferation. These data identify both microbial and host mechanisms which may be targeted pharmacologically to mitigate the iron-dependent growth of 2° mold infection.
IL-10 regulates BCL-3 mediated adaptation to repeated TLR4 stimulation in macrophages
Authors: Bongartz Hannes , Bradfield Clinton , Gross Julia , Nita-Lazar Aleksandra , Fraser Iain , Meier-Schellersheim Martin 
Published: November 2024

Abstract
Due to its potential to initiate pathological inflammation, Toll-like receptor 4 (TLR4) mediated activation of macrophages has been studied intensely, revealing a multitude of regulatory components and mechanisms. Systematically varying dose and duration of priming and re-exposure of bone marrow derived macrophages (BMDM) to the TLR4 ligand KLA, we quantified dose-dependent adaptation and sensitization at multiple stages of the signaling response, including TLR4 internalization, MAPK activation, and nuclear translocation of transcription factors. We find that the anti-inflammatory cytokine IL-10 neither interferes with MyD88-dependent MAPK, NFkB nor TRIF-dependent IRF signaling upon restimulation after priming. Instead, IL-10 is found to affect nuclear translocation of the transcriptional regulator BCL-3 while showing a surprisingly strong dependence on the KLA exposure history. Moreover, chromatin immunoprecipitation of BCL-3 reveals that BCL-3 occupies kb sites in the TNFa and IL-6 genes in hypo-responsive macrophages in an IL-10 dependent manner.
This work was supported by the intramural program of the NIAID, NIH.
Functional Phenotype and Ontogeny of Olfr2 Expressing Vascular Macrophages in Atherosclerotic Plaques
Authors: Orecchioni Marco , Armstrong Suthahar Sujit , Nettersheim Felix , Alimadadi Ahmad , Wang Erpei , Billitti Monica , Resto-Trujillo Natalya , Roy Payel , Hedrick Catherine , Ley Klaus 
Published: November 2024

Abstract
Olfactory receptor 2 (Olfr2) and its human ortholog OR6A2 have emerged as a critical modulator of inflammation, with its presence detected in approximately 30% of vascular macrophages and its depletion shown to reduce atherosclerosis in mice. Single-cell RNA sequencing identified several distinct subsets of macrophages in the atherosclerotic plaque. This study investigates the functional phenotype and ontogeny of Olfr2-expressing vascular macrophages. Olfr2+ macrophage number increased in response to a western diet (WD) in the aorta and they expressed a variety of pro-inflammatory cytokines, including Il1b, and Il6, as well as Ccl5, Cx3cl1, and Ccl22. By extracting differentially expressed genes from bulk RNA-seq data of Olfr2+ vs. Olfr2- vascular macrophages, we defined a signature that mapped with mouse plaque single-cell data of monocytes, aortic intima resident macrophages (MacAir) and Trem2-Gpnmb foamy macrophages. Similar results were found in human plaque, with monocytes and foamy macrophages being significantly enriched. Olfr2 expression also increased in oxLDL-mediated foamy macrophages in vitro. Adoptive transfer experiments confirmed that Olfr2+ macrophages are monocyte-derived, accumulate in the aortic plaque within 72 hours, and display enhanced proliferative potential. Collectively, our findings indicate that Olfr2 expression is boosted by diet in monocytes-derived foamy macrophage subsets that by sensing octanal promote inflammation and atherosclerosis.
γδ​​​​​​​ T17 cells are master regulators of the acute antiviral response in HSV-1 infected corneas
Authors: Rodenberg Rachel , Spadafora Domenico , Fitzpatrick Steffani , Daly Grant , Lausch Robert , Barrington Robert 
Published: November 2024

Abstract
Herpes stromal keratitis (HSK), caused by herpes simplex virus 1 (HSV-1), is the leading cause of infectious blindness in developed countries. Innate-acting γδ T17 cells are critical for protection against ocular HSV-1 infection. It is well established these cells can facilitate neutrophil influx into the cornea, but whether γδ T17 cells regulate other innate immune cells following ocular HSV-1 infection remains unclear. Natural killer (NK) cells also participate in the innate immune response generated against HSV-1 by directly killing infected cells through secretion of granzymes and promoting the antiviral response by production of IFN-γ. Our data strongly suggest that γδ T17 cell secretion of IL-17A promotes NK cell accumulation in the infected cornea. This is demonstrated in several ways: 1) in mice that lack γδ T cells (TCRδ-/-) there are fewer antiviral NK cells following corneal HSV-1 infection compared to wild-type (WT) mice; 2) administering IL-17A to TCRδ-/- mice restored the NK cell population; and 3) neutralization of IL-17A or 4) inhibiting γδ T17 cell influx to corneas in WT mice diminished NK cell accumulation leading to increased viral titers. In contrast, IL-17A production is enhanced in the absence of IFN-γ. In sum, this study identifies a novel mechanism by which IL-17A production by γδ T17 cells promotes antiviral NK cell responses that, in turn limit further IL-17A production via INF-γ secretion by NK cells in the HSV-1 infected cornea.
SKIN RESIDENT T CELL AND NPY1R+ NEURON INTERACTIONS DURING WOUND REPAIR
Authors: Lam Kendra , Colin Joseph , Almaguer Berenice , Jameson Julie 
Published: November 2024

Abstract
Wound repair involves complex cellular interactions to induce proper healing. The transmission of pain signals and immune cell function occurs in unison during tissue repair. The cells and factors responsible for this neuroimmune crosstalk are important to delineate. Knowing γδ T cells and NPY play key roles in wound repair, we aimed to ascertain if γδ T cells and NPY1R+ neurons exhibit crosstalk. We identified that NPY1R+ T cell and neuron subsets including γδ T cells reside in the skin, and that there are γδ T cell-neuronal interactions in the epidermis and dermis of both non-wounded and wounded skin. We then compared dermal interactions in obese and lean mice in wounded and non-wounded conditions, as a model for diabetic neuropathy. There are decreased NPY1R+ neuron-γδ T cell interactions in wounded and non-wounded mice fed a high-fat diet for 14 weeks, indicating obesity and type 2 diabetes negatively impact crosstalk during wound healing. Last, to determine the impact of the γδ TCR on NPY1R+ neuron interactions in wound healing, we compared epidermal T cell-neuron interactions in TCRd-/- mice during wound repair. We found a decrease in interactions between αβ T cells and NPY1R+ neurons in TCRd-/- mice, suggesting γδ T cells communicate with NPY1R+ more frequently during wound repair. Further work will help uncover the factors and receptors that γδ T cells and NPY1R+ neurons utilize to communicate during wound healing, and the consequences of disrupting this relationship.
Regulation of IL-1β-mediated inflammatory responses by neural networks
Authors: Hepler Tyler , Hashimoto Okito , Tynan Aisling , Tracey Kevin , Chavan Sangeeta 
Published: November 2024

Abstract
Sickness behavior associated with inflammation impairs quality of life and increases morbidity and mortality. However, the neural mechanism that mediates inflammation-associated changes are poorly understood. Here, we identify the bed nucleus of the stria terminalis (BNST) as a critical node in mediating responses to cytokines. Administration of recombinant interleukin-1β (rIL-1β) significantly increases serum IL-6 (control vs. rIL-1β; 22.6±4.7 vs 1,487±341.4 pg/ml; p=0.0079). A significant decrease in core body temperature (pre vs. post; 38.1±0.2 vs. 36.1±0.4oC; p&lt;0.0001) and activity level (pre vs. post; 0.2±0.03 vs. 0.03±0.01 A.U.; p=0.0002) is also observed with rIL-1β administration. We visualized rIL-1β-responsive neurons using activity-dependent cell labeling, and identified distinct neuronal populations in the BNST that are active in response to IL-1β. Selective activation of IL-1β-responsive neurons using a chemogenetic approach is sufficient to induce significant increase in serum IL-6 levels (control vs. IL-1β; 26.2±74.1 vs. 272.5±8.3 pg/ml; p=0.01). Reactivation of IL-1β-responsive neurons also induces significant decrease in core body temperature (pre vs. post; 37.6±0.3 vs. 36.1±0.6oC; p=0.02) and activity levels (pre vs. post; 0.1±0.01 vs. 0.06±0.01 A.U.; p=0.02). Our data defines a neural population in the BNST that is sufficient for mediating IL-1β-associated responses, revealing its role in the coordination of inflammatory responses.
Clinical implications for CAR T cell persistence and functional phenotype in patients treated with CD19 CAR T cell therapy.
Authors: Bochnacka Oliwia , Quinn Suzanne , Dalheim Annika , Scurti Gina , Hossain Nasheed , Nishimura Michael 
Published: November 2024

Abstract
Understanding the critical relationship between cancer and the immune system has led to novel immune therapies including chimeric antigen receptor (CAR) T-cell therapy. In the clinic, CAR T cells have been most effective against hematological malignancies. CD19 is expressed in almost all stages of B cell development as well as in most B cell malignancies, making it an ideal antigen to target using CAR T cells. Despite the promising efficacy of CD19 CAR T cell therapy, there remains a subset of patients who are resistant to therapy. At Loyola University Chicago, we initiated a CD19 CAR T cell clinical trial in 2020. A total of 6 patients were treated on protocol, with a 50% durable complete response rate. Importantly, no treated patients experienced severe side effects. Unique to the CD19 CAR T cells, is the incorporation of a CD34 marker gene (CD34t) in the CAR retroviral construct used for T cell transduction. The use of CD34t allows us to track infused CAR T cells in the blood. With the ability to distinguish CD19 CAR T cells from the endogenous T cell population, we have developed an immunofluorescence staining panel of 39 functional cell surface proteins to study the treated patient blood samples. We found that CD19 CAR T cells persist regardless of clinical outcome. We are studying the changing CAR T cell phenotype in patients with varying clinical outcomes to identify phenotypic trends that may have clinical implications and could serve as relevant clinical biomarkers.
In vitro and in vivo proof-of-concept studies of extracellular protein degradation using novel M6PR-targeting chimeras (MTACs).
Authors: Molz Lisa , Liu Hu , Zhu Nanqun , Karra Srinivasa , Tozzo Effie 
Published: November 2024

Abstract
The mannose-6-phosphate-receptor (M6PR) traffics proteins from extracellular and intracellular locales to late endosomes/lysosomes for degradation and is ubiquitously expressed. This natural endogenous function of the mannose-6-phosphate-receptor (M6PR) may be harnessed for degradation of soluble circulating proteins and cell specific degradation of membrane proteins.
Here we describe the design and development of novel M6PR targeting chimeras (MTACs). MTACs are heterobifunctional molecules containing Avilar’s novel, potent, small molecule M6PR-binding ligands conjugated to ligands that bind a target protein of interest for degradation. For proof-of-concept studies, MTACs were designed to target the extracellular protein IgG, the second most abundant circulating protein with a long plasma half-life. Reduction of pathogenic IgG autoantibodies is a clinically validated strategy for treating multiple debilitating autoimmune diseases. In vitro characterization of tridentate and monodentate MTACs revealed potent biochemical binding to M6PR and IgG, and MTAC-mediated cellular uptake and degradation of IgG via the endolysosomal pathway in K562 and LNCaP cells. In vivo MTAC-mediated depletion of human IgG was demonstrated using a heterologous rat PK/PD model.
Vgd1 and Vgd3 T cells are abundant gd T cells populations in high-grade serous ovarian carcinoma, non-small/small cell lung cancer, and show a heterogeneous g-chain usage
Authors: Chaurio Gonzalez Ricardo , Payne Kyle , Anadon Carmen , Lopez-Bailon Luis , Yu Xiaoqing , Mine Jessica , Bari Shahla , Plappert Noah , Perez Bradford , Innamarato Patrick , Melendez Joanna , Eysha Mostafa , Semidey-Hurtado Jonathan , Carter Bethany , Kroeger Jodi , Antonia Scott , Rodriguez Paulo 
Published: November 2024

Abstract
γδ T cells, in contrast to the conventional MHC-restricted αβ, recognize and kill tumor cells in an MHC-unrestricted manner, highlighting their potential for anti-tumor therapies. However, the phenotypes of γδ T cells infiltrating human tumors and their role in anti-tumor immunity remain poorly understood. To assess this, we characterized by Immuno-Seq/iRepertoire the repertoire of γδ-chains used by intra-tumoral γδ T cells in human cancers from FACS-sorted T cells in freshly dissociated high-grade serous ovarian cancers (HGSOCs), non-small cell lung cancer (NSCLC), and small cell lung cancer (SCLC). Our analysis confirmed that δ1 and δ3 TILs are abundant in all diseases. Interestingly, γδ TILs show heterogenous Vγ-chain repertoires, although TCRVγ9 was the dominant γ-chain. We then encapsulated and run 10X (single-cell) analyses, with sorted, bar-coded, δ1 plus δ2 TILs. All tumors showed distinctive clusters of stem-like cells expressing TCF7, IL7R and CD27 (δ1stem); plus effector T cells expressing high levels of granzymes, IFNG and perforin and NK receptors (δ1Eff). Importantly, we found low expression of PD-1, with only a small cluster of δ1 TILs with TOX expression in lung cancer. Therefore, our data indicate that: 1) γδ T cells quasi-universally infiltrate human carcinomas; 2) δ1 T cells outnumber δ2 T cells in tumors; 3) δ1 TILs use a variety of Vγ chains, with dominant Vγ9 and 4) γδ T cell tumor infiltration is associated with better outcome.
NKG2A expression on tumor reactive CD8 T cells is induced by IL-12
Authors: Fesneau Olivier , Rosales Wesley , Samson Kimberly , Rajamanickan Venkatesh , Haugh William , Huang Jianguo , Duhen Thomas 
Published: November 2024

Abstract
Targeting the NKG2A/HLA-E pathway is a promising strategy to restore CD8 T cells functions, but several aspects of the biology of NKG2A+ CD8 T cells remain to be understood to fully benefit from it. Here, we characterized the NKG2A+ tumor infiltrating CD8 T cells (CD8 TILs) from patients with head and neck squamous cell carcinoma (HNSCC) and colorectal cancer (CRC). We showed that NKG2A was expressed by 10 to 15% of CD8 TILs, most of them resembling the previously described CD39+CD103+ (DP) CD8 TILs. scRNA-seq analysis revealed that NKG2A was upregulated as cells differentiated into DP CD8 TILs and acquired cytotoxic properties. NKG2A- and NKG2A+ DP CD8 TILs shared a significant TCR repertoire overlap, suggesting local upregulation of NKG2A. Using RNAscope we demonstrated that NKG2A+ DP CD8 TILs were enriched at the invasive margin and in the tumor bed where they might interact with HLA-E-expressing tumor cells. In contrast to previous results, we showed that CD4 T cells and APCs were required for NKG2A induction and this was dependent on CD40/CD40L interaction and IL-12 secretion. Altogether, our study provides evidence that NKG2A is expressed by tumor reactive CD8 TILs. It also highlights the dual role of IL-12, promoting cytotoxic T cell responses while inducing the expression of immune regulatory markers. The latter might negatively impact the anti-tumor immune response and should be taken into consideration in the future.
Analysis of transcriptional changes associated with anti-tumor effects of iNKT immunotherapy
Authors: Ferguson Sarah , Gumperz Jenny , Brand Joshua , Dinh Huy 
Published: November 2024

Abstract
iNKT cells are innate T cells that can dramatically influence outcomes of immune responses, and that are associated with anti-tumor immunotherapeutic effects in murine models and human clinical trials. Using a human xenograft model of EBV-driven B cell lymphoma, we found that administering a combination of human CD4+ iNKT cells and monocyte-derived DCs markedly reduced tumor mass, while equivalent doses of iNKT cells or DCs alone had little anti-tumor effect. NanoString array analysis of tumor and spleen from mice given a combination of iNKT cells and DCs identified a number of gene expression changes in both tissues compared to untreated lymphoma-bearing mice, suggesting that the interaction of iNKT cells with DCs activates gene expression pathways that promote anti-tumor responses. Ongoing RNA-seq analyses are aimed at further investigating transcriptional changes resulting from iNKT-DC interactions. Gene expression signatures of DCs and iNKTS cultured alone or mixed and immediately lysed will be compared to those of iNKT cells and DCs co-cultured for 24 hours. Differential gene expression analyses will be performed to identify specific transcripts that are up or down-regulated after iNKT-DC co-incubation. Transcriptional changes resulting from iNKT-DC interactions will be compared to those induced in iNKT cells or DCs alone after exposure to other activating stimuli. Together, these analyses will identify candidate pathways that promote anti-tumor outcomes.
Relevance of resident memory T (TRM) cells in antitumor immunity and response to cancer immunotherapy
Authors: Mami-Chouaib Fathia , Damei Isabelle , Corgnac Stéphanie 
Published: November 2024

Abstract
CD8+CD103+ TRM cells accumulate in human lung tumors and are associated with a favorable prognosis and response to immune checkpoint blockade (ICB). However, a large fraction of human solid tumors is only weakly infiltrated by TRM, and the reason of this TRM desert is not explained. Defining the molecular signals that give rise and maintain TRM cells within the tumor microenvironment (TME) and the mechanisms that potentiate antitumor TRM functions are important challenges. Data obtained in pre-clinical mouse tumor models demonstrate that knocking-out CD103 impairs antitumor T-cell immunity and response to ICB and therapeutic peptide vaccine. Immunization of melanoma-bearing mice with the cancer vaccine results in a dramatic decrease in the percentage of CD103+ TRM cells among CD8+ tumor-infiltrating T lymphocytes (TIL). Algorithm analyses applied to spectral cytometry data of TIL show that upon activation with the peptide vaccine a TRM subset gives rise to tumor-specific effector T cells. The mechanisms associated with the decrease in CD103 expression on CD8+ TIL and lead to a TRM differentiation shift will be presented as well as the role of TGF-beta in CD8+ TRM formation and persistence. Our data demonstrate that tumor CD8+CD103+ TRM are capable of mounting a potent antitumor immunity, and suggest that a fine balance between diverse signals in the tumor ecosystem is required to induce their differentiation and thereby optimize response to immunotherapy.
Low-dose mIL-2/CD25 alters the composition and reduces the frequency of autoreactive T cell subsets in inflamed islets from NOD mice
Authors: Põder Janika , Malek Thomas 
Published: November 2024

Abstract
To enhance Treg-targeted IL-2-dependent therapy for autoimmunity, we developed a fusion protein of mouse (m) IL-2 linked to CD25. mIL-2/CD25 has a long half-life, acts as a selective agonist for the high-affinity IL-2R, at a low dose (LD) preferentially targets Tregs and is more effective than IL-2 at limiting autoimmunity in models of type 1 diabetes (T1D) and lupus. To explore the mechanism for T1D suppression, we assessed the effect of LD-mIL-2/CD25 on the global islet immune landscape in female NOD mice with scRNA-seq and flow cytometry. In the pancreas, LD mIL-2/CD25 reduced CD4+Foxp3- and CD8+ T cells, especially effector-phenotypic and highly proliferating CD8+ T cells, but increased Tregs, particularly ST2+ eTregs, and ILC2s. In addition, a CD8+ T cell subset expressing an exhaustion-related gene signature was identified. TCR repertoire analysis revealed multiple dominant clonotypes associated with effector-phenotypic CD4+Foxp3- and CD8+ T cells in untreated diseased NOD mice, which were reduced by LD mIL-2/CD25. Consistent with this finding, peptide-MHC tetramer staining showed that LD mIL-2/CD25 reduced autoantigen-reactive CD4+ and CD8+ T cells in islets. Our findings suggest that Tregs regulate autoimmunity in NOD mice in part by limiting the expansion of autoreactive T cells. Moreover, mIL-2/CD25 may also function through Tregs to promote exhaustion of CD8+ T cells, and this may reflect an active mechanism by which Tregs regulate some autoreactive T cells.