bluuebunny/crossref_metadata_2025_split huggingface dataset:
Myeloid-specific

ablation of Cblb is necessary to bolster defense against pulmonary fungal infection
Authors: Sharma Jaishree , Nanjappa Somashekarappa Gowda 
Published: August 2023

Abstract
Fungal pneumonia is a severe clinical problem, especially in patients with compromised immunity. The long-term therapeutic and prophylactic use of antifungal drugs in high-risk patients has promoted the emergence of multidrug-resistant fungi. Myeloid cells, mainly alveolar macrophages, are critical for orchestrating the immune responses, limiting pathogen growth, and resolving pathology during lung fungal infection. Thus, it is essential to understand mechanisms and uncover the host-fungus interface regulating early immune responses in the lung. Here, we studied the myeloid cell-specific role of the host element, CBLB, an E3 ubiquitin ligase, for antifungal immunity in the lungs. Using a mouse model of pulmonary blastomycosis, we found that targeted ablation of Cblb in myeloid cells (LysMcre/creCblbfl/fl), but not in all the cells, was essential to mediate protective immunity. Myeloid-specific Cblb deletion enhanced survival, reduced fungal burden, and reduced inflammation following lethal lung fungal infection. Mechanistically, myeloid-specific ablation of Cblb led to enhanced numbers of alveolar macrophages and increased activation and ROS production by neutrophils. Interestingly, we found an augmented type 17 innate lymphocytes in LysMcre/creCblbfl/fl mice than in controls, suggesting their potential crosstalk with Cblb-deficient myeloid cells. Our study suggests that myeloid-specific ablation of Cblb is necessary to enhance immunity against pulmonary fungal infections and proposes this as a potential therapeutic avenue.
Supported by grants from American Lung Association (IA-697123, to SGN), NIH (R01 AI53522, to SGN), and UIUC (Faculty Start-up Funds, to SGN)
Investigating the lipid-immune signaling axis in modulating the inflammatory response to Salmonella enterica serovar Typhimurium infections
Authors: Taddeo Jessica R , Beld Joris , Tukel Cagla , Tam Vincent 
Published: August 2023

Abstract
Salmonella enterica serovar Typhimurium (STM) is an intracellular pathogen that causes gastroenteritis and can disseminate systemically through survival in phagocytes. Eicosanoids-derived bioactive lipids play critical roles in both the induction and resolution of inflammation during bacterial infections or other insults. A subset of these lipids are endogenous ligands for nuclear receptors/transcription factors Peroxisome proliferator-activated receptors (PPAR), which play major roles in fatty acid metabolism and modulation of immune functions (including macrophage MΦ polarization). To elucidate the role of PPARα, we infected streptomycin-pretreated C57BL/6 (wildtype) and Ppara−/− mice with STM (colitis model). We determined that Ppara−/− mice have increased eicosanoid production, decreased inflammatory gene expression and histopathology scores in cecal tissue, and reduced systemic dissemination of bacteria, compared to C57BL/6 mice. These data suggest that the activation of PPARα results in a deleterious immune response favoring STM infection. When polarizing MΦ to various subsets, STM infection enhanced M2b and M2c polarizations (anti-inflammatory and immunoregulatory) in a PPARα-dependent manner. Using liquid chromatography-mass spectrometry (LC/MS), we also determined that STM infection alters the fatty acid profiles in polarized MΦ. Specifically, C57BL/6 M2b-polarized STM-infected MΦ have decreased ceramide levels compared to uninfected control, which would inhibit programmed cell death and enhance bacterial intracellular survival. These findings provide important insights into how the reprogramming of immune metabolism affects host-pathogen interactions during bacterial infections.
This work was supported by National Institute of Allergy and Infectious Disease R01 AI168550 (VT)
Candida albicans pathogenesis in the context of mucosal type II interferonopathy
Authors: Lubkin Ashira , Oikonomou Vasileios , Millet Nicolas , Swidergall Marc , Lionakis Michail S 
Published: August 2023

Abstract
Interferon-gamma (IFNγ) is a powerful cytokine that is crucial for adequate host defense. However, unchecked IFNγ can cause autoimmunity in several organs and can be directly toxic to host cells. Indeed, excessive IFNγ, or type II interferonopathy, occurs in several diverse disease states, including STAT1 gain-of-function mutations, autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), Downs syndrome, and AIDS. We have recently shown that in APECED, which is caused by deficiency of autoimmune regulator (AIRE), excessive IFNγ leads to greater susceptibility to oral Candida albicans infection. C. albicans is both a normal member of the human microbiome and the most common human fungal pathogen. The ability of C. albicans to morph between commensal and pathogenic states is tied to its complex transcriptional regulation of virulence traits in response to environmental cues. Thus, we hypothesize that excessive IFNγ in the oral mucosa leads C. albicans to transition to a more virulent state. We are using transcriptomics to define the response of C. albicans to excess IFNγ. Further, we have found that during infection of oral epithelial cells, C. albicans becomes more invasive in the presence of IFNγ. Thus, we are also using metabolomics to investigate soluble mediators released by these cells in response to IFNγ. These experiments will shed light on C. albicans virulence regulation in response to autoinflammatory conditions. Further, this study will provide new insights into the dualistic nature of IFNγ — providing protection against many pathogens, while facilitating disease from a specific pathogen.
This work was supported by the Division of Intramural Research of the NIAID (ZIA AI001175).
An essential role for CD4+ T cells during Staphylococcus aureus craniotomy infection
Authors: Kak Gunjan , Horn Christopher M , Heim Cortney E , Kielian Tammy L 
Published: August 2023

Abstract
Treatment of brain tumors, epilepsy, or hemodynamic abnormalities requires a craniotomy to access the brain. Despite prophylaxis, infectious complications after craniotomy range from 1–3% with approximately half caused by Staphylococcus aureus (S. aureus). Using our novel mouse model of craniotomy infection, a critical role of CD4+ T cells was established since bacterial burdens were significantly increased in the brain, galea, and bone flap of RAG1−/− mice as well as following CD4+ T cell depletion. Bacterial burdens were also increased following anti-VLA-4 & LFA-1 treatment, further establishing a beneficial role for peripheral CD4+ infiltrates. Adoptive transfer of in vitro skewed Th1 or Th17 cells into RAG1−/− mice returned infectious burdens back to wild type (WT) levels. Ex vivo staining of transferred CD4+ T cells recovered from the brains of RAG1−/− mice revealed a dominant IFN-γ signature, even following Th17 adoptive transfer, suggesting that Th1 responses are critical for infection containment. Further evidence of Th1 and Th17 involvement was demonstrated by exacerbated infection in Tbet−/− and RORγt−/− mice, respectively. Interestingly, phenotypes were not evident in either IL-17A/F−/− KO or IFN-γ−/− mice, whereas adoptive transfer of CD4+ T cells from either IL-17−/− or IFN-γ−/− animals mitigated bacterial outgrowth in RAG1−/− mice, suggesting the combined action of Th1- and Th17-derived mediators. scRNA-seq identified an increased IFN-γ-regulated signature (gbp1/5, isg15, irf1, stat1) in select microglial and infiltrating granulocytic clusters. This suggests that T cell-innate immune cell crosstalk is necessary for bacterial containment during craniotomy infection.
R01 AI169788
Programming of checkpoint blockade responsive exhausted T cells by rheostatic IL-2 signals during priming by dendritic cells
Authors: Sarkar Surojit , Toumi Ryma , Xiao Hanxi , Pulliam Thomas , Reed Jim , Nghiem Paul , Kalia Vandana 
Published: August 2023

Abstract
Stem-like progenitor exhausted CD8 T cells are critical for maintaining long-term resistance during chronic infections and cancer, and represent an important checkpoint blockade immunotherapy target for functional reinvigoration and disease control. Here, we show a fate-deterministic role of rheostatic IL-2 signals and differential DC priming in programming the development of stem-like progenitor exhausted CD8 T cells during chronic viral infection. In vivo fate-tracking studies reveal that strong IL-2 signals during priming drive terminal differentiation. In contrast, tempered IL-2 signals are associated with TCF-1Hi stem-like precursors, which give rise to progenitor exhausted CD8 T cells, capable of long-term persistence and potent responsiveness to anti-PD-1 therapy in later stages of chronic viral infection. In the context of human tumors as well, single cell RNA-seq analyses of total or antigen-specific tumor infiltrating lymphocytes show an inverse relationship of IL-2 signaling signature with T cell stemness, as well as checkpoint blockade therapy outcomes in melanoma. Our studies further show that the rheostatic control of exhausted T cell fates by differential IL-2 signals is physiologically mediated through differential cell surface expression of IL-2Rα during early stages of T cell activation, which in turn is pioneered during priming by distinct dendritic cell subsets. Notably, moderation of in vivo IL-2 signals during priming promotes the development of stem-like TCF-1Hi lineage, thus supporting a unique strategy for improving clinical immunotherapy outcomes by enhancing the development of long-lived, therapy-responsive stem-like cells with vigorous effector expansion capabilities.
Pediatric Cancer Research Foundation to SS, the Rachel Lynn Henley Foundation to VK, the Hopes and Smiles For Children Foundation to VK, In Concert for Cancer Foundation to VK and the National Institutes of Health (CA254168 to TP; CA225517 to PN; AI132819, AI103748 to SS; 5P30CA015704; AI154363 to VK)
Pak2-mediated phosphorylation promotes RORγt ubiquitination and inhibits colonic inflammation
Authors: Kathania Mahesh , Kumar Ritesh , Lenou Elviche Taskem , Basrur Venkatesha , Theiss Arianne L , Chernoff Jonathan , K Venuprasad 
Published: August 2023

Abstract
Dysregulated Interleukin (IL)-17 expression and its downstream signaling is strongly linked to inflammatory bowel diseases (IBD). However, the molecular mechanisms by which the function of RORγt, the transcription factor of IL-17, is regulated remains elusive, which is absolutely necessary to develop advanced strategies to inhibit IL-17-mediated inflammation. By mass spectrometry-based approach, we have identified that Pak2, a Ser/Thr kinase, directly associates with RORγt. Pak2 recognizes a conserved KRLS motif within RORγt and phosphorylated the Serine (S) 316 within this motif. Genetic deletion of Pak2 in Th17 cells resulted in reduced RORγt phosphorylation, increased IL-17 expression and severe colitis upon adoptive transfer to Rag1−/− mice. Similarly, reconstitution of RORγt-S316A mutant in Rorc−/− Th17 cells resulted in enhanced IL-17 expression and colitis severity. Mechanistically, we demonstrate that Pak2-mediated phosphorylation causes a conformational change resulting in exposure of the ubiquitin ligase Itch interacting PPLY motif and degradation of RORγt. Thus, we have uncovered a novel mechanism by which the activity of RORγt is regulated that can be exploited therapeutically.
This work was supported by funds from the National Institutes of Health (R01-DK115668 and R01-AI155786) and Cancer Prevention Research Institute of Texas (RP160577 and RP190527), a translational pilot project grant from the Harold C. Simmons Comprehensive Cancer Center, UT Southwestern Medical Center (23001045) to K.V., and R01-DK117001 to K.V. and A.L.T.
Unraveling myeloid cell-mediated mechanisms of resistance to immune checkpoint blockade in bladder cancer
Authors: Tran Michelle A , Farkas Adam M , Lee Karen , Horowitz Amir , Beaumont Kristin , Sebra Robert , Sfakianos John , Galsky Matthew , Bhardwaj Nina 
Published: August 2023

Abstract

Background
Only 15–25% of bladder cancer (BC) patients respond to PD-(L)1 immune checkpoint blockade (ICB) therapy. We previously used pre-treatment tumor to derive an ICB response gene signature that was enriched in adaptive immune genes and an ICB resistance signature enriched in innate immune and inflammatory genes.

Methods
We performed single-cell RNA sequencing (scRNAseq) on 27 bladder tumors and 3 normal-adjacent tissue specimens to resolve these signatures. We profiled tumor macrophages (MΦs) underlying the resistance signature with flow cytometry (FC). To recapitulate them, we used healthy donor blood monocytes differentiated with M-CSF, skewed with predicted ligands, and characterized by FC, RTqPCR, and bulk RNAseq. To examine for peripheral biomarkers, we conducted scRNAseq on paired blood and urine and performed O-Link and ELISA on plasma.

Results
Our ICB response and resistance signatures were enriched in distinct MΦ subsets: immunostimulatory (is)MΦs and pro-tumorigenic (pt)MΦs, respectively. ptMΦs upregulated SPP1, TREM1, and CLEC5A and pro-inflammatory and hypoxic programs whereas isMΦs upregulated antigen presentation and complement machinery. ptMΦs were enriched in tumor versus normal tissue. When we tested ptMΦ predicted drivers, IL-1β induced expression of Clec5a and Trem1 protein and transcription of SPP1, TREM1, and CLEC5A. In the first-ever BC urine scRNAseq, we discovered urine contains ptMΦs. Corresponding inflammatory cytokines (IL-1β, IL-6, IL-8) were elevated in advanced patient plasma compared to early stage and healthy donors. In conclusion, we identified pro-inflammatory SPP1+CLEC5A+TREM1+ MΦs that may underlie ICB resistance, be targeted via IL-1β and monitored via plasma and urine.

Supported by grants from NIH (F30 CA275269-01, R01 CA249175-01).
ON104, a novel antibody targeting oxidized Macrophage Migration Inhibitory Factor (oxMIF) shows efficacy in preclinical models of chronic inflammation
Authors: Ferhat Maroua , Landlinger Christine , Kerschbaumer Randolf , Tam Frederick W K , Nicoletti Ferdinando , Thiele Michael 
Published: August 2023

Abstract
Macrophage Migration Inhibitory Factor (MIF) is a pleiotropic inflammatory cytokine, ubiquitously present in various tissues and the circulation of healthy subjects. MIF is a primary counter-actor of glucocorticoids that emerged as a pivotal regulator of chronic inflammation. During inflammation, MIF converts into oxidized (ox)MIF which was described as the pathogenic and druggable isoform of MIF. ON104, a fully human antibody, was engineered to specifically neutralize oxMIF without triggering immune cell effector functions. ON104 was tested in three experimental models of chronic inflammatory diseases (CIDs): Glomerulonephritis (GN) in WKY rats, Collagen-Induced Arthritis (CIA) in DBA/1j mice; and adoptive T-cell transfer colitis in SCID mice. At disease onset, ON104 was administered at different dose levels twice a week for 1, 3, and 8 weeks in GN, CIA, and colitis, respectively. Clinical evaluations, blood and tissue sampling were done for all individual animals.
In the GN model, ON104 significantly reduced proteinuria, hematuria, and CD68+ macrophage infiltration within the inflamed kidneys. During CIA, ON104 ameliorated the severity of arthritis as indicated by reduced paw thickness, and decreased disease score. Furthermore, treatment with ON104 substantially attenuated clinical signs of colitis by preventing body weight loss and restoring stool consistency compared to the vehicle-treated controls.
Our findings substantiate the role of oxMIF in the pathogenesis of various CIDs. Thus, ON104 has the potential to become a well-tolerated therapeutic antibody for treating patients with CIDs.
Systemic administration of anti-CD20 indirectly reduces B cells in the inflamed meninges in a chronic model of central nervous system autoimmunity
Authors: Kerfoot Steven , Tesfagiorgis Yodit , Kemal Eden A , Craig Heather C , Parham Kate A 
Published: August 2023

Abstract
Anti-CD20 B cell depleting therapies have demonstrated that B cells are important drivers of disease progress in Multiple Sclerosis, although the pathogenic mechanisms are not well understood. A population of B cells accumulates in the inflamed meninges in MS and also in some chronic animal models of disease, typically adjacent to demyelinating lesions. The role of these meningeal B cells in disease is not completely understood, nor is their susceptibility to anti-CD20 therapy. Here, we administered anti-CD20 to 2D2 IgHMOG spontaneous experimental autoimmune encephalomyelitis mice in the chronic phase of disease, after the establishment of meningeal B cell clusters. Compared to the circulation, lymph nodes, and spleen, B cell depletion from the CNS was delayed and not evident until 7d post administration of anti-CD20. Further, we did not find evidence that anti-CD20 accessed meningeal B cells directly, but rather that depletion was indirect and the result of ongoing turnover of the meningeal population and elimination of the peripheral pool from which it is sustained. The reduction of B cell numbers in the CNS coincided with less demyelination of the spinal cord white matter and, surprisingly, an increase in the number of T cells in the meninges but not parenchyma.
YT is the recipient of an endMS Doctoral Studentships from the Multiple Sclerosis Society of Canada (MSSOC). KAP is the recipient of an endMS Post-Doctoral Fellowship from MSSOC. This study was funded by operating grants from the Canadian Institutes of Health Research and the MSSOC.
Stable Expression of FOXP3 and Core Tolerogenic Genes by Engineered Human Tregs
Authors: Zarin Payam , Sassone-Corsi Martina , Doherty Abigail , Chicoine Adam , DeVault Victoria , Li Xiang , Yam Jennifer , Wickham Thomas , Chen Tiffany , Depis Fabien 
Published: August 2023

Abstract
FOXP3+ Regulatory T cells (Tregs) play a critical role in the prevention of fatal autoimmunity and maintaining tissue homeostasis. The functional stability of Tregs remains critical in their contribution towards immune tolerance rather than runaway immunity, particularly for cell therapy whereby an inflammatory microenvironment may impact the expression of FOXP3 and associated tolerogenic genes. To address potential Treg instability, we generated human Engineered Tregs (EngTregs) through a gene editing approach of PBMC isolated CD4+ T cells, leading to stable FOXP3 expression and a rapamycin-activated signaling complex that provides tunable IL-2 signal, thereby effectively divorcing FOXP3 expression from existing regulatory elements known to promote Treg instability under inflammatory conditions. Using transcriptomic analysis, we evaluated the stability of cultured sorted Tregs (cTregs) compared to EngTregs. We found EngTregs expressed higher levels of key “Core Treg” and “FOXP3 Synergy” genes based on bulk and single cell RNA-seq analysis as confirmed by flow cytometry. Additionally, favorable expression pattern of key Treg stability markers, CD27, CD70 and IKZF4 (EOS) was observed in EngTregs compared to cTregs. Conversely, cTregs express higher levels of cytotoxic genes including GZMA and PERFORIN1 compared to EngTregs along with other inflammatory genes. Finally, we observed that these correlations have a functional implication as demonstrated by higher expression of key tolerogenic markers in EngTregs compared to cTregs. This work strongly supports EngTregs as stable, tolerogenic FOXP3+ T cells, thereby providing an invaluable asset for treating life-threatening autoimmune and autoinflammatory diseases.
Harnessing the Full Potential of Regulatory T Cells by mRNA-induced Antigen Specificity
Authors: Scott David W , Janssens Ibo , Van Delen Mats , Flumens Donovan , Dams Amber , Roex Gils , Van Den Bos Jasper , Campillo-Davò Diana , Packialakshmi Balamurugan , Cools Natalie 
Published: August 2023

Abstract
Regulatory T cells (Tregs) are crucial in inducing and maintaining tolerance, and already have shown clinical potential. In this study, we describe an efficient non-viral engineering method by TCR-encoding mRNA electroporation, while also exploring the potential of further promoting on-target effects by disrupting the endogenous TCR using a fully RNA-based CRISPR/Cas9 technology.
Endogenous TCR disruption prior to TCR-transfection resulted in more than 90% blockade of the endogenous TCR expression and a similar high (&gt;85%) but more homogenous expression of the transgenic TCR. Importantly, following antigen-specific TCR-induced stimulation, both engineered Tregs significantly increased the expression of multiple activation and suppression markers. Moreover, the engineered Tregs outcompeted polyclonal Tregs in suppressing antigen-specific effector T cell proliferation and performed better in blocking moDC maturation, even in the presence of antigen-activated effector T cells. Secretome screening revealed a distinct anti-inflammatory profile for TCR-Tregs. On the other hand, TCR-engineered KO Tregs co-expressed anti-inflammatory cytokines together with some pro-inflammatory cytokines, but qualitatively different from their effector Th counterparts. Hence, an anti-inflammatory profile with Th cell features is seen with TCR-engineered KO Tregs. In conclusion, we show that this versatile approach can be used to induce the functional expression of a transgenic TCR, but also for CRISPR/Cas9-mediated disruption of endogenous TCRs with high efficiency using an RNA-based platform. Comparison between polyclonal Tregs and TCR engineered Tregs in EAE using an HLA-transgenic mouse model is now being investigated.
National Multiple Sclerosis Society (RG-1612-26484)
Determining antitumor responses to variable cadherin expression
Authors: Jordon Alexander R D , Fugere Céline , Borys Samantha , Brossay Laurent 
Published: August 2023

Abstract
Cancer immunotherapy has dramatically transformed the treatment of many cancers. Despite this, several limitations exist. For example, in some patients’ cancer can become resistant to immunotherapy and relapse is common. Therefore, it is essential to identify new targets for intervention. Our laboratory previously showed that KLRG1, which has two main ligands E-cadherin and N-cadherin, is a potential target. However, cadherin has many roles including a role in epithelial mesenchymal transition. To examine the KLRG1-cadherin axis in antitumor responses we used CRISPR/Cas9 and retroviral overexpression methods to develop a library of B16–F10 melanoma and 4T1 breast cancer cell lines with variable cadherin expression. The appropriate expression of E- and N-cadherin on the cells in this library was confirmed by both flow cytometry and Western blot. We then demonstrated that the modulation of cadherins does not alter cell proliferation rates in vitro. Importantly, in both models, we find that E-cadherin expressing cell lines cause significantly more mortality than E-cadherin deficient parental cell lines. Together, these findings suggest that the KLRG1-cadherin axis play an important role in antitumor responses.
This work was supported by the NIH research Grant R01 AI46709 (L.B.)