bluuebunny/crossref_metadata_2025_split huggingface dataset:
IL-21 blocks B cell death from BCR-delivered TLR9 agonists and upregulates both AID and T-bet (LYM6P.773)
Authors: Naradikian Martin , Oropallo Michael , Marshak-Rothstein Ann , Cancro Michael 
Published: January 2023

Abstract
A growing literature implicates the B cell receptor (BCR), Toll-like receptors (TLR), and survival cytokines or costimulation in the etiology of SLE and other humoral autoimmune diseases. Our lab has found that BCR-delivered TLR9 ligands yield a unique post-proliferative cell death that limits responses to DNA-linked antigens. Furthermore, death via this mechanism is rescued by the survival cytokine BLyS. However, here we show that BLyS is not unique in its capacity to rescue this cell death. Indeed, T cell-derived survival signals including CD40L, IL-4, and IL-21 similarly rescue B cells from BCR delivered TLR9 agonists. All of these stimuli also induce AID, which is required for class switch recombination and somatic hypermutation. However, IL-21 uniquely upregulates T-bet in the context of TLR9 stimulation. Further, we show that IL-21 induces antibody secretion to the IgG2b and IgG2c isotypes. As these isotypes have been implicated in humoral autoimmunity, our findings suggest that IL-21 may be involved in the production of DNA autoantibodies.
Examining the local response in the mucosa: how microfold cells act as first responders during a bordetella infection (MPF3P.811)
Authors: Goodfield Laura , Muse Sarah , Harvill Eric 
Published: January 2023

Abstract
During a respiratory tract infection, the first interaction between the pathogen and host occurs at the mucosal interface. Microfold (M) cells, specialized antigen sampling cells associated with the mucosa and recently identified in the respiratory tract (Kim, D.Y, et al., 2011), are considered the gateway of the initial immune response because they are constantly binding and taking up soluble and inert antigens as well as commensals and pathogens. To determine how M cells contribute to the generation of an appropriate immune response against an invading pathogen, we investigated the response of villous M cells to the respiratory pathogen B. bronchiseptica using a co-culture system and in the murine nasal cavity. Since M cells must quickly respond to initiate a mucosal immune response following the binding of an antigen, we measured changes in mRNA encoding the cytokines and chemokines interleukin-4, interleukin-17, macrophage inflammatory protein 1 α, and tumor necrosis factor-α in culture as well as the ability of B. bronchiseptica to adhere to M cells. To directly measure the response of M cells to B. bronchiseptica in vivo, laser capture microdissection was used to isolate M cells, and gene expression changes were compared during different stages of B. bronchiseptica infection. These results will inform our understanding of the initial interactions between Bordetella and host cells, which will impact the development of an appropriate immune response.
Multi-drug resistance efflux pumps confer additional resistance against host innate immune defense via induction of genes for biofilm formation and virulence (MPF3P.803)
Authors: Gupta Goutam , Wren Melinda , Ganguly Kumkum , Pardington Paige 
Published: January 2023

Abstract
Multi-drug resistance, resistance-nodulation-differentiation efflux pumps in gram-negative bacteria are membrane protein complexes that extrude a wide variety of drugs before they can act on their bacterial target. However, they are not merely pumps for physical transport of drugs but their expression level is also up-regulated by the presence of drugs inside the bacteria, which, in turn, facilitates the release of quorum-sensing molecules and subsequent induction of genes responsible for biofilm formation and virulence. It is our hypothesis that efflux mediated biofilm formation and virulence expression offer gram-negative bacteria additional resistance not only against drugs but also against host innate immune defense. In order to test our hypothesis, we performed real-time qPCR studies on Fluidigm platform to monitor the expression of (i) the efflux genes and their regulators, (ii) the genes for synthesis of quorum sensing molecules and their regulators, and (iii) the genes for biofilm formation and virulence. For this, we used Pseudomonas aeruginosa and its efflux and quorum sensing mutants. The gene expression studies were performed for planktonic cells and nascent and mature biofilms in the presence and absence of a drug, ciprofloxacin. We were able to show efflux mediated expression of biofilm and virulence genes, which resulted in the abrogation of host innate immune defense in a cellular assay.
Normoxic induction of HIF-1α in macrophages is required for protection from the fungal pathogen H. capsulatum. (MPF3P.807)
Authors: Fecher Roger , Deepe George 
Published: January 2023

Abstract
Histoplasma capsulatum (Hc) is the most common cause of respiratory infection among endemic mycoses. Macrophages (MΦ) are the primary phagocytic cell that engulfs Hc. Recent data has demonstrated that the transcription factor hypoxia inducible factor 1α (HIF-1α) can be induced by pro-inflammatory cytokines or LPS in normoxia. Because HIF-1α regulates a number of factors that are potentially vital in MΦ responses to Hc, we hypothesized that myeloid HIF-1α exerts a critical role in conferring protection to Hc. Indeed, the absence of myeloid HIF-1α led to significantly higher fungal burden at 7 days post-infection and a 75% reduction in survival following Hc infection. In vitro, the HIF-1α isoform i.1 (HIFi.1) and downstream target genes were transcriptionally induced in Hc-infected bone marrow derived MΦ in normoxia. To determine if HIFi.1 induction occurs in vivo, mice were infected with a Hc strain expressing GFP. At 7 days post infection, GFP+ (infected) and GFP- (uninfected) MΦ were isolated from lungs; surprisingly both GFP+ and GFP- phagocytes exhibited induction of HIFi.1 and its target genes. Transwell assays demonstrated that a soluble mediator released from infected cells was partly responsible for HIF i.1 induction. Genetic deletion of TNFR1 or dectin-1 abrogated the in vitro induction of HIFi.1. We propose that dectin-1 mediated MΦ TNF-α production is responsible, at least in part, for induction of HIFi.1; myeloid HIF-1α is instrumental in the immune response to Hc.
Intricate properties of memory B cell development after oral immunization (MUC4P.836)
Authors: Lycke Nils , Bemark Mats , Komban Rathan , Stensson Anneli 
Published: January 2023

Abstract
Recent debate over the ability to generate memory B cells in the gut immune system prompted us to explore a novel model with transgenic NP-hapten-specific GFP+ B cells and oral immunizations with NP-conjugated to cholera toxin (NP-CT). We found that orally immunized mice developed exceptional long term memory B cell populations that resided in B cell follicles in Peyer’s patches, mesenteric lymph nodes and spleen. A majority of the GFP+ memory cells, defined by CD80, CD73 and PD-L2 expression, were IgM+, but also a significant number were IgA+ and in spleen IgG+ memory B cells were frequent. Upon analysis of the NP binding VH186.2 IgA gene sequences between 20-40 % carried high affinity mutations and VH186.2 IgG (bone marrow) and IgA (intestine and bone marrow) gene sequences were clonally related. A challenge immunization after 1.5 years, either orally or i.p, elicited a strong NP-specific gut lamina propria (LP) IgA and serum IgG response, starting from a calculated frequency of 1 NP-memory / total 10.000 B cells in these mice. Gut LP IgA plasma cells increased after challenge from 0.05% long-lived NP-specific IgA plasma cells to &gt;5% NP-specific IgA plasma cells of all LP IgA plasma cells. The boosted IgA plasma cells carried more mutations and higher affinities (&gt;60%) than long-lived memory B cells, demonstrating unexpectedly effective selection and maturational processes in gut IgA memory responses.
Bacterial surface (S-) layer glycosylation orchestrates oral microbial communities by tuning the host immunity (MUC4P.841)
Authors: Settem Rajendra Prasad , Honma Kiyonobu , Sharma Ashu 
Published: January 2023

Abstract
Human mucosal surfaces begin to get colonized after birth with microbial communities, which co-exist in the host by modulating the host immune responses. In this process, some microbes decorate with unique sugars to mold the host immunity conducive to their survival. Recently we have shown that oral pathogen Tannerella forsythia (Tf) exploits a similar strategy to facilitate its survival in the oral cavity, and subsequently cause the alveolar bone destruction by dampening Th17-dependent neutrophil infiltration. The present study was aimed to assess the role of Tf surface O-glycans in the manipulation of host immunity and test if this might impact the colonization and persistence of other co-habitants, such as Porphyromonas gingivalis (Pg). Periodontitis mouse model was utilized to compare the ability of Tf or its O-glycan deficient mutant ED1 in impacting Pg colonization and induced alveolar bone loss. In comparison to the ED1: Pg infected group, higher alveolar bone loss was observed in the Tf: Pg group. This paralleled with osteoclastic numbers. Elevated numbers of neutrophils observed in the ED1: Pg group over Tf: Pg group, associated with bacterial colonization levels. Our study suggests that Tf tunes host immunity to promote its colonization and of its co-habiting partner P. gingivalis, leading to increased alveolar bone loss and this scenario parallels with clinical settings where the increased periodontitis severity has been observed in patients harboring Pg and Tf.
Colonic goblet cell intrinsic, Myd88 dependent, microbial sensing rapidly regulates antigen delivery to the lamina propria immune system (MUC4P.853)
Authors: Knoop Kathryn , McDonald Keely , McCrate Stephanie , McDole Jeremiah , Newberry Rodney 
Published: January 2023

Abstract
The delivery of antigens across the intestinal epithelium is an early and critical event in homeostatic responses to innocuous antigens and inflammatory responses to potential pathogens. The factors and mechanisms regulating antigen delivery to the lamina propria (LP) immune system are largely unknown. We recently demonstrated when small intestinal goblet cells (GCs) secrete, they form goblet cell-associated antigen passages (GAPs) and deliver luminal antigens to LP dendritic cells (DCs). Here we report in the basal state, GAP formation is driven by acetylcholine (Ach) signaling via the muscarinic ACh receptor 4 (mAChR4) expressed by GCs. In contrast to conventionally housed mice, GAPs were present in the colon of mice with a reduced microbiota or altered microbial sensing. Formation of colonic GAPs in these mice was dependent upon mAChR4 signaling. Moreover, in mice with a reduced microbial load, luminal microbial stimuli inhibited GAP formation within minutes. In vitro studies revealed GC intrinsic sensing of the microbiota rapidly inhibited ACh induced GC secretion, and conditional deletion of Myd88 in GCs in conventionally housed mice resulted in the spontaneous formation of colonic GAPs, and the delivery of luminal antigen to colonic LP DCs. These findings identify a role for microbial sensing by colonic GCs to rapidly regulate antigen delivery to the LP immune system, and preventing delivery of luminal antigens when the luminal microbial load is abundant and complex.
Gestational hypothyroidism in mice makes female offspring more resistant to pneumococcal pneumonia (MUC5P.869)
Authors: Nieto Pamela , Peñaloza Hernán , Salazar Francisco , Kalergis Alexis , Riedel Claudia , Bueno Susan 
Published: January 2023

Abstract
Gestational hypothyroidism affects fetus neurodevelopment and leads to cognitive deficiencies. In rodents, this condition also affects the central nervous system and produces differences in the amount and type of immune cells in spleen. We wanted to study if the offspring gestated under hypothyroidism have alterations in the performance and/or activation state of the immune cells during an infection. To evaluate this, we studied the immune response against the pathogen Streptococcus pneumoniae in female mice gestated in hypothyroid mothers (hypo mice) and in control mothers (untreated mice), which were infected intranasally to cause pneumonia. A survival curve showed that hypo mice were more resistant to the infection. Hypo mice also showed an increase in the IL-17 levels in BALF. Other parameters such as immune cell populations, histopathology, and bacterial load were also evaluated. For the first time, our study addresses whether there is a correlation between gestational hypothyroidism and the functionality of the offspring immune cells during an infection.
Both local antigen and inflammation are required to establish resident memory CD8 T cell populations in the lung parenchyma and airways (MUC5P.866)
Authors: McMaster Sean , Wilson Jarad , Wang Hong , Kohlmeier Jacob 
Published: January 2023

Abstract
Microenvironment cues regulating the establishment and maintenance of a pathogen-specific resident memory T (TRM) cell population in the lung parenchyma and airways are poorly understood. We show that local inflammation is sufficient to “pull” systemic antigen-specific effector T cells into the lung and airways acutely, but these cells are not maintained and do not develop into long-lived TRM. However, administration of both local antigen and inflammation is sufficient to foster the establishment of long-lasting TRM populations in both the lung parenchyma and airways; moreover, local antigen and inflammation is able to compensate for the defect in airway-resident T cell memory (TARM) recruitment in mice infected with influenza virus via the intramuscular route as compared to the intranasal route. We also show that mice administered local antigen and inflammation are more protected upon heterologous challenge as compared to mice receiving either local inflammation or local antigen alone. This observation correlates well with our finding that the airway and lung parenchyma TRM populations are not systemically maintained. Follow-up experiments suggest a hypothesis that the lung TRM population could be the replicative reservoir for the TARM population. Our findings could further the development of cellular-mediated vaccines against respiratory pathogens by defining the parameters for the establishment of pathogen-specific TRM populations in the lung parenchyma and airways.
Compensation-free dead cell exclusion: multi-beam excitation of the far-red DNA binding viability dye DRAQ7. (TECH2P.873)
Authors: Edward Roy , Dimmick Ian 
Published: January 2023

Abstract
Reliable exclusion of dead/damaged cells is often needed for immunological assays by flow cytometry and cell sorting. DRAQ7 is a far-red biomarker of cell viability developed for this purpose. This water-soluble molecule binds dsDNA with high affinity but is membrane impermeant, only labeling cells with compromised membranes (Akagi, J. et al., 2013 and Smith, P.J. et al., 2013). Multi-colour analysis is limited by the available spectrum and preference for bright fluors, increasing compensation issues. DRAQ7 aids panel design by its emission: not overlapping Brilliant Violet dyes, FITC or R-PE and minimally with APC. Due to its specificity and affinity it provides clear s:n in membrane-compromised cells. DRAQ7 is optimally excited at 599 / 644 nm, however it is usefully excited down to blue wavelengths. In principle, this, so called, multi-beam excitation means DRAQ7 occupies multiple fluorescence channels. Conversely, this actually enables selection of a unique population of dead cells; plotting two independently excited far-red channels against each other one can choose a suitable pairing that gives the best separation of a double-positive population for a chromophore panel. This double-positive population is defined as a dead-cell gate of events then entirely excluded from other channels (avoiding any compensation). This can be achieved during acquisition or post-acquisition. The technical principles of this simple approach and exemplar data will be presented.
Identification of CXCR2 as an important regulator of granulocytic myeloid-derived suppressor cell mobilization during tumor progression (TUM4P.912)
Authors: Bian Zhen , Shi Lei , Liu Yuan 
Published: January 2023

Abstract
Expansion of myeloid-derived suppressor cells (MDSCs) is associated with cancer progression and suppression of T cell function. However, further defining MDSCs and their role is hindered by the fact that no specific cell marker is available to distinguish MDSCs from non-suppressive leukocytes. Here we report a separation method that clearly separate granulocytic MDSC (G-MDSC) from non-suppressive granulocytes. In this study, mouse bone marrow cells from healthy and tumor bearing mice were collected, followed by Percoll density gradients centrifugation. Naive CD11b+Ly6G+Ly6Clow granulocytes from the fraction between 50-60% Percoll effectively inhibited the proliferation of T cells via production of ROS and arginase. These suppressive cells were expanded during tumor formation and the inhibitory function was enhanced in tumor bearing mice, indicating that G-MDSC was enriched at the 50-60% Percoll interface. Unlike G-MDSC, high density granulocyte (~60-70% Percoll fraction) failed to suppress T cells proliferation but had a high phagocytic activity and chemotatic capacity toward CXCL1, suggesting that they were mature neutrophils. Further study found that CXCR2 in G-MDSC from tumor bearing mice was significantly upregulated, leading to a massive migration of G-MDSC into spleen and tumor site. Collectively, our findings provide a new way to discriminate G-MDSC from mature granulocytes and reveal a critical role of CXCR2 in G-MDSC mobilization during tumor formation.
MDSC-derived and TIL-endogenous NO plays a crucial role in the induction and stability of Th17 cells in human ovarian cancer (TUM4P.915)
Authors: Obermajer Natasa , Wong Jeffrey , Edwards Robert , Chen Kong , Scott Melanie , Khader Shabaana , Kolls Jay , Odunsi Kunle , Billiar Timothy , Kalinski Pawel 
Published: January 2023

Abstract
Intratumoral Th17 numbers and local production of IL-17 have been shown to positively correlate with survival of ovarian cancer (OvCa) patients. We demonstrate that the tumor-associated expression of IL-17A in OvCa patients positively correlates with the expression of NOS-2. Our data indicate that the nitric oxide synthase-2 (NOS2/iNOS)- driven NO produced by patients’ myeloid-derived suppressor cells (MDSCs) promotes the development of RORgt(Rorc)+IL-23R+IL-17+ Th17 cells. In addition to the positive impact of the exogenous (MDSC-produced) NO, we show that the development of Th17 cells from naive-, memory-, or tumor-infiltrating CD4+ T cells, driven by IL-1β/IL-6/IL-23/NO-producing MDSCs or by recombinant cytokines (IL-1β/IL-6/IL-23), is associated with the induction of endogenous NOS2 and NO production, and critically depends on NOS2 activity within CD4+ T cells. Inhibition of NOS2 activity or the canonical cGMP/cGK pathway of NO signaling abolishes the de novo induction of Th17 cells and selectively suppresses IL-17 production by established Th17 cells isolated from OvCa patients. The current results provide rationale for targeting NO, NOS2, cGMP, and cGK as new targets to manipulate Th17 responses in cancer.
Mast cells and mast cell-derived IL-13 play an important role in MDSC activation, migration, and accumulation. (TUM4P.925)
Authors: Damle Sheela , Martin Rebecca , Saleem Sheinei , Folgosa Lauren , Zellner Hannah , Nguyen Kim , Ryan John , Bear Harry , Irani Anne Marie , Conrad Daniel 
Published: January 2023

Abstract
Until recently, the interaction between mast cells (MCs) and CD11b+Gr-1+ myeloid derived suppressor cells (MDSCs) was limited to recruitment of MDSCs to the tumor site. However, we report that MCs are also needed for the activity of MDSCs. Adoptive transfer (AT) of MDSCs failed to promote B16 melanoma colonization in MC deficient mice. MDSCs in these mice also failed to localize to the liver and stayed mainly in peripheral blood. MC sufficient mice accumulated MDSCs in the liver and retained significantly lower levels of MDSCs in circulation after AT. Recently, Ma et al. (Cancer Res. 2013 Jul 1; 73(13);3927-37.) has shown that MC derived IL-13 has been show to promote the growth of pancreatic ductal carcinoma. Given this, we investigated the role that IL-13 plays in the MDSC/MC interaction. Our data shows that IL-13 KO mice crossed to ADAM10Tg mice, which carry a mutation resulting in the over-accumulation of MDSCs, have significantly decreased MDSC accumulation in the naïve state, especially evident in the granulocytic subset. In a model of natural MDSC accumulation (Lewis Lung Carcinoma, i.v.), IL-13 KO mice had reduced tumor colonization to the lungs. They also had reduced MDSC accumulation in the liver and spleen, but increased number in circulation. Taken together, our data indicates that MCs and IL-13, an important MC-derived cytokine, play an important role in MDSC migration and accumulation and represent important drug targets for the control of MDSCs in tumor.
Impact of non-oncogenic acute viral infection on anti-tumor CD8+ T cell responses (TUM5P.938)
Authors: Zloza Andrew , Huelsmann Erica , Lacek Andrew , Lusciks Eugene , Nabatiyan Arman 
Published: January 2023

Abstract
Background: Factors dictating clinical emergence of cancer are not fully understood. Specifically, how non-oncogenic acute viral infections affect unrelated anti-tumor CD8+ T cell responses (those in uninfected tissues) is unknown. Methods: Therefore, we infected B6 mice with non-lethal Influenza (A/PR/8-OVA, 80,000 EID50, intranasally) 3 days prior to challenge with B16-F10 melanoma (120,000 cells, subcutaneously, right flank). To track self/tumor antigen, virus-expressing antigen, and unrelated foreign antigen CD8+ T cells, we transferred pmel (gp100-specific, Thy1.1+Ly5.1-), OT-I (OVA-specific, Thy1.1+Ly5.1+), and P14 (LCMV gp33, Thy1.1-Ly5.1-) CD8+ T cells, respectively, into Thy1.1-Ly5.1+ B6 mice. Results: Influenza infection significantly worsened normal host anti-tumor responses and decreased host survival with cancer from ~32 to ~15 days (P&lt;0.01). Interestingly, pmel (but not OT-I and P14) CD8+ T cells circulating to the infected lung underwent cell death resulting in their eventual loss from the tumor microenvironment (up to 90%; P&lt;0.001). Similar findings were observed with infections 15 or 30 days before or 7 days after B16 challenge and when Balb/c or humanized NSG mice were challenged with breast cancer or melanoma in the context of LCMV or HIV. Conclusions: Our findings may explain the clinical phenomenon emerging from epidemiologic studies reporting increased unrelated cancer in patients with non-oncogenic viral infections.