bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rapid, simple, column-free isolation of CD11b+ cells from various mouse organs (INC6P.323)
Authors: Valdez Yanet , Tabatabaei-Zavareh Nooshin , de Jong Susan , Morin Peter , Karas Evan , Kollar Anita , Fairhurst Maureen , Eaves Allen , Thomas Terry 
Published: January 2023

Abstract
CD11b+ cells are distributed throughout the body and act as sentinels for the immune system. The frequency of CD11b+ myeloid cells varies in different organs, ranging from approximately 48 % of the brain myeloid (microglia) cells, 30% of bone marrow and 6% of total nucleated cells in gut and spleen. The integrin CD11b heterodimerizes with CD18 to form the mature complex Mac-1. Mac-1 facilitates a variety of immune cell responses including phagocytosis, adhesion, migration and chemotaxis, all functions that are crucial for normal tissue homeostasis and inflammatory responses. CD11b+ cells are difficult and time consuming to isolate, due to their variable frequency using traditional methods. Here, we describe EasySepTM, a simple, column-free immunomagnetic method for the positive selection of highly purified CD11b+ cells from brain, bone marrow, spleen and gut. Labelled CD11b+ cells are retained in a tube in an EasySepTM magnet while unwanted cells are poured off. Selection can be fully automated using RoboSepTM. Starting with a frequency of 7.6 ± 1.3 % CD11b+ cells in spleen, purities of 93.9 ± 4.5 % (n=41) were achieved. Fully competent cells, as shown by functional assays (phagocytosis, ROS), were isolated from the above tissues. This EasySepTM technology provides a fast and simple method to access highly purified functional CD11b+ cells from a variety of tissues, facilitating their study ex-vivo.
Intoxication reduces the number of alveolar macrophages and increases apoptosis after burn and pulmonary infection (INC6P.307)
Authors: Ippolito Jill , Ramirez Luis 
Published: January 2023

Abstract
Alcohol intoxication is involved in 50% of all burn injuries and results in an increased risk of lung infection and mortality, relative to non-drinking patients. Experimental models of intoxication, burn and Pseudomonas aeruginosa (PA) intratracheal (i.t.) infection have revealed amplified lung inflammation, relative to controls. Since lung inflammation is mediated in part by alveolar macrophages (AMs), we chose to examine AMs in this combined injury model. Mice were subjected to ethanol 30 min prior to a 15% total body surface area scald injury, followed by an i.t. PA infection (3,000 CFUs). At 24 h, bronchoalveolar lavage cells were analyzed by flow cytometry for absolute numbers of AMs. Relative to shams, burn and infected mice had 60% less AMs (p&lt;0.05), while combined injured, intoxicated, burn and infected mice had 80% less AMs (p&lt;0.05). Between burn groups, combined injury had 60% less AMs, relative to burn and infection alone. Lungs from shams revealed 3 apoptotic cells per low powered field. This was increased 2-fold after burn and infection and by 8-fold with combined injury (p&lt;0.05). Combined injured mice also had 5-fold more apoptotic cells than burn and infection alone (p&lt;0.05). These data suggest a decrease in AM numbers may be due to an increase in AM apoptosis. Overall, the loss of AMs may contribute to the exacerbated lung inflammation observed with combined injury. [R01AA012034 (EJK), T32 AA013527 (EJK), F31 AA022566 (JAI), and the Falk Foundation (EJK)].
TIGIT facilitates liver regeneration via limiting natural killer cell over-activation (INM6P.341)
Authors: Sun Rui , Bi Jiacheng , Zheng Xiaodong , Tian Zhigang 
Published: January 2023

Abstract
Innate immune cells, particularly natural killer (NK) cells, are known to negatively regulate liver regeneration, and inhibition of NK cell overactivation could facilitate liver regeneration. However, the molecular mechanisms that limit NK cell overactivation during liver regeneration are still elusive. In our study, we show that T cell Ig and ITIM domain (TIGIT), a coinhibitory receptor, is selectively up-regulated on NK cells, along with high expression of its ligand, poliovirus receptor (PVR/CD155), on hepatocytes during liver regeneration. Moreover, TIGIT absence impairs liver regeneration in vivo, along with NK cell overactivation and higher IFN-γ production by NK cells. We also show that NK cell-derived higher IFN-γ production is the major account of impaired liver regeneration caused by the absence of TIGIT. Adoptive transfer of Tigit-/- NK cells into NK-deficient Nfil3-/- mice sufficiently led to impairment of liver regeneration. On the other hand, silencing PVR in hepatocytes rescued impaired liver regeneration caused by TIGIT deficiency in vivo, while blockade of TIGIT in NK-hepatocyte coculture increased IFN-γ production by NK cells in vitro. Therefore, these results reveal that TIGIT is a safeguard molecule to facilitate liver regeneration through negatively regulating NK-hepatocyte crosstalk.
IL-1β secretion is dependent upon a calcium-mediated interaction with calmodulin (INM6P.329)
Authors: Ainscough joseph 
Published: January 2023

Abstract
IL-1β secretion is a central event in the inflammatory process. Unlike most other cytokines, which are secreted via the classical pathway, 2 independent signals are required for IL-1β release. The first induces the intracellular up-regulation of pro-IL-1β, and the second drives secretion of the bioactive molecule. The release of pro-IL-1β is a complex process involving proteolytic cleavage by caspase-1. However, the exact mechanism of secretion is poorly understood. Here, we sought to identify novel proteins involved in IL-1β secretion and intracellular processing, in order to gain further insight into the mechanism of release. A human proteome microarray containing 19,951 unique proteins was used to identify globally proteins that bind Human recombinant pro-IL-1β. Probes with a signal to noise ratio (SNR) of &gt;3 were defined as relevant biologically. In these analyses, calmodulin was identified with a mean SNR of ~11; one of the strongest interactions recorded. Using an ELISA-based protein binding assay, the interaction between recombinant calmodulin and pro-IL-1β, but not mature IL-1β, was confirmed and shown to be calcium dependent. Finally, using small molecule inhibitors it was demonstrated that both calcium and calmodulin were required for IL-1β secretion by the human monocytic cell line THP1. These data suggest that following calcium influx into the cell, pro-IL-1β interacts with calmodulin and that this interaction is important for the secretion of IL-1β.
IL-33-mediated immune responses contribute to renal pathology and endothelial damage in Orientia tsutsugamushi-infected mice. (INM6P.332)
Authors: Shelite Thomas , Liang Yuejin , Wang Hui , Xu Guang , Sun Jiaren , Gong Bin , Walker David , Soong Lynn 
Published: January 2023

Abstract
Endothelial cells (EC) are the main target for Orientia tsutsugamushi infection, and EC dysfunction is a hallmark of severe scrub typhus in patients. However, very little is known regarding the molecular basis of EC dysfunction and its impact on infection outcomes. To address these issues, we recently developed a mouse model of severe scrub typhus and examined gene expression kinetics for IL-33, a DAMP molecule preferentially expressed in EC. Following the infection, mice showed a tissue-specific expression profile for IL-33 and its ST2 receptor, and both genes were highly expressed in the kidneys at the onset of disease (day 6) and prior to host death (day 10). The attenuated disease in IL-33-/- mice implied a role of IL-33/ST2 signaling in disease pathogenesis. We found that WT mice displayed exacerbated EC activation and apoptosis, as judged by the altered expression of angiopoietin (elevated Ang2/Ang1 ratio), increased endothelin-1 (ET-1) expression, and increased vascular damage. In IL-33-/- mice, however, EC stress and apoptosis were markedly attenuated, partially via the expression of molecules known to promote endothelial integrity (Ang1, eNOS, BCL2). The effect of Ang1-based treatment on disease outcome is being examined. These data indicate that IL-33-mediated immune responses contribute to renal pathology and endothelial damage in severe scrub typhus. This study also points to potential therapeutic targets in the treatment of Orientia-induced acute kidney injury.
The role of SARM1 in Toll-like receptor and viral-induced neuronal apoptosis (INM7P.347)
Authors: Khan Burhan , Mukherjee Piyali , Taylor Katherine , Woods Tyson , Winkler Clayton , Peterson Karin 
Published: January 2023

Abstract
Neuronal apoptosis is a key aspect of several neurological diseases but the mechanisms leading to apoptosis are not well understood. Recent studies have suggested that activation of Pattern Recognition Receptors (PRRs) of neurons leads to neuronal apoptosis, even in the absence of pathogen insult. For example, combined stimulation of neurons through toll-like receptors (TLRs) TLR7 and TLR9 results in significant apoptosis. Using neurons from transgenic mice, we found that TLR-induced apoptosis required endosomal localization of TLRs but was independent of MyD88 signaling. Instead, apoptosis required sterile alpha armadillo motif (SARM1), which localized to the mitochondria following TLR activation and was associated with mitochondrial accumulation in neurites. Deficiency in SARM1 inhibited both mitochondrial accumulation and TLR-induced apoptosis. These studies indicate a previously unknown pathway for TLR-mediated neuronal apoptosis and provide a mechanism for how innate immune responses in the CNS directly induce neuronal damage. SARM1 also induces neuronal apoptosis in a Bunyavirus infection model through another PRR pathway, suggesting that SARM1 may be a common mechanism for innate-immune driven neuronal apoptosis. Our current studies are focused on understanding the interaction of SARM1 with other cellular proteins to identify: 1) the mechanism that regulates SARM1 localization to mitochondria, and 2) the mechanism by which SARM1 influences mitochondrial trafficking.
Macrophage-specific deletion of PPARγ impairs lung function and immune responses in an mouse model of Pseudomonas aeruginosa infection (INM8P.363)
Authors: Curtis Brenda , Palmer Jessica , Hoo Melissa , Thomassen Mary , Kovacs Elizabeth 
Published: January 2023

Abstract
Alveolar macrophages (AM) play a pivotal role in response to infection by first releasing pro-inflammatory molecules and phagocytosing pathogens and then resolving inflammation and clearing apoptotic neutrophils. This is accomplished largely based on their ability to switch from a pro-inflammatory to an anti-inflammatory phenotype. PPARγ is a transcription factor important for promoting resolution. To study the role of AM PPARγ in regulating respiratory function and immune responses in the context of bacterial infection, we utilized macrophage-specific PPARγ knockout mice (Mac PPARγ KO). Wild-type (C57Bl/6) or Mac PPARγ KO mice were given intratracheal Pseudomonas aeruginosa (40,000 CFU) or PBS control. After 24 hours, plethysmography was performed, followed by euthanasia. Compared to other groups, infected Mac PPARγ KO mice had impaired respiratory function, with a 5-fold increase in PenH (p&lt;0.05) and a 50% reduction in minute volume and breath rate (p&lt;0.05). Lungs of infected Mac PPARγ KO mice had dramatically increased cellularity, alveolar collapse, and interstitial thickening. Bacterial dissemination to the spleen was observed. Lastly, interleukin-6 levels were 40-fold higher in infected Mac PPARγ KO, compared to infected controls. Together, these findings suggest AM-specific PPARγ loss prolongs inflammation and weakens the ability to clear P. aeruginosa infection.
Lipid metabolism regulator, Acetyl-coA carboxylase: a possible therapeutic target for dendritic cell-based immunotherapy (IRC10P.414)
Authors: Nguyen Thuy , Kim Jung-Sik , Kang Seong-Jun , Park Chung-Gyu 
Published: January 2023

Abstract
Dendritic cells (DCs) are proficient antigen-presenting cells that regulate the activation and suppression of the innate and adaptive immunity. There has been a growing interest in the regulation of DC function by tailoring metabolic requirements in DC-based immunotherapy of cancer. Here we report that targeting acetyl-CoA carboxylase, a key regulator of de novo fatty acid synthesis and lipid oxidation, facilitates the down-modulation of DC functions during the maturation process. DCs treated with ACC blockade, CP-640186, dramatically down-regulated the expression of co-stimulatory molecules including CD80, CD86 and CD40 together with MHC II(major histocompatibility complex II) during LPS-mediated maturation by a dose-dependent manner. Coincidently, the secretion of pro-inflammatory cytokines (IFN-r, IL-6, IL-12) was decreased as well. Blocking fatty acid synthesis by targeting ACC down-regulated the expression of CCR7, a homing receptor of DCs, but up-regulated the expression of CCR5, a chemokine receptor largely expressed in immature DCs. Our data elucidate the role of fatty-acid synthesis in DC maturation and have implications for the development of metabolism based-approaches in DC-mediated immunotherapies.
Dietary vitamin D suppresses delayed type hypersensitivity without effecting bacterial burden during experimental tuberculosis infection (IRC10P.410)
Authors: Reeme Allison , Robinson Richard 
Published: January 2023

Abstract
Mycobacterium tuberculosis (Mtb), the causative agent of the disease tuberculosis (TB), is a significant and extremely successful intracellular pathogen. Protective immunity against Mtb is initiated by phagocytic cells in the lung that activate CD4+ T cells to produce cytokines essential for combating the infection (e.g. IFNγ). However, a negative consequence of this robust T cell response is damaging immunopathology in the host lungs. Recently there is a renewed interest in using vitamin D (VitD) as a host-directed therapy to help treat TB. We sought to understand the effects of VitD on the immune response using an in vivo mouse model of Mtb infection. When C3HeB/FeJ mice were fed a VitD high (VitDHI) or VitD deficient (VitDLO) diet and aerosol infected with the Mtb strain H37Rv, we did not see a significant difference in bacterial burden in the lung or spleen. However, VitDHI mice had significantly less recruitment of lymphocytes to the lung granuloma and decreased production of IFNγ, indicating that VitD is inducing an anti-inflammatory immune environment. Currently, we are using VitD receptor knockout mice to further understand the specific cell types VitD is exerting its effects on to modulate the immune response. Overall the results from these experiments will help define VitD’s effects on the immune response during Mtb infection and help determine if VitD is a reasonable therapeutic option to reduce TB-associated immunopathology.
In vivo activation of the aryl hydrocarbon receptor during viral infection up-regulates immunoregulatory factors in dendritic cells (IRC10P.411)
Authors: Franchini Anthony , Jin Guang-Bi , Boule Lisbeth , Lawrence B. 
Published: January 2023

Abstract
Activation of the aryl hydrocarbon receptor (AHR) substantially dampens host resistance to infection with influenza A viruses. We previously reported that AHR activation negatively regulates the expansion and differentiation of CD8+ cytotoxic T lymphocytes (CTL), and reduces the ability of dendritic cells (DC) to prime naïve CD8 T cells. However, the mechanism by which AHR activation dampens DC function in vivo is unknown. Given that AHR is a ligand inducible transcription factor, we used unbiased gene expression profiling to identify DC specific pathways modulated by in vivo AHR activation. Lung DCs from mice that were exposed to the prototype AHR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or control were sorted and used for unbias transcriptome analysis with RNA-seq. This approach revealed multiple gene targets of AHR in DCs, which may play a role in modulating DC function. For example, indoleamine-2,3-dioxygenase (Ido1) was highly induced. Accompanying increased Ido1 expression were significant increases in the proportion of plasmacytoid DCs (pDCs) and regulatory T cells. Expanding our findings into an in vitro human system, monocyte-derived DCs from healthy human donors exposed to TCDD and influenza A virus presented a similar suite of altered genes, including elevated Ido1. This is further evidence that AHR activation in DCs intrinsically changes gene expression, inducing an immunoregulatory phenotype that exerts strong influence on the antiviral immune response.
Distinct glucocorticoid sensitivity of Th17 cytokines in murine T hybridomas and primary cells (IRC11P.428)
Authors: Banuelos Jesus , Shin Soon , Lu Nick 
Published: January 2023

Abstract
Cytokine suppression contributes to the anti-inflammatory actions of glucocorticoids (GCs). However, IL-17 has been reported to be resistant to GC regulation in asthma, nasal polyps, and Crohn’s disease. In contrast, a negative GC-response element has been identified in the promoter region of the IL-17 gene and IL-17 was sensitive to GC suppression in a murine asthma model. To clarify whether IL-17 and other Th17 cytokines are sensitive to GC regulation, we generated Th17 hybridoma cells by fusing in vitro differentiated murine (bl/6) Th17 cells with BW5147 thymoma. Two clones selectively expressing IL-17A, IL-17F, IL-22 and GM-CSF were stable for over 30 generations. Additional clones selectively expressing IFN-γ or IL-4, but not Th17 cytokines and BW5147 cells that produce little or no cytokines were used as controls. Dexamethasone, a potent GC, significantly inhibited anti-CD3 and anti-CD28-stimulated IFN-γ, IL-4, as well as IL-17A, IL-17F, and IL-22 in various clones. Interestingly, IL-17A and IL-17F, but not IL-22, were resistant to GC suppression in in vitro differentiated Th17 cells. Furthermore, hybridomas, but not in vitro differentiated Th17 cells underwent GC-induced apoptosis. IL-6 partially rescues hybridomas from GC-induced apoptosis, however it did not modulate IL-17 suppression by GCs. Our findings support that IL-17A and IL-17F are capable of being downregulated by GCs and Th17 cell microenvironment and cellular milleu engender GC resistance.
Progressive differentiation of follicular B helper T cells regulates the germinal center response (IRC11P.430)
Authors: Herman Edward , Weinstein Jason , Lainez Begoña , Licona-Limón Paula , Esplugues Enric , Flavell Richard , Craft Joseph 
Published: January 2023

Abstract
Initiation of a thymus-dependent humoral immune response requires interaction of activated B cells and follicular B helper T (Tfh) cells, a specialized subset of CD4 T cells. Differentiation of Tfh cells results in their localization to the B-cell follicle and germinal center (GC) of secondary lymphoid organs and expression of the transcription factor Bcl6. Here we show Tfh cells differentiate further within the follicle and GC, with development of discrete subsets that regulate distinct aspects of B-cell maturation. Using mice expressing reporter genes for IL-21 and IL-4, we found that Tfh cells initially produce IL-21 mRNA and protein and support selection of high-affinity B cell mutations. Gradually, Tfh cells began to produce IL-4 and while ceasing production of IL-21, and demonstrated the ability to induce class-switch recombination and development of antibody-secreting cells. These changes in phenotype and function occurred as these Tfh21- and Tfh4-subsets exhibited differences in global gene expression as well as localization within the GC. These data support a model in which ongoing Tfh-cell differentiation orchestrates the formation of an effective antibody response.
Regulation of T cell responses by a novel tissue-resident-macrophage derived immunomodulatory molecule-CRIg (IRC11P.434)
Authors: Yuan Xiaomei , Dong Yi , Fu Wenxian 
Published: January 2023

Abstract
Costimulatory and coinhibitory molecules of B7 family and TNF superfamily play critical roles in almost all aspects of T cell biology. Recently, a novel immuno-modulatory molecule -complement receptor of the immunoglobulin superfamily (CRIg), has been identified. CRIg is exclusively expressed on CD11b+ F4/80hi tissue-resident macrophages. In addition to act as a receptor for complement component C3 fragments, CRIg has also been implicated as a B7 family member to suppress T cell proliferation. Our previous studies suggest that CRIg has a regulatory role in dampening the autoimmunity in type 1 diabetes. Here we attempted to study how CRIg regulates T cell activities. Using in vitro T cell culture system, we found that suppression of conventional T (Tconv) cells by CRIg, in a form of Fc fusion protein, was elicited at the early activating, rather than the late dividing phase of T cells. These data suggest that CRIg may preferentially influence T cell activation signaling cascades. More strikingly, we found that CRIg can promote the conversion of inducible regulatory T (iTreg) cells, by synergizing with TGFb signaling. Together, these data define novel mechanisms of regulating adaptive immunity by tissue-resident macrophages.
Neuronal circuits modulate antigen flow through lymph nodes (IRC11P.420)
Authors: Hanes William , Olofsson Peder , Tsaava Tea , Levine Yaakov , Chavan Sangeeta , Tracey Kevin 
Published: January 2023

Abstract
Initial exposure to an antigen, by infection or vaccination, is a critical step in developing protection against subsequent infection. Antigen injected into the dorsum of the mouse hind foot flows to the popliteal lymph node, then to the sciatic lymph node, through the lymphatics and eventually reaching the blood stream. In mice immunized to Keyhole-Limpet Hemocyanin (KLH), flow of IrDye-labeled KLH was restricted through the popliteal and sciatic lymph nodes. Imaging one hour after antigen administration revealed a significant decrease in sciatic and popliteal KLH levels in immunized mice compared to naïve animals (antigen signal in naïve 0.75 ± 0.15 vs sensitized 0.23 ± 0.11, p&lt;0.05). Flow cytometric analysis of lymph node cellular populations revealed that antigen was associated mainly with a CD21+CD23+ mature subset of B-cells (51.9 ± 1.6% of total antigen positive cells)). Blocking neuronal activity with bupivacaine at the lymph nodes of immunized animals resulted in restoration of antigen flow, with an increase in antigen signal (control 0.27 ± 0.016 vs bupivacaine 0.38 ± 0.037, p&lt;0.05). Conversely, direct activation of neuronal signals by noninvasive magnetic stimulation resulted in a significant decrease of antigen trafficking compared to sham controls (sham 1.39 ± 0.18 vs stimulated 0.47 ± 0.11, p&lt;0.001). Taken together, these studies reveal a neuronal circuit that modulates antigen trafficking through a pathway involving a mature subset of B-cells.