bluuebunny/crossref_metadata_2025_split huggingface dataset:
Receptor-interacting protein 3 (RIP3) deficiency inhibits immune cell infiltration and attenuates organ injury in sepsis (INC6P.344)
Authors: Sharma Archna , Matsuo Shingo , Yang Weng-Lang , Wang Zhimin , Wang Ping 
Published: January 2023

Abstract
Sepsis is defined as a systemic inflammatory response syndrome (SIRS), leading to multiple organ dysfunction and lethality. RIP3-dependent necrosis is implicated in driving TNF-α- and sepsis-induced mortality in mice. However, it is unknown if RIP3 deficiency has any impact on immune cell trafficking, which attributes to organ damage in SIRS. To study this, male wild-type (WT) and RIP3-deficient (RIP3-KO) mice were subjected to cecal ligation and puncture (CLP), a sepsis model. Blood and tissue samples were collected at 20 h after CLP for analysis. In our severe sepsis model, the mean survival time of septic RIP3-KO mice was significantly extended from 41 h to 68 h, and their plasma levels of TNF-α, IL-6, AST, ALT and LDH were significantly decreased, compared to septic WT mice. In the lungs, septic RIP3-KO mice preserved a better integrity of microscopic structure with reduced apoptosis, and decreased the levels of IL-6, MIP2 and KC, compared to WT. In the liver, the levels of MIP2 and KC were also decreased in septic RIP3-KO mice. Particularly, the total number of neutrophils in the lungs and liver of septic RIP3-KO mice decreased by 64.6% and 52.2%, respectively, compared to septic WT mice. In addition, the number of natural killer (NK) and CD8T cells in the liver decreased by 63.9% and 50.3%, respectively, in septic RIP3-KO mice. Thus, targeting RIP3 may provide a therapeutic strategy in attenuating organ injury caused by excessive immune cell infiltration during sepsis.
Defining aspects of noncognate splenic iNKT cell help which contribute to humoral memory (INC6P.403)
Authors: Leadbetter Elizabeth , Vomhof-DeKrey Emilie , Lanthier Paula , Veerapen Natacha , Besra Gurdyal 
Published: January 2023

Abstract
Successful induction of pathogen-specific humoral B cell memory responses depends upon help from CD4+ T cells. We find that invariant natural killer T (iNKT) cells, glycolipid-specific, CD1d-restricted innate lymphocytes, can provide both cognate (direct) and noncognate (or indirect) helper signals to enhance B cell humoral responses. In fact, a subset of iNKT cells, termed iNKT follicular helper cells, are the cells which specifically act to promote germinal centers, facilitate optimal B cell memory, and plasma cell differentiation. While both cognate and noncognate iNKT cell help induce class-switched, antigen-specific humoral immune responses, only noncognate iNKT cell help drives formation of humoral memory and plasma cell development. We find that differences in the strength of the TcR signal received by iNKT cells, very early effector cytokine production by iNKT cells, T follicular helper cells, and T follicular regulatory cells all contribute to the observed differences in the ultimate humoral outcome. These findings help to identify the optimal requirements for iNKT induction of humoral memory, which has important implications for the application of glycolipid molecules as humoral adjuvants.
NK cell produced HMGB1 regulates DC maturation and Tim-3 expression (INC6P.339)
Authors: Kaus James , Guanqiao Li , Lotze Michael 
Published: January 2023

Abstract
NK cells critically mediate the maturation and activation of DCs in generating effective immune responses as well as their ability to produce cytokines, kill, and induce autophagy. Central to this maturation process is the NK cell mediated release of HMGB1, a pro-inflammatory cytokine, that plays an extracellular role in regulating immune responses as well as an intracellular role in promoting autophagy, a mechanism that allows stressed cells to survive mutagenic conditions. We examined the critical role of murine wild type (NKWT) and HMGB1 ablated (NKH) NK cells in stimulating DC maturation, using NK:DC co-cultures. In comparison with DCs cultured with NKWT cells, significant decreases in CD80, CD86, CD83, CD40, CCR7 and MHCII expressions were found using NKH co-cultures (p&lt;.05). Furthermore, all-thiol rHMGB1 was able to restore the maturation of NKH to comparable levels (p&gt;.05). Significant changes (p&lt;.01) in the percent of cells expressing high levels of maturation markers CD80, CD86, CD83, MHCII were observed. Decreased expression of maturation markers under experimental conditions confirms that HMGB1 release by NK cells is necessary to induce DC maturation. In addition, anti-Tim-3 antibody coincubation increased CD86, CD80, and CD40 expression, even in the absence of HMGB1. These results demonstrate the importance of HMGB1 in stimulating DC maturation, triggering Tim-3 expression, and influencing DC function even with blockade of Tim-3 receptor.
Control of microtubule-dependent activation of nucleic acid sensors for antiviral host defenses (INM6P.419)
Authors: Chiang Hao-Sen , Zhao Yun , Song Joo-Hye , Liu Song , Wang Ninghai , Terhorst Cox , Sharpe Arlene , Basavappa Megha , Jeffrey Kate , Reinecker Hans-Christian 
Published: January 2023

Abstract
Viral targeting of dynein-based transport mechanisms plays an important role for intracellular movements and replication of viral pathogens. We show that the activation of the microtubule and dynein motor complex-associated guanine nucleotide exchange factor GEF-H1, encoded by Arhgef2, is essential for sensing of foreign RNA by RIG-I-like receptors. GEF-H1-deficient macrophages have a profound defect in the induction of IFN-β following detection of synthetic dsRNAs including HMW and LMW poly(I:C) and 5′ppp-dsRNA. The recognition of viral RNA and synthetic dsRNA in the MAVS pathway required the nucleotide exchange activity of GEF-H1. Furthermore, microtubule networks were required for the activation and interaction of GEF-H1 with TBK1 for IRF3 phosphorylation and subsequent induction of Ifnb1 gene expression. Generation of GEF-H1 deficient mice revealed a pronounced signaling defect that prevented antiviral host responses to encephalomyocarditis virus and influenza A virus. In conclusion, our findings identify GEF-H1 as an antiviral signaling component that directs utilization of TBK1 in the MAVS-dependent nucleic acid detection pathways for the sensing of ssRNA virus infection and induction of IFN-β expression and secretion.
Identification and characterization of potent, selective small molecule RIP2 kinase inhibitors (INM6P.422)
Authors: Lipshutz David , Haile Pamela , Cassillas Linda , Desai Biva , Capriotti Carol , Swift Barbara , Berger Scott , Sun Helen , Mahajan Mukesh , Nagilla Rakesh , Reilly Michael , Gough Peter , Bertin John , Votta Bart 
Published: January 2023

Abstract
NOD1 and NOD2 are cytoplasmic PRRs, which bind peptides derived from bacterial peptidoglycans. Signaling via these receptors involves recruitment and activation of RIP2 kinase, which results in NF-κB- and MAPK-dependent pro-inflammatory cytokine production. To enable a chemical biology approach to evaluate the role of RIP2 in inflammation we screened an in-house compound collection. Optimization of an aminoquinoline template led to the discovery of GSK’214 and related analogs. GSK’214 potently and reversibly inhibited human, rat and mouse RIP2 enzymatic activity (IC50 3-10 nM). Selectivity was demonstrated against a panel of 273 commercially-available kinases. NOD2-mediated signaling induced by muramyl dipeptide (MDP) was used to characterize functional activity, in vitro and in vivo. GSK’214 inhibited cytokine (TNFα, IL1β, IL-6 and IL-8) production (IC50 20 nM) by human monocytes stimulated by MDP, but not by ligands specific for TLR2, TLR4, TLR7, TLR9, IL1R and TNFR. Using an acute mouse peritonitis assay, orally administered GSK’214 inhibited neutrophil influx and cytokine release induced by MDP, but not LPS. Next, we used these inhibitors to probe the role of RIP2 in a murine TNBS-colitis model of intestinal inflammation. Selective inhibition of RIP2 kinase activity reduced disease severity to a degree comparable to prednisolone. These results demonstrate the potential of these inhibitors to elucidate the contribution of RIP2 kinase-dependent signaling in inflammation.
E3 ubiquitin ligase Cbl-b inhibits NLRP3 inflammasome activation by targeting NLRP3 for ubiquitination (INM6P.405)
Authors: Liu Qingjun , Guo Hui , Xiao Yun , Zhao Yixia , Zhang Jian 
Published: January 2023

Abstract
NLRP3 inflammasome has been shown to be involved in antiviral and antibacterial innate immunity, as well as in the response to adjuvants critical for adaptive immunity. However, inappropriate activation of NLRP3 contributes to deleterious inflammatory syndromes. Although recent studies show that NLRP3 undergoes ubiquitination, the mechanisms for negatively regulating the activation of the NLRP3 inflammasome in response to PAMPs or endogenous DAMPs are largely unknown. In this study, we found that Cbl-b selectively inhibits NLRP3 inflammasome activation, leaving the other key innate response pathways. Further indication of a negative regulation by Cbl-b on NLRP3 inflammasome activity is the finding that LPS injection induces a significant mortality in Cbl-b-/- mice compared to wild-type (WT) controls, and this increased mortality in Cbl-b-/- mice is blocked by an IL-1 receptor antagonist. At the molecular levels, Cbl-b binds to NLRP3 upon LPS priming and ATP stimulation independently of ASC. NLRP3 undergoes ubiquitination in WT macrophages upon NLRP3 inflammasome stimulation, whereas its ubiquitination is impaired in Cbl-b-/- macrophages. Interestingly, NLRP3 inflammasome stimulation also induces Cbl-b tyrosine phosphorylation, which is critical for its ubiquitin ligase activity, thus initiating a negative feedback that controls NLRP3 inflammasome activity. Therefore, we provide a previously-unappreciated function for Cbl-b in regulating NLRP3 inflammasome activation.
DNA sensing by innate immune receptors (INM6P.408)
Authors: Xiao Tsan , Jin Tengchuan , Chuenchor Watchalee , Ravilious Geoffrey , Jiang Jiansheng , Smith Patrick 
Published: January 2023

Abstract
Nucleic acids are amongst the most potent stimuli of host cells. Interferons and proinflammatory cytokines induced by nucleic acids have pleiotropic effects on the function of both innate and adaptive immune systems. Several families of innate immune receptors at different subcellular locations have evolved to recognize DNA and RNA molecules from both invading microbes and damaged host tissues. Recent findings on DNA sensing by cytosolic receptors AIM2, IFI16 and DHX36 helicase will be discussed. Structure-function studies of these receptors reveal a common mechanism of sequence-independent recognition through electrostatic attractions. As such, highly charged small molecules or peptides are potential candidates for therapeutic intervention of inflammatory conditions involving nucleic acid sensing. This work is supported by the Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, USA.
Profiling ileal immune response genes in obese rats induced by high-fat diet and adenovirus infection (INM7P.432)
Authors: Sang Yongming , Schade Andrew , Blecha Frank 
Published: January 2023

Abstract
Recent studies have shown that an interaction among high-fat diet (HF), gut microbiota and the immune system evokes chronic inflammation in both intestinal and adipose tissues. To profile signature genes regulated by HF and intestinal infection, we induced obese rats by feeding a HF plus a human adenovirus 36 (Ad-36) infection. Compared to control groups, rats in the HF and HF plus Ad-36 groups showed significant increases in body weight, adiposity index, as well as intestinal permeability at 8 weeks post treatment. RNA-Seq analysis was used to examine the comparative transcriptome of ileal samples from animals of each group. Analysis of differentially expressed genes (DEGs) identified 3,998 genes significantly regulated among treated and control rats. Genome-wide examination of innate immune genes was used to cluster significantly up- and down-regulated DEGs, which might mediate chronic inflammation involved in obesity stimulated by HF and Ad-36 infection. In addition, we identified other prominent DEGs pertinent to transcription factors, steroid/cholesterol /lipid metabolism, as well as many functionally uncharacterized transcripts. Interestingly, pathway analysis suggested that the circadian rhythm and intestinal immune network for IgA production were significantly regulated pathways related to obesity. In summary, our findings identified early gene regulation occurring in the intestine during obesity and viral infection that may influence intestinal inflammation.
Diabetes exacerbates infection severity due to hyperinflammation rather than neutrophil defects (INM7P.425)
Authors: Bruhn Kevin , Pantapalangkoor Paul , Nielsen Travis , Tan Brandon , Bonomo Robert , Duncanson Frederick , Everson Michael , Doherty Terence , Lin Lin , Spellberg Brad 
Published: January 2023

Abstract
Patients with diabetes mellitus are at increased risk of dying from Gram negative bacterial infections. This excess mortality is often attributed to immune suppression, including neutrophil dysfunction. However, diabetic hosts also exhibit increased inflammatory responses to LPS. Here we show that lethal infections with Acinetobacter baumannii in diabetic mice are driven by LPS interactions with TLR4, as opposed to neutrophil defects. Mice made diabetic either by streptozotocin (Type 1) or diet induced obesity (Type 2) were hypersusceptible to lethality from A. baumannii bacteremia, whereas mice made neutropenic by anti-Ly6G administration were not more susceptible to infection. Diabetic infected mice developed severe septic shock, with hypothermia, medical acidosis, and marked elevations in inflammatory cytokines, compared to non-diabetic mice. Diabetic mice with impaired or absent TLR4 function, however, were protected against lethal infection and did not develop sepsis. Collectively, these results suggest that enhanced lethality of A. baumannii infection in diabetic mice is not driven by diabetes-related abrogation of neutrophil function. Rather, diabetic mice are more prone to progression to severe sepsis syndrome and are protected from this progression by impairment of TLR4 function. Short term abrogation of TLR4 signaling is a promising therapeutic approach to the treatment of A. baumannii, and possibly other Gram negative infections in diabetic hosts.
Aberrant insulin signaling results in mTOR suppression and immune dysfunction during diabetic infections. (INM7P.427)
Authors: Thurlow Lance , Richardson Anthony 
Published: January 2023

Abstract
Diabetics have defects in several aspects of immunity that result in increased infection rates; however, the underlying mechanisms of immune dysfunction are not well defined. Previous studies in this area concluded that infections in diabetics are hyper-inflamed as indicated by increased cytokine production. Conversely, despite high levels of pro-inflammatory cytokines, our research shows that innate immune cells from diabetics cannot produce inflammatory effectors such as reactive oxygen species (ROS) and nitric oxide (NO). ROS/RNS generation by immune cells is essential for killing bacteria and requires cells to undergo a metabolic shift reminiscent of Warburg metabolism. This shift is required for NADPH production, the sole reductant needed for ROS/RNS generation. Using murine infection models and cell culture, we show that Toll-like receptor signaling induces Warburg metabolism through activation of the mammalian target of rapamycin (mTOR). Moreover, we show that mTOR signaling is suppressed in diabetics and results in decreased ROS/RNS generation and worse infection outcomes. Finally, we demonstrate that mTOR inhibition in diabetes is caused by the activation of AMP-activated protein kinase (AMPK) as a result of deficient insulin signaling. Thus, diabetic infections are not hyper-inflamed. Instead, immune dysfunction in diabetic infections is caused by aberrant insulin signaling resulting in mTOR suppression and diminished antimicrobial effector production.
Type I IFN signaling promotes Coxiella burnetii infection (INM8P.446)
Authors: Hedges Jodi , Robison Amanda , Christensen Kelly , Ramstead Andrew , Jutila Mark 
Published: January 2023

Abstract
Coxiella burnetii is an obligate intracellular pathogen and the cause of Q fever. The strain Nine Mile Phase I (NMI) is virulent whereas a laboratory passaged avirulent form is referred to as Phase II (NMII). Interferon (IFN)-γ is critical for protection from C. burnetii, but a role for type I IFN has not been determined. Type I IFN supports host protection from a related pathogen, Legionella pneumophila, thus we hypothesized that it would be similarly protective in C. burnetii infection. When murine macrophages were treated with IFN-α in vitro at infection, replication of NMII decreased, however, treatment 24 hours post-infection enhanced replication. Infection of spleen cells from type I IFN receptor (IFNAR)-deficient mice in vitro yielded fewer NMII genomic equivalents compared to wild type, with no change in IFN-γ expression. Thus, type I IFN signaling early in infection protected against NMII replication, but later signaling promoted replication. In vivo studies using the virulent NMI similarly suggested that IFNAR signaling promoted pathogenesis. IFNAR-deficient mice infected with NMI were significantly protected from weight loss compared to wild-type mice, although they had larger spleens and similar C. burnetii genomic equivalents. The data suggest that type I IFN signaling promotes infection with C. burnetii, contrary to the case with L. pneumophila. These results support abrogation of this pathway as a potential therapeutic approach to Q fever.
Autophagy enhances NFκB activity and boosts early anti-fungal immunity (INM8P.447)
Authors: Kanayama Masashi , Inoue Makoto , He You-Wen , Shinohara Mari 
Published: January 2023

Abstract
To avoid excessive inflammation, molecular signaling that triggers immune responses has to be well restrained by various cellular and molecular regulatory mechanisms. However, in infections, such restraints must be removed to exert quick and efficient anti-microbial responses. Here, we report a novel role of autophagy in systemic Candida infection by enhancing early immune responses through increased nuclear factor kappa B (NFκB) activity. First, we found that Atg7 conditional knockout (CKO) mice with LysM-Cre were susceptible to Candida infection. Our ex vivo experiments showed that autophagy did not affect Candida-phagocytosis and inhibition of Candida expansion by macrophages and neutrophils. We also found that phagocytosed conidia were not contained in LC3-positive vesicles. The data suggested that it is unlikely that autophagy played a role in “direct” Candida killing by myeloid cells. Unexpectedly, instead, enhancement of NFκB activation was observed in a certain macrophage subset. Enhancement of NFκB activation was achieved by autophagy-mediated sequestration of an NFκB inhibitor. This allows the macrophages to release chemokines to effectively attract neutrophils to infected sites, resulting in better resistance to Candida in wild-type mice compared to Atg7 CKO mice. Thus, autophagy protects mice from systemic Candida infection by a mechanism distinct from those that protect mice from bacteria and viruses.
Targeting of the surfactant protein A receptor SP-R210L variant by influenza A virus in macrophages. (INM8P.440)
Authors: Yang Linlin , Hu Sanmei , Chroneos Zissis 
Published: January 2023

Abstract
Alveolar macrophages (AM) are critical for controlling IAV infection by limiting virus release and by initiating innate and adaptive immunity. Surfactant proteins A (SP-A) and D (SP-D) are a primary line of defense against IAV. Previous studies demonstrated that the SP-A receptor SP-R210 mediates clearance of SP-A-opsonized pathogens. Macrophages express two variants, SP-R210L and SP-R210S, though SP-R210L is predominant on AMs. Here, we studied the role of SP-R210 in IAV infection. We show that disruption of SP-R210L in vitro blocked IAV infection with H3N2 and H1N1 IAV strains. Flow cytometry and microscopy experiments showed that SP-R210L is required for endocytic trafficking of IAV to the nucleus. Furthermore, SP-R210L-mediated infection of macrophages was coupled to the production of TNFα. Interestingly, SP-R210L-deficient macrophages were hyper-responsive to the TLR7 ligand imiquimod. Accordingly, IAV inhibited SP-R210L expression in a time-dependent manner. To address these results in vivo, we studied transgenic mice with conditional disruption of SP-R210 in macrophages; SP-R210-deficient mice were significantly more susceptible to IAV infection as indicated by increased morbidity, decreased survival, and severe lung histopathology compared to WT littermates. Taken together, these findings support the model that by co-opting SP-R210L in AMs, IAV causes a functional ‘knock-down’, reducing the beneficial function of SP-R210L and leading to excessive lung inflammation.
Signaling and function of STAT3 in NK cells during murine cytomegalovirus infection (INM8P.442)
Authors: Tarrio Margarite , Kallal Lara , Sexl Veronika , Biron Christine 
Published: January 2023

Abstract
Cytokines induced during viral infection regulate NK cell functions to optimize antiviral defense and health. Type 1 IFNs (IFNαβ) and IL-12 employ STAT molecules to mediate their effects. In this study, we characterized STAT3 signaling and function within NK cells during infections of mice with murine cytomegalovirus (MCMV). Basally, NK cells activated STAT3 in response to IL-10, IFNα, and IL-12. During MCMV infection, with high circulating IL-12 and IFNα but not IL-10 levels on D1.5, NK cells had elevated STAT3 and both pSTAT4 and pSTAT3. The pathways for the changes were characterized as IFN-dependent induction of STAT3 with IL-12-activated pSTAT4 and pSTAT3. In comparison to wild-type (wt) littermate controls, mice with an NKp46-cell-specific STAT3 deletion within NK cells had comparable D1.5 responses and viral titers in spleens and livers through D4 of infection. In addition, NK cell numbers were maintained or increased as the infection progressed. Surprisingly, mice specifically lacking STAT3 in NK cells but not wt succumbed to high dose MCMV challenge conditions at D5.5. Thus, IL-12-dependent STAT3 activation is part of an ongoing response to infection, and STAT3 within NK cells plays a critical role in host survival. Ongoing studies are delineating the STAT3-dependent mechanism within NK cells functioning for host protection.