bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tumor progression locus 2 inhibits regulatory T cell development and immunosuppressive functions (LYM8P.640)
Authors: Li Xin , Acuff Nicole , Peeks Angela , Kirkland Rebecca , Hardwick Kara , Nagy Tamas , Watford Wendy 
Published: January 2023

Abstract
Regulatory T cells (Tregs) are a specialized subset of immunosuppressive T cells that function to maintain peripheral tolerance. We previously demonstrated that the serine-threonine kinase, tumor progression locus 2 (Tpl2, also designated Cot/Map3k8), regulates TCR signaling and inflammatory cytokine secretion in CD4+ T cells. Herein, we demonstrate that Tpl2 is preferentially expressed by Tregs and regulates their development and functions. Tpl2-/- mice exhibited increased proportions of thymic natural Tregs (nTregs) as well as peripheral splenic Tregs, in vivo. Enhanced Treg development was due to a T cell autonomous defect, since Treg development from tpl2-/- naïve T cells cultured in vitro was similarly enhanced, and peripheral Treg proportions were also increased within the tpl2-/- donor compartment of mixed bone marrow chimeras. This defect depended upon TCR signal strength and was enhanced at low antigen concentrations. Importantly, tpl2-/- Tregs had increased expression of the immunosupressive cytokines, IL-10 and IL-35, and provided better protection than wild-type Tregs in vivo in a T cell transfer model of colitis. These results demonstrate that Tpl2 has an important physiological role in limiting Foxp3 expression, Treg development and functions. Therefore, Tpl2 inhibition could potentially be used to deviate pathological immune responses in a variety of autoimmune diseases towards a protective, tolerogenic response through preferential Treg induction.
Blood transcriptional signatures of early tuberculosis infection and disease in cynomolgus macaque model (MPF6P.658)
Authors: Gideon Hannah , Skinner Jason , Flynn JoAnne , Lin Philana 
Published: January 2023

Abstract
Mycobacterium tuberculosis (Mtb) remains one of the successful human bacterial pathogen, yet little is known about the factors that influence infection outcome. The Cynomolgus macaque model of tuberculosis (TB) recapitulates many of the manifestations of human TB and provides an excellent platform to study early disease progesssion, which is not tractable in the humans. Macaques (n=38) were infected with low dose of Mtb (Erdman strain, ~25CFU) and whole blood samples were obtained for transcriptional analysis at serial time-points (pre infection to 6 months post infection) when the animals were declared to have clinically active TB or latent infection. Linear Mixed Modeling Analysis (LMMA) revealed increased activity of transcript clusters associated with interferon response, inflammation, hematopoiesis, early immune regulation, innate, and acquired immune responses between 20-56 days post Mtb infection in all animals. Despite considerable overlap in the transcriptional signatures of animals that developed active TB (n=16) or remained with latent infection (n=22), cross correlation of transcript cluster activity and disease activity (scan score) revealed significant immunologically relevant associations especially with interferon and lymphocyte responses that differed in composition and timing between active and latent cohorts. In summary, this study elucidates novel associations of infection outcome and the timing and composition of transcriptional signatures in TB.
IL2RA variants that reduce IL-2 signaling through its receptor aggravate cutaneous leishmaniasis (MPF6P.700)
Authors: Dessein Alain , Oliveira Pablo , Dessein Helia , Romano Audrey , Cabantous Sandrine , pereira Valeria , de Brito Maria , Rodrigues Virmondes , Rafati Sima 
Published: January 2023

Abstract
Leishmania infection may be asymptomatic, cause mild or severe skin ulcers (cutaneous leishmaniasis, CL), limited or disseminated lesions, or lethal visceral disease. We previously reported an association between IL-2 receptor mutations and susceptibility to visceral leishmaniasis in children infected with Leishmania donovani. Here, we evaluated the possible role of IL-2 signaling in human CL. We first showed that transcript abundance was high for several genes of the IL-2 pathway in skin lesions caused by Leishmania braziliensis. We then carried out a genetic analysis, focusing on major genes of the IL-2 pathway. We used a family-based approach and found that polymorphisms of several genes appeared to be associated with CL in a Brazilian population. Moreover, two polymorphisms of the IL2RA gene were significantly and independently associated with CL. We confirmed this result in a second Brazilian sample and in Iranians infected with Leishmania tropica: IL2RA rs10905669 T (Pcombined = 6 x 10-7) and IL2RA rs706778 T (Pcombined = 2 x 10-9) were associated with greater susceptibility to lesion development. These alleles were also correlated with a poor IFN-g response and poor FoxP3+ Treg cell activation. Thus, IL-2 plays a crucial role in protection against the cutaneous ulcers caused by Leishmania, and the IL-2 pathway is a potential target for strategies aiming to control Leishmania-related diseases.
Type I IFN enhances Coxiella burnetii infection (MPF6P.646)
Authors: Hedges Jodi , Robison Amanda , Christensen Kelly , Kimmel Emily , Lucas Erin , Ramstead Andrew , Jutila Mark 
Published: January 2023

Abstract
Coxiella burnetii is an intracellular pathogen and the cause of Q fever. A role for type I IFN in C. burnetii pathogenesis has not been determined but this pathway supports host protection from a related pathogen, Legionella pneumophila. Spleen cells from type I IFN receptor (IFNAR)-deficient mice supported diminished replication of the avirulent Nine Mile phase II strain (NMII) in vitro compared to spleen cells from wild type mice, and addition or induction of type I IFN enhanced replication, suggesting that type I IFN promoted C. burnetii infection. The inverse relationship between type I IFN and inflammasome activation was explored as a potential mechanism of type I IFN-dependent enhanced C. burnetii infection. When human cells were infected with NMII in vitro inflammatory transcripts, such as IL-1β, were strongly induced. This transcript expression was reversed by exogenous induction of the type I IFN pathway. In vivo infection with the virulent phase I strain (NMI) of C. burnetii confirmed in vitro predictions; IFNAR-deficient mice were significantly protected from weight loss compared to wild-type mice, and treatment with rIFNa exacerbated disease. In contrast, IL-1β- and ASC-deficient mice had enhanced bacterial replication in the lungs. The data suggest that type I IFN signaling may dampen IL-1β expression, which is important for protection from C. burnetii.
Elucidating a role for eosinophils and discovering novel vaccine targets in Pneumocystis pneumonia using transcriptomics (MPF6P.651)
Authors: Eddens Taylor , Elsegeiny Waleed , Horne William , Zheng Mingquan 
Published: January 2023

Abstract
Pneumocystis (PC) pneumonia remains the most common serious opportunistic infection in patients with HIV/AIDS and is re-emerging in non-HIV infected immunosuppressed patients. Clearance of PC occurs through CD4+ T-cells with antibodies acting as one documented mechanism of clearance, although protective antibody epitopes have yet to be elucidated. To address the role of CD4+ T-cells, we used transcriptomics in mice over the course of PC infection and we observed a strong eosinophil signature in CD4-replete animals by RNA sequencing and by BAL analysis. Hypereosinophilic CD4-depleted mice receiving pIL5 via hydrodynamic injection had increased eosinophils in the lung and decreased PC burden. Finally, mice deficient in eosinophilopoiesis, had no change in burden with pIL5 treatment following CD4-depletion, further implicating eosinophils as novel mediators of immunity against PC. To identify protective antibody epitopes as potential vaccine candidates, we used a life-cycle based approach and sequenced RNA from the ascus and troph life forms of PC and found GSC-1 (ascus) and Meu10 (troph) were differentially regulated. Both GSC-1 and Meu10 generate strong humoral immune responses. Meu10 peptide vaccination decreases PC burden in CD4+ T-cell depleted animals, while immunization with recombinant GSC-1 failed to provide complete protection in a challenge model. Future studies will evaluate a multivalent life-cycle based vaccine in both a PC challenge model and a co-housing model.
Rescuing cells from apoptosis after induction in cells protected by Toxoplasma infection (MPF7P.720)
Authors: Crouse Elle , Nash Paul 
Published: January 2023

Abstract
CTLL-2, a cell line commonly used to quantify IL-2 levels, undergo apoptosis when IL-2 is not present. We previously demonstrated that infection with Toxoplasma gondii prevents this programmed cell death. We also demonstrated that Toxoplasma infection prevents apoptosis induced by beauvericin. In this study, we undertook to determine whether the protection from apoptosis provided by Toxoplasma infection is partial and only protects to the level of our assay determination, or whether it is complete and the cells could continue to grow once the apoptotic inducer is removed. Of course this is an artificial system since the parasite would normally quickly kill the cell. However, the results could be relevant when cytotoxic T lymphocytes are killing infected cells when at the same time Toxoplasma treatment drugs are employed. More importantly, the information gained from this study will help us better understand inhibition of apoptosis by Toxoplasma and of apoptotic mechanisms. Since apoptosis is involved in so many processes, understanding and potentially being able to modulate its occurrence could be valuable. Here we present the fate of cells that had apoptosis induced, were treated to prevent Toxoplasma from killing the cells, and then maintained under normal conditions.
Exacerbated endothelial activation and type 1 immune responses, but impaired type 2 responses, contribute to severe scrub typhus (MPF7P.706)
Authors: Soong Lynn , Wang Hui , Shelite Thomas R , Liang Yuejin , Saito Tais , Mendell Nicole L , Zhang Wenbo , Sun Jiaren , Gong Bin , Walker David 
Published: January 2023

Abstract
Dysregulated immune responses can contribute to pathogenesis of severe scrub typhus, a neglected tropical disease caused by Gram/LPS-negative Orientia tsutsugamushi bacteria. However, the molecular basis of immune alterations is poorly defined. We have developed a murine model of severe scrub typhus and examined innate regulators in O. tsutsugamushi-infected mice. We found a selective impairment in type 2 cytokines (IL-4, -7, -13), but a significant activation of IL-1β, IL-6, IL-10, TNF-α, IFN-γ, as well as CXCR3- and CXCR1-related chemokines at 2, 6, and 10 days in the lungs, liver and spleen. The elevated angiopoietin (Ang)-2 expression and Ang2/Ang1 ratios suggested the presence of excessive inflammation and the loss of endothelial integrity. These alterations, together with extensive recruitment of myeloperoxidase (MPO)-expressing neutrophils and an influx of T cells, contributed to pronounced apoptosis and acute tissue damage and death. However, targeted depletion of CXCR3 by itself was insufficient to offset progressive disease, although lung edema and vascular leakage were much attenuated in CXCR3-/- mice. We believe that this is the first report of endothelial dysregulation and selective alterations in a panel of immune regulators during early O. tsutsugamushi infection in intravenously inoculated C57BL/6 mice. Our findings shed new light on the pathogenic mechanisms associated with severe scrub typhus and suggest potential targets for therapeutic investigation.
Rac GTPases have opposite roles in T cells and dendritic cells in mucosal immune function (MUC8P.730)
Authors: Cruz Morales Elisa , Song Yan , Marin Romela , Khor Bernard , Nguyen Deanna 
Published: January 2023

Abstract
Variants in RAC1 and RAC2, which encode small Rho-family GTPases, are associated with risk for inflammatory bowel disease. We have shown reduced regulatory T-cell (Treg) proportions in Rac2 knockout (KO) mice with additional dendritic cells (DC)-specific deletion of Rac1 compared to WT. Since Rac is crucial to TCR signaling, we hypothesized that a T-cell specific deletion of Rac1 would also result in defects in Treg development. Treg proportions and susceptibility to dextran sulfate sodium(DSS)-induced intestinal inflammation were assessed in mice deficient in Rac2 in addition to either T cell- or DC-specific deletion of Rac1 (cDKO). Surprisingly, elevated Treg proportions were observed in T-cell specific cDKO mice compared to WT as well as higher Treg proportions within the cDKO compared to the WT compartment in mixed bone marrow chimera. Furthermore, cDKO naïve CD4+ T cells were able to differentiate into Tregs, unlike cDKO DCs, which led to defective Treg generation of WT naïve CD4+ T cells in in vitro Treg generation assays. In vivo Treg generation was reduced in DC-specific cDKO mice compared to WT controls with ovalbumin (OVA) feeding following transfer of CD4+ T cells transgenic for OVA. Lastly, exacerbated DSS colitis was observed in DC-specific, but not T-cell specific cDKO mice, compared to WT. In conclusion, deficiency of Rac proteins result in contrasting effects on mucosal immune function in DCs compared to T cells. The precise mechanism is under investigation.
An NK1.1-expressing regulatory leukocyte in gut-associated lymphoid tissue controls IgA production (MUC8P.733)
Authors: Karmele Erik , Waggoner Stephen 
Published: January 2023

Abstract
Immunoglobulin A (IgA) is the most abundant immunoglobulin in the body and is responsible for maintaining homeostasis at mucosal surfaces. Gut microbes elicit continuous germinal center (GC) reactions within gut associated lymphoid tissues (GALT), which in turn contribute to anti-microbial IgA production by promoting maturation and IgA class switching of B cells that populate the gut lamina propria. Dysregulation of IgA levels is a common disease of unknown etiology. Thus, a better understanding of the mechanisms regulating IgA is critical. We previously found that NK1.1+ cells suppress GC responses and generation of long-lived humoral immunity during systemic viral infection of mice. In the GALT, these NK1.1+ cells may be conventional natural killer cells, group 1 innate lymphoid cells, or natural killer T cells. Surprisingly, systemic depletion of NK1.1+ cells in uninfected specific pathogen free (SPF) mice resulted in an increase in the numbers of GC B cells and T follicular helper cells within GALT (e.g. mesenteric lymph nodes) but not peripheral lymph nodes. In addition, the concentration of IgA in fecal pellets was increased following depletion of NK1.1+ cells when compared to non-depleted control mice. Together our data suggests that an NK1.1-expressing leukocyte inhibits IgA production by impairing GALT GC responses against the gut microbiota. This novel immunoregulatory mechanism may control intestinal homeostasis and prevent development of IgA nephropathy.
An inhibitory receptor leukocyte mono-immunoglobulin-like receptor 3/CD300f deficiency aggravates DSS-induced colitis (MUC8P.737)
Authors: Matsukawa Toshihiro , Teshima Takanori , Okumura Ko , Kitaura Jiro 
Published: January 2023

Abstract
Leukocyte mono-immunoglobulin-like receptor 3 (LMIR3) (also called CD300f) is an inhibitory receptor. LMIR3 is mainly expressed in myeloid cells, including mast cells. The binding of LMIR3 to its ligand ceramide inhibits IgE- and mast cell-dependent allergic responses. It was recently reported that ATP-induced activation of mast cells via P2X7 purinoceptor contributes to the initiation and progression of intestinal inflammation. Here we show that LMIR3 is a negative regulator of dextran sodium sulfate (DSS)-induced colitis, a murine model of ulcerative colitis (UC). Notably, Lmir3-/- mice were highly susceptible to DSS-induced colitis. We found a prominent increase in inflammatory cells, including degranulated mast cells, in the colonic lamina propria of DSS-treated Lmir3-/- mice. Mast cell-deficient KitWsh/Wsh mice engrafted with LMIR3 deficient bone-marrow-derived mast cells (BMMCs) exhibited increased susceptibility to DSS-induced colitis as compared with those with wild-type (WT) BMMCs. Importantly, ceramide-LMIR3 binding suppressed activation of ATP-stimulated BMMCs. Consistently, treatment with ceramide liposomes improved DSS-induced colitis in WT mice, but not in Lmir3-/- mice. Taken together, these results indicated that LMIR3 negatively regulated DSS-induced colitis by inhibiting ATP-mediated mast cell activation through ceramide-LMIR3 binding. Accordingly, LMIR3-targeted ceramide liposomes would provide novel therapeutic strategies for UC.
CCR9 regulates colitis development by inhibiting regulatory T cell development (MUC8P.725)
Authors: Evans-Marin Heather , Cao Anthony , Yao Suxia , Chen Feidi , Gonzalez Maria , He Chong , Liu Han , Wu Wei , Dann Sara , Cong Yingzi 
Published: January 2023

Abstract
T cells reactive to microbiota regulate the pathogenesis of inflammatory bowel disease (IBD). As T cell trafficking to intestines is regulated through interactions between highly specific chemokine-chemokine receptors, great efforts have been made to develop intestinal specific immunosuppression based on blocking these key processes. CCR9, a gut-trophic chemokine receptor expressed by lymphocytes and dendritic cells, has been implicated in regulation of IBD through mediating recruitment of T cells to inflamed sites. However, the role of CCR9 in inducing and sustaining inflammation in the context of IBD is poorly understood. In the current study, we demonstrate that CCR9 inhibits Treg cell development, which contributes to its regulation of intestinal inflammation. While CCR9-/- mice are more resistant to disease compared to wild type (WT) mice upon DSS insults, CCR9 deficiency does not affect effector T cell induction of colitis in a microbiota antigen specific T cell-mediated model. Interestingly, CCR9-/- mice demonstrate a high level of Foxp3+ Tregs, and ligation of CCR9 by its ligand CCL25 inhibited Treg cell differentiation in vitro. Furthermore, partial depletion of Tregs in CCR9-/- mice increased susceptibility to DSS insults to a level similar to WT mice. Collectively, our data indicates that CCR9 signaling inhibits Treg cell development, which contributes to its regulation of colitis development, in addition to acting as a gut-homing molecule.
IL-10 engages macrophages to shift Th17 cytokine dependency and pathogenicity during T cell-mediated colitis (MUC8P.729)
Authors: Geiger Terrence , Li Bofeng , Gurung Prajwal , Malireddi RK , Vogel Peter , Kanneganti Thirumala-Devi 
Published: January 2023

Abstract
Polymorphisms attenuating IL-10 signaling confer genetic risk for inflammatory bowel disease. Yet how IL-10 prevents mucosal autoinflammation is incompletely understood. We demonstrate using lineage-specific deletions of IL-10Rα that IL-10 acts primarily through macrophages to limit colitis. A macrophage-selective deficiency of IL-10Rα increases the severity of colitis induced by T cell transfer into Rag1-/- mice to the same extent as germline deletion of the gene. Colitis mediated by T cell transfer is Th17 cell dependent regardless of the expression of IL-10Rα. Disease development requires IL-6 to support pathologic Th17 cell generation in wild type mice. However, specific ablation of macrophage IL-10Rα provokes excessive IL-1β production that overrides Th17 IL-6 dependence, amplifying the colonic Th17 response and disease severity. IL-10 not only inhibits pro-IL-1β production transcriptionally in macrophages, but suppresses caspase-1 activation and caspase-1 dependent maturation of pro-IL-1β to IL-1β. Therefore lineage-specific effects of IL-10 skew the cytokine dependency of Th17 development required for colitis pathogenesis. Coordinated interventions may be needed to fully suppress Th17-mediated immunopathology.
Dynamics of intestinal IgA expression after early-life antibiotic treatment. (MUC9P.738)
Authors: Ruiz Victoria , Teitler Isabel , Ou Amy , Weber Laura , Chess Edith , Battaglia Thomas , Cadwell Ken , Blaser Martin 
Published: January 2023

Abstract
Class IgA secretory antibodies (SigA) are an early line of antigen-specific immune defenses that are influenced by commensal microbial populations. Perinatal antibiotic exposures may affect the host microbiota and have long-term consequences on host immunity. We hypothesize that early-life pulsed antibiotic treatment (PAT) perturbs the intestinal microbiota leading to alterations in sIgA expression. To test this hypothesis, C57BL/6 mice were exposed to 5 therapeutic doses of the β-lactam, amoxicillin, or the macrolide, tylosin. Exposure to either the tylosin or amoxicillin delayed sIgA expression. sIgA expression recovered in β-lactam-exposed mice but was abrogated with macrolide exposure. Compared to age-matched controls, 1 or 3 tylosin pulses decreased sIgA levels by 40% and 75%, respectively. Alterations in sIgA production were associated with decreased bacterial richness (α-diversity), altered community structure (β-diversity), and changes in microbial compositional, including increased abundance of Enterobacteriaceae and Bacteroides uniformis. Whereas tylosin had multiple immunological effects on conventional mice, essentially none were detected in germ-free animals; indicating that an antibiotic-perturbed microbiota influences developing immunity. These results provide evidence that early-life alterations of the intestinal microbiota by PAT modulates sIgA production, phenotypes that may persist after microbial recovery, with long-term consequences to host immunity.
Comparison of flow cytometers using the side population technique to identify stem cells (TECH2P.900)
Authors: Guenther Garret , Telford William 
Published: January 2023

Abstract
Traditional methods using the side population analysis to identify stem cells involved the use of a flow cytometer with an ultra violet laser excited dye. This presents some challenges for researchers in relation to costs, and even though other dyes such as DyeCycle Violet (DCV) are now available that use the more economical violet laser, the cost of a flow cytometer to provide this type of analysis is still an obstacle. More recently, new benchtop flow cytometers have become available that can provide the quality of a high-end flow cytometer at a much more affordable cost. In this analysis, we sought out to identify stem cells and progenitors using the side population technique in murine bone marrow using the ACEA NovoCyte flow cytometer which maximizes optical design through its use of shared PMTs. Samples were stained and evaluated with (DCV), Lineage markers, Sca, and c-kit. To identify stem cells using the side population technique, we first gated on Lineage- cells and subsequently on Sca+, c-kit+. To confirm the correct identification of these cells, this population was back gated comparing DCV red fluorescence vs DCV blue fluorescence and verified the cells are localized to the expected tail-region. This study is evidence that quality data can be generated on a budget-friendly flow cytometer and allows researchers with limited resources to use flow cytometry.