bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dissecting the relationship between migration, TCR signaling, and thymic selection in situ (111.44)
Authors: Melichar Heather , Ross Jenny , Herzmark Paul , Robey Ellen 
Published: January 2023

Abstract
T cell receptors (TCRs) on developing thymocytes are screened for appropriate recognition of MHC molecules and self-reactivity before differentiating into mature T cell subsets. However, how thymocyte interactions and TCR signaling relate to developmental outcomes during these selection events in vivo is unclear. We use a thymic slice model, fluorescent calcium reporters and fusion proteins, in conjunction with two-photon imaging techniques, to examine the impact of TCR:peptide affinity and antigen presenting cell type on thymocyte sampling, signaling, and developmental outcome in situ. Low affinity antigen interactions result in multiple, transient interactions, and TCR signals are short and interrupted. In contrast, negative selecting ligands are associated with longer, stable interactions with more sustained signaling. Further, we examined the behavior of male-specific, on-time HY transgenic thymocytes to characterize the behavior and signaling characteristics of cells undergoing positive and negative selection to endogenous antigen in a physiological setting. In sum, these experiments provide insight into the role of cell-cell interactions and TCR signaling in shaping the processes of positive and negative selection in situ.
Differential regulation of V(D)J recombination in multipotent progenitors in bone marrow versus thymus (111.8)
Authors: Borghesi Lisa , Vallejo Abbe , Kardava Lela , Yang Qi , Aites Jennifer 
Published: January 2023

Abstract
V(D)J recombinase activity is expressed in bone marrow (BM) multipotent progenitors (MPPs) that retain the potential to differentiate into B cells in BM and T cells in thymus. However, it remains unclear whether MPPs in each organ exhibit distinct transcriptional requirements for recombinase activity, or whether thymic MPPs imprint the requirements of upstream BM MPPs from which they originate. Here we use V(D)J recombination reporter mice to examine transcriptional requirements in BM versus thymic multipotent subsets. Within BM, the frequency of recombinase+ cells increases from 5% in MPPs to 15% in common lymphoid progenitors (CLPs) and 20% in pre-pro B cells. Within thymus, 10% of early thymic progenitors (ETPs) are recombinase+. While the E47 transcription factor is expressed in both BM and thymus, E47 deficiency selectively ablates V(D)J recombination in the former. BM MPPs, CLPs and pre-pro B cells from E47 knockouts lack detectable recombinase activity. In the thymus, E47 deficiency reduces ETP numbers but V(D)J recombinase activity is maintained. Loss of the related E protein HEB does not diminish recombinase activity in ETPs while functional inhibition of E47:HEB complexes leads to a 40% reduction in TCRβ rearrangements. Thus, V(D)J recombinase activity is regulated differently in MPPs in BM and thymus. E protein transcription factors are essential for recombinase activity in multipotent BM MPPs and CLPs but are dispensable in multipotent thymic ETPs.
Driving HIV-specific vaccine responses in immune suppressed recipients (113.20)
Authors: McEwen Lisa , Bui Cac , Paterson Yvonne , Harn Donald 
Published: January 2023

Abstract
Malaria, TB and HIV-1 remain tremendous disease burdens in much of the world’s population. Vaccines for these diseases are desperately needed. However, vaccine efficacy is likely to be compromised due to the systemic Th2 biasing and immune suppression caused by infection with parasitic helminthes. Our lab and others have shown that helminth infection suppresses Th1-type vaccine-specific responses. Although Sub-Saharan populations will benefit most from these vaccines, a large percentage of these populations live in helminth endemic regions. A goal of our research is to find vaccine vectors that drive significant HIV-specific immune responses in helminth infected recipients, despite the immune status of the individual. In the current study, we demonstrate that administration of an HIV vaccine (Listeria expressing HIV-1 gag protein) drives significant HIV-specific CTL and Th1 responses in mice chronically infected with the helminth parasite Schistosoma mansoni. These results suggest that Listeria vector vaccines likely will drive HIV-specific responses in helminth-infected human populations. Kinetic studies show that the HIV-vaccine-specific responses generated in schistosome infected mice are durable and capable of inducing CD4+ and CD8+ T cell memory. Further, Listeria vectors should be considered in the development of new generation HIV-1, malaria or TB vaccines to be administered to populations in sub-Saharan Africa where helminth infection is endemic.
Vaccine-elicited CD8+ T cells protect against respiratory syncytial virus-induced lung airway obstruction (113.8)
Authors: Lee Sujin , Stokes Kate , Celis Esteban , Moore Martin 
Published: January 2023

Abstract
Respiratory syncytial virus (RSV) is the most important pathogen for lower respiratory tract illness in infants. CD8+ T cells clear RSV and may contribute to future RSV vaccines. RSV itself inhibits CD8+ T cell responses. Compared to CD8+ T cells responding to RSV infection, vaccine-elicited anti-RSV CD8+ T cells are less well studied, and mechanisms for achieving optimally functional anti-RSV CD8+ T cells are not defined. We developed a novel RSV vaccination strategy using combined toll-like receptor (TLR) and CD40 stimulation in BALB/c mice as a model for studying antigen-defined RSV-specific CD8+ T cells. Vaccination of mice with TriVax, a mixture of a M2 82-90 peptide representing an immunodominant RSV CD8 epitope, the TLR agonist poly (I:C), and a costimulatory anti-CD40 antibody resulted in robust, functional CD8+ CTL responses. Using a mucus-inducing RSV challenge strain (rA2-line19F), we demonstrate that TriVax vaccination protected against RSV infection and RSV-induced airway mucus expression, mechanical airway obstruction, and cellular lung inflammation. Compared to RSV challenge alone, TriVax vaccination followed by challenge resulted in increased CD8+ T cell cytokine expression and the rapid appearance of RSV-specific CD8+ T cells in the lung. Despite a large number of cytokine expressing and virus-specific CD8+ T cells in the lung, TriVax vaccination did not cause immunopathology. High-quality vaccine-elicited anti-RSV CD8+ T cells were protective in the lung.
Epigenetic effects of transmethylation micronutrients on gene expression in T cells from patients with active lupus. (116.12)
Authors: Ray Donna , Richardson Bruce 
Published: January 2023

Abstract
T cell DNA demethylation contributes to the development of human lupus. However, the mechanisms causing T cell DNA demethylation in lupus patients are incompletely understood. The replication of DNA methylation patterns during mitosis depends on DNA methyltransferase (Dnmt) enzymatic activity and the methyl donor S-adenosylmethionine (SAM), and is inhibited by S-adenosylhomocysteine (SAH). SAM levels are determined by various dietary nutrients including folate and methionine, and previous work demonstrates that patients with active lupus patients have decreased Dnmt activity. We hypothesized that lupus patients will be more susceptible to environmental changes in folate and methionine due to their already low levels of T cell Dnmt activity compared to controls, resulting in aberrant overexpression of genes normally suppressed by DNA methylation. We cultured PHA-stimulated lupus T cells in media containing physiologic high and low concentrations of folate or methionine, and compared effects on methylation sensitive gene expression (CD11a, CD70, KIR and perforin) by flow cytometry. Lupus T cells cultured in lower concentrations of folate and methionine overexpress CD70, KIR and perforin compared to those cultured in standard RPMI 1640 media, relative to control T cells. These results suggest that folate and methionine may contribute to abnormal T cell gene expression in lupus, and that dietary supplementation may have a beneficial effect.
Identification of a tissue resident alveolar macrophage population susceptible to and permissive of Coxiella burnetii infection. (117.14)
Authors: Calverley Matthew , Harmsen Allen 
Published: January 2023

Abstract
Intracellular bacteria, such as the US biological Class B bacterium Coxiella burnetii, are known to infect alveolar macrophages (AM). However, it is not known if a particular subset of AM, within a relatively heterogeneous population of lung macrophages, is particularly susceptible to bacterial infection. Nor is it known whether certain AM are less efficient at resolving infection. We hypothesized that, as a result of tightly regulated immune function, tissue resident AM would prove susceptible to C. burnetii infection and provide a required niche for bacterial persistence. Dye labeling techniques allowed discrimination of AM subsets during the course of infection. Infected cell populations were identified immunohistochemically and through the use of fluorescent protein expressing C. burnetii. We report here that the tissue resident AM are the predominantly infected AM subset, even at low infective doses; these cells remain infected at least nine days after the onset of infection, indicating a permissive niche for bacterial infection. These tissue resident AM undergo a distinguishing phenotypic change during the progression of infection and account for the cell-type predominating in the early uptake of Coxiella. Identification of the AM subset, both susceptible to and permissive of Coxiella burnetii infection, marks an important step in understanding Coxiella pathogenesis providing insight into general pulmonary innate immune function in response to intracellular bacteria.
Heme oxygenase-1 expression were decreased in monocytes from Systemic Lupus Erithematosus patients (119.6)
Authors: Mackern Juan , Llanos Carolina , Herrada Andrés , Leandro Carreno , Gómez Roberto , Anegon Ignacio , Jacobelli Sergio , Kalergis Alexis 
Published: January 2023

Abstract
Systemic Lupus Erithematosus (SLE) is a complex disease characterized by alterations affecting innate and adaptive immune cells. In SLE, monocytes and dendritic cells (DCs) present a stimulatory phenotype that promotes the activation of T cells. One of the molecules that control monocyte function is Heme oxigenase-1 (HO-1) which possesses anti-inflammatory capacity. The aim of this work was to assess HO-1 levels in monocytes of SLE patients (SLEp) and HC. PBMCs were obtained from 43 SLEp and 30 HC. CD14+ monocytes and CD4+ cells were sorted by FACS and HO-1 expression was measured by RT-PCR. HO-1 protein expression was determined by FACS. Parameters of CD4+ T cell activation such as CD69 and CD25 expression and IL2 production after staphylococcal enterotoxin A (SEA) stimulation were used as surrogate to evaluate the stimulatory capacity of monocytes from PBMCs cultures. MHCII and CD86 expression was analyzed by FACS. HO-1 mRNA levels were decreased in monocytes from SLEp compared to HC (P&lt;0.0075, t test). When HO-1 protein expression were assessed by FACS similar results were observed (relative MFI expression to HC; 0.73 ± 0.05; P&lt;0.0001, t test). When CD4+ activation in response to SEA was evaluated, no differences were found. MHCII and CD86 expression in CD14+ cells showed no differences between SLEp and HC. In conclusion, we found a significant decrease in HO-1 expression in monocytes from SLEp, suggesting that an imbalance in HO-1 could play a role in SLE pathogenesis.
The role and mechanisms of type I IFN pathways in suppressing acute colitis (120.15)
Authors: Zhang Lixia , Guo Beichu 
Published: January 2023

Abstract
Inflammatory bowel diseases (IBD), such as Crohn’s disease and ulcerative colitis, is characterized by chronic inflammation of the gastrointestinal tract. The etiology and pathogenesis are poorly understood, dysregulated immune responses in the gut and a loss of tolerance to intestinal microflora are thought to cause IBD in human patients and in animal models. Recent studies show that he IL-23/Th17 axis plays an important role in the development of IBD. Our recent studies revealed a novel immunoregulatory or immunosuppressive function of TLR-mediated type I IFN (IFNβ/IFNα) pathways on the innate and antigen-specific immune response. The goal of this study is to investigate the role and mechanisms of type 1 IFN in the homeostasis of the intestine and development of colitis. Our results show that I IFN receptor (IFNAR) deficient mice were highly susceptible for the development of DSS-induced colitis as indicated by body weight loss. IFNAR KO mice had significantly higher macroscopical and histopathological colitis scores compared to wt mice. We also found increased infiltration of innate immune cells and Th cells including Th17 cells in intestine and colon tissues in IFNAR KO mice. Furthermore, IFNAR KO mice had enhanced expression of proinflammatory cytokines and chemokines in intestinal tissues after DSS-induced colitis. Together, our studies imply the physiological role of endogenous type I IFNs in inhibiting the development of inflammatory bowel disease.
Loss of p27 enhances gastrointestinal eptithelial malignancy in mice with Smad4 deficiency in T cells. (120.17)
Authors: Kim Byung-Gyu , Choi Sung Hee , Robinson Janet , Letterio John 
Published: January 2023

Abstract
We previously reported that selective loss of Smad4 signaling in T cells (Smad4KO) leads to spontaneous gastrointestinal inflammation and cancer. The expression of p27Kip1 is significantly decreased in the colon of Smad4KO mice, thus we investigated the role of p27Kip1 in the maintenance of intestinal mucosal immune and epithelial homeostasis. Smad4KO and p27Kip1 germ line mutation (Smad4/p27DKO) mice develop spontaneous gastrointestinal inflammation and carcinoma at 1-2 months of age and 3 months of age, respectively. Smad4/p27DKO had little in vitro TGF-β-induced Foxp3+CD4+ T cells (iTregs) and decreased natural Foxp3+CD4+ T cells (nTregs) in vivo, whereas Smad4KO had little iTregs and p27KO had decreased nTregs. Furthermore, Smad4/p27DKO have an increased number of mucosal CD4+ T cells and an increased percentage of memory T cells. The production of pro-inflammatory cytokines including IL-6, IFN-γ and TNF-α was greatly increased in the colon mucosa of Smad4/p27DKO mice. The colon epithelium of Smad4/p27DKO mice exhibit an oncogenic signature with a significant elevation in expression of iNOS, p-stat1 and p-stat3 and a repression of the tumor suppressor gene, 15-PGDH when compared with WT, p27KO and Smad4KO mice. Thus, our data indicate that p27Kip1 deficiency promotes gastrointestinal epithelial malignancy through both enhancement of the activation of pathogenic T cells and through promotion of the expression of pro-inflammatory mediators by mucosal epithelial cells.
Bacteroides fragilis decrease intestinal Enterobacteriacease colonization and reverse metronidazole-induced bacterial translocation (120.22)
Authors: Chen Lee-Wei , Wu Ying-Ying , Hsu Ching-Mei 
Published: January 2023

Abstract
Bacteroides, one of the major bacterial groups in the intestine, maintain a complex relationship with the host. In this study, the role of B. fragilis in intestinal permeability and immunity were examined in C57BL/6 mice receiving oral treatment of metronidazole for 6 days. Metronidazole treatment significantly decreased the number of B. fragilis but increased that of Enterobacteriaceae in intestinal mucosa and lumen. Metronidazole treatment increased intestinal permeability and pathogenic Klebsiella pneumoniae translocation. However, metronidazole did not change the bacterial killing activity and expression of TLR4, TLR2, MyD88, RegIIIβ, RegIIIγ, CRP-ductin, RELMβ, as well as defensin proteins in intestinal mucosa. Supplementation of dead B. fragilis in drinking water reversed the effects of metronidazole on intestinal permeability, Enterobacteriaceae colonization, and bacterial translocation (BT), suggesting that B. fragilis maintains intestinal permeability and decreases BT through reducing the colonization of Enterobacteriaceae rather than enhancing the expression of antibacterial proteins or the bacterial killing activity. This is further supported by that B. fragilis monoassociated mice showed a decrease of K. pneumoniae translocation and no change in bacterial killing activity as well as expression of defensin and nondefensin protein expression except of CRP-ductin when compared with germ free mice.
Gut-tropic DC precursors contribute to intestinal immune tolerance (120.4)
Authors: Villablanca Eduardo , Torregrosa-Paredes Patricia , Cassani Barbara , Wagers Amy , Mora Rodrigo 
Published: January 2023

Abstract
Gut-associated tolerogenic and inflammatory dendritic cells (DC) develop from at least two different bone marrow (BM)-derived precursors. Tolerogenic, but not inflammatory gut-associated DC, express CD103 and synthesize retinoic acid (RA), which contributes to the induction of Foxp3+ regulatory T cells (TREG). How CD103+ DC and/or precursors are recruited to the intestinal mucosa and then acquire RA synthesizing capacity remains unknown. Here, we show that the BM contains a distinct population of DC precursors expressing the gut-specific α4β7 and CCR9 receptor (gut-tropic pre-DC). Interestingly, vitamin A-depleted mice exhibited a marked decrease in gut-tropic pre-DC in the BM, which correlated with the absence of RA-producing DC in gut-associated tissues. Furthermore, gut-tropic pre-DC lacking β7 integrins exhibited decreased capacity to home to Peyer’s patches and the small bowel lamina propria. Importantly, gut-tropic pre-DC were required to give rise to RA-producing DC in the mesenteric lymph node and to efficiently induce antigen-specific TREG. Consistent with a pro-tolerogenic role of gut-tropic pre-DC, selective depletion of DC derived from β7-expressing cells promoted an inflammatory milieu, as shown by exacerbated inflammation in experimental colitis and by impaired oral immune tolerance. Thus, gut-tropic pre-DC give rise to RA-producing tolerogenic DC in the intestinal mucosa and are required to maintain immune homeostasis during inflammation and steady state
Adaptive immunity impaired by defective lymphocyte motility and activation in mice lacking the actin-bundling protein L-plastin (121.18)
Authors: Morley Sharon Celeste , Todd Elizabeth , Deady Lauren 
Published: January 2023

Abstract
Germinal center development, critical for long-term humoral immunity, requires the migration of T and B lymphocytes to defined locations within and around the B cell follicle. Lymphocyte motility is dependent upon the linkage of extracellular adhesive and chemotactic cues to intracellular changes in the actin cytoskeleton. The molecular mechanisms linking extracellular cues and the actin cytoskeleton are not yet fully defined. We have recently identified the actin-bundling protein L-plastin as a requisite intermediary in both B and T lymphocyte motility. L-plastin is required for establishment of polarity following chemokine stimulation, for the integrin-mediated enhancement of transmigraton, and for the phsophorylation of the integrin-associated kinase Pyk-2. L-plastin is also required for mature synapse formation in T cells. In the absence of L-plastin, cell-intrinsic disruptions of T and B cell motility and activation lead to reduced follicular helper T cell generation, diminished germinal center formation and diminished antibody production to both thymus-dependent and thymus-independent antigens. Probing the specific defects in T and B cell location, activation, and survival in L-plastin-deficient mice will reveal novel insights into critical cell:cell interactions during the germinal center reaction. L-plastin thus serves as a critical bridge between adhesive and chemotactic signals and the morphological changes in actin required for T and B cell function.
Effective detection of phosphorylated Stat5 in FoxP3-positive regulatory T cells (121.24)
Authors: Lee Sun Min , O'Donnell Erika , Haron Jay , Reynolds Sue , Prudencio Joy , Golts Natalie 
Published: January 2023

Abstract
Multicolor flow cytometric analysis is an invaluable tool for analyzing unique cell types within heterogeneous primary cell samples, like peripheral blood mononuclear cells (PBMCs). To identify these specific populations, antibodies against cell surface markers are typically used. However, obtaining greater in-depth knowledge often requires analysis of intracellular events, like phosphorylation of signaling molecules or expression of transcription factors. A challenge to staining cells for both cell surface and intracellular markers is the lack of compatibility of the cell fixation and permeabilization agents with the antibodies required for cell surface and intracellular staining. In this report, we developed an optimized protocol for the simultaneous detection of the cell signaling molecule Stat5 together with transcription factors and cell surface markers. We evaluated human PBMCs for phospho-Stat5 signal induction in Treg cells upon stimulation with IL-2. Using the optimized buffer protocol with antibodies at their optimal concentrations for this intracellular staining condition, we detected the phosphorylation of Stat5 in CD4+/CD25bright/FoxP3+ Treg cells in response to IL-2 stimulation. We also observed differential phosphorylation of Stat5 in various lymphocyte subsets, including memory and naive T cells, when human whole blood was stimulated with IL-2, lysed, fixed, permeabilized, and stained for CD3, CD4, CD45RA, CD45RO, T-bet, and pStat5 expression.
Copper zinc superoxide dismutase 1-deficient mice show increased susceptibility to MOG 35-55-induced EAE (123.12)
Authors: Massilamany Chandirasegaran , Gangaplara Arunakumar , Rathnaiah Govardhan , Steffen David , Lee Jaekwon , Reddy Jay 
Published: January 2023

Abstract
Oxidative stress has been implicated in the pathogenesis of various inflammatory disorders including autoimmune diseases. On the contrary, low levels of reactive oxygen species (ROS) could be beneficial and act as secondary messengers for many physiological processes including cell signaling. The steady state formation of pro-oxidants is normally balanced by their consumption of antioxidants such as superoxide dismutase (SOD), glutathione peroxidase and catalase, thus attaining redox homeostasis. Here we have addressed the role of SOD deficiency in the mediation of central nervous system (CNS) autoimmunity. We demonstrate that copper zinc SOD1-deficient C57BL/6 mice develop more severe EAE induced with myelin oligodendrocyte glycoprotein (MOG) 35-55 as compared to wild type mice. Contrary to the expectations, SOD1-deficient mice showed significant reduction in the expansion of antigen-specific T cells including interferon-γ and interleukin-17A production. Likewise, using major histocompatibility complex class II/IAb MOG 35-55 dextramers, we demonstrate that CNS infiltrates from SOD1-deficient animals contain low frequencies of MOG-specific CD4 T cells. The data suggest that SOD1 deficiency might lead to the elevated levels of ROS and alter EAE severity by modulating T cell responses.