bluuebunny/crossref_metadata_2025_split huggingface dataset:
Essential role for Toll-like receptor 9 in the pathogenesis of liver inflammation in a murine model of Crohn’s disease-like ileitis (P3147)
Authors: Omenetti Sara , Brogi Marco , Garg Rekha , Goodman Wendy , Klinman Dennis , Pinzani Massimo , Pizarro Theresa 
Published: January 2023

Abstract
Liver inflammation, a common extraintestinal manifestation of inflammatory bowel disease (IBD), was reported to unexpectedly precede ileitis onset in SAMP1/YitFc (SAMP) mice, a spontaneous model of IBD. We hypothesized that since SAMP possess an inherent epithelial barrier defect that can facilitate bacterial translocation, ultimately into the liver, toll-like receptor (TLR) activation of resident immune cells may occur, leading to liver inflammation. To test this, SAMP X MyD88KO mice were generated and liver inflammation assessed. TLRs were measured by qPCR and TLR9+ cells identified by FACS in isolated non-parenchymal liver cells (NPLs). Cytokines were measured by ELISA from NPLs +/- a TLR9 agonist, and SAMP treated with a TLR9 agonist or antagonist evaluated. Deletion of MyD88 resulted in complete amelioration of SAMP liver inflammation as early as 4, and up to 40, wks vs age-matched WT SAMP (both p&lt;0.001). TLR9 mRNA levels were highest for TLR expression in SAMP vs AKR (parental) control livers (↑2.4-fold, p&lt;0.001) and TLR9 expressed in both NPL F4/80+ MΦs and CD4+ T cells, with specific TLR9 activation modulating TNF, IL-12, and IL-10 (p&lt;0.05). Finally, administration of a TLR9 agonist exacerbated (p&lt;0.001), while a TLR9 antagonist ameliorated, SAMP liver inflammation (vs vehicle, p&lt;0.05). These data indicate a pivotal role for TLR9 in the pathogenesis of liver inflammation in ileitis-prone SAMP mice and suggest its importance in hepatic involvement in patients with IBD.
Clinical and immunological profile of patients with corneal ulcers due to microbial keratitis (P3157)
Authors: Jimenez-Martinez Maria C , Santacruz Concepcion , Estrada-Garcia I Iris , Perez-Tapia Mayra , Garfias Yonathan 
Published: January 2023

Abstract
Aim: To describe clinical and immunological characteristics of human microbial keratitis. Methods. 28 patients with bacterial and fungal keratitis were included. Characteristics of ocular lesions such as size of corneal ulcer (epithelial defect), infiltration, hypopyon, were considered; immunological evaluation included determination of IL-1b, IL-6, IL-8, IL-10, IL-12 and TNF-a by cytometric bead arrays in samples obtained from infected and non-infected eyes; frequency of CD4, CD8, CD19 and CD56 lymphocytes were also determined on peripheral blood mononuclear cells (PMBC). Results. Non-significant differences were observed in area of epithelial defect, stromal infiltration, nor hypopyon, when compared; however, we observed a characteristic immunological profile apparently related to keratitis origin: IL-8 &gt;IL-6 in bacterial keratitis; IL-8&gt;IL-6 with increased frequency of circulating CD3-CD56+ cells in gram-negative keratitis; and IL-8=IL-6 &gt;IL-1b in fungal keratitis. Conclusions. This study is the first in evaluate clinical and immunological characteristics of patients with corneal ulcers, despite we did not observe a clinical profile clearly related with the type of keratitis, characterization of immunological status could aid to understand immune pathophysiological mechanisms underlying each type of keratitis. IL-6 and IL-8 could be a target for therapy in severe microbial keratitis
The effect of an autoimmune-associated allelic variant of protein tyrosine phosphatase nonreceptor 22 on B cell tolerance in a murine model. (P4066)
Authors: Metzler Genita , Rawlings David 
Published: January 2023

Abstract
The C1858T allelic variant of tyrosine phosphatase PTPN22 (resulting in an R620W change of protein product, LYP) is strongly associated with multiple autoimmune diseases. Contradictory findings suggest the LYP-R620W variant exhibits either gain- or loss-of-function activity in antigen-receptor signaling. To model this risk allele in mice, we generated knock-in mice expressing the analogous R619W mutation in the murine ortholog, PEP. While the variant protein exhibits normal stability, its expression significantly alters lymphocyte function and aged knock-in mice exhibit effector T and, transitional, germinal center and age-related B cell expansion, autoantibodies and systemic autoimmunity. Further, PEP-R619W impacts B cell selection and, strikingly, B lineage-restricted variant expression is sufficient to promote autoimmunity. In the current project, we initiated a detailed study of the impact of variant expression on B cell tolerance mechanisms. Our studies were designed to test the hypothesis that while negative selection is intact, peripheral B cells expressing the variant exhibit increased positive selection, culminating in a skewed, autoreactive naïve B cell repertoire that promotes autoimmune pathogenesis. Preliminary data utilizing single cell BCR cloning and deep sequencing technologies to assess autoreactivity and V(D)J family usage in naïve B cell subsets, as well as characterization of negative and peripheral selection using in vivo models, will be presented.
Linear synthetic peptides induce anti-HER-2 antibody responses with Trastuzumab- (Herceptin) like biological activity (P4308)
Authors: Showalter Loral , Czerniecki Brian , Koski Gary 
Published: January 2023

Abstract
The monoclonal antibody Trastuzumab, which recognizes a conformation-dependent epitope on the extracellular domain of HER-2, has revolutionized the treatment of HER-2pos breast cancers. High costs and short half-life of Trastuzumab spurs the search for ways to induce therapeutic anti-HER-2 antibodies in vivo. Linear T cell epitopes have already been discovered for HER-2, and these have shown promise in vaccine studies where strong cell-mediated responses have been generated. Identifying conformation-independent B cell epitopes would allow the construction of linear T-B vaccine constructs that would generate both cellular and humoral anti-HER-2 responses with potential anti-cancer activity. We produced a synthetic peptide library consisting of 62 overlapping 20-mers spanning the HER-2 extracellular domain. These were used to vaccinate Balb/c mice. Resulting antisera were screened for reactivity against peptide and recombinant extracellular domain protein via ELISA, and whole native protein by immuno-fluorescent microscopy and FACS analysis. Sera reactive by all these criteria were compared with Trastuzumab for ability to disrupt HER-2 and HER-2/HER-3 dimer signaling with and without heregulin. We identified linear peptides that induced antibodies capable of blocking AKT as well as HER-3 phosphorylation in breast cancer lines similar to Trastuzumab. This suggests that a linear epitope/vaccination approach may be feasible for generating effective anti-HER-2 antibody responses.
Expression of CXCR5 and its natural ligand CXCL13 in breast cancer (P6278)
Authors: Singh Rajesh , Gupta Pranav , Grizzle William , Singh Shailesh , Lillard, Jr. James 
Published: January 2023

Abstract
Despite recent advancements in understanding of breast cancer (BrCa) molecular features and their impact on clinical behavior, BrCa systemic therapy decision-making is still guided by a constellation of clinico-pathological features, which are derived from histo-pathological analysis of primary BrCa tissues. Chemokines and their corresponding receptors play a crucial role in metastasis and organ specific homing of cancer cells in different cancers including BrCa. CXCL13 is a homeostatic chemokine acts through its cognate receptor, CXCR5. Here, we have provided the evidence that CXCR5 and CXCL13 are significantly elevated in both BrCa cell lines (MCF-7 and MDA-MB-231) and clinical samples. Flow cytometric analysis of BrCa cell lines (MCF-7 and MDA-MB-231) show significantly higher expression of CXCR5 in comparison to normal epithelial cells (MCF-10A). This finding was further confirmed at mRNA expression level by real-time PCR analysis. To determine the clinical significance of CXCR5 expressed by BrCa cell lines, breast cancer tissue microarray was stained for CXCR5 and CXCL13. Aperio scan scope scanning system was used to numerically analyze immuno-histochemical staining of CXCR5 and CXCL13. Our results show significantly higher expression of CXCR5 and CXCL13 (p &lt; 0.001) in cancerous tissues compared to normal breast tissues. These data demonstrate both biological and clinical significance of CXCR5 and CXCL13 expression that may contribute to BrCa metastasis.
NK inhibitory receptor Ly49G2 recognition of MHC I Dk in efficient virus control (P6058)
Authors: Coats Ebony , Xie Xuefang , Deshmukh Umesh , Brown Michael 
Published: January 2023

Abstract
NK cells bearing inhibitory receptor Ly49G2 preferentially respond to and efficiently control murine cytomegalovirus (MCMV) in MA/My and C57L mice with Dk. Thus the role of Ly49G2 and Dk in virus control is under study. One hypothesis to explain the necessity of Ly49G2 and Dk is “missing self” recognition—release of inhibition due to downregulation of MHC I—by Ly49G2 of Dk during infection. To confirm Ly49G2 binding to Dk, we generated reporter cells expressing chimeric Ly49G2. We also performed licensing and cytotoxicity assays to assess Ly49G2+ NK cell licensing and inhibition of Dk, thus confirming Dk as the cognate ligand of Ly49G2. Next we examined Ly49G2 recognition of Dk on infected cells. Recent data has shown MCMV immunoevasin gp34 bound to MHC I stabilizes interactions between inhibitory receptors and MHC I, including the BALB allele of Ly49G2. However as BALB.K mice with Dk have 3-fold higher MCMV titers than MA/My or C57L mice with Dk, we examined the role of gp34 in Ly49G2c57l recognition of Dk. We show preferential association of gp34 to Dk, not Kk. Further, binding assays also confirm gp34 stabilization of Ly49G2mamy binding to Dk on infected cells. As cis Dk expression with Ly49G2 may impact trans binding, we are examining the role of cis binding of Dk. Additionally, signaling may be impacted by gp34, causing “missing self” recognition by Ly49G2. As such, Ly49G2 phosphorylation is being assessed, as well as phosphorylation of downstream inhibitory molecules.
Out of sequence “signal 3” suppresses CD8 T cell proliferation (P6040)
Authors: Urban Stina , Welsh Raymond 
Published: January 2023

Abstract
Type 1 interferons (IFN), the first cytokines to be discovered, have been shown to regulate many aspects of the immune system. Type 1 IFN can act directly on CD8 T cells as a “signal 3” cytokine, which augments proliferation and differentiation of T cells into effectors in response to antigen. Exposure of CD8 T cells to IFN-inducers at the same time as antigen had stimulatory effects, whereas pre-exposure to IFN-inducers was inhibitory, indicating that the timing of IFN exposure was of essence. Pre-exposure of CD8 T cells to the type 1 IFN inducer and TLR agonist poly(I:C) had transient inhibitory effects on CD8 T cell proliferation. CD8 T cells lacking type 1 IFN receptors were resistant to the suppressive effects of poly(I:C), indicating a direct involvement of IFN. Poly(I:C)-pretreated CD8 T cells were refractory to further IFNβ stimulation, as measured by phosphorylation of STAT molecules, including STAT1, 3, 4, and 5. Reduced phospho-STAT levels were not due to a decrease in total STAT protein. Poly(I:C)-pretreated CD8 T cells phosphorylated STAT molecules in response to other cytokines, including IL-6 and IL-15. A decrease in type 1 IFN receptors and increased expression of SOCS1 likely contribute to the unresponsiveness to IFNβ stimulation. These data suggest that IFN-pretreated CD8 T cells are unable to get the positive effects that type 1 IFN provides as a “signal 3” cytokine, and this may partially explain why vaccines function poorly in virus-infected individuals.
Early IL-17 production by intrahepatic T cells is important for adaptive immune responses in viral hepatitis (P6042)
Authors: Jie Zuliang , Hou Lifei , Liang Yuejin , Sun Jiaren 
Published: January 2023

Abstract
This study was conducted to examine the interactions among the innate and adaptive immune components of the liver parenchyma during acute viral hepatitis. Mice were i.v. infected with a recombinant adenovirus. We found a transient, but significant, accumulation of IL-17-producing cells in the liver within the 24 h of infection. In vivo neutralization of IL-17 alleviated the liver injury. Further investigations showed that IL-17 neutralization halted the intrahepatic accumulation of CTL and Th1 cells. Additionally, the serum ALT levels, liver injury and the functions of intrahepatic CTL and Th1 cells were significantly suppressed in IL-17R-/- mice compared to the controls. A majority of the IL-17-producing cells in the liver were γδ T cells. Notably, intrahepatic IL-17+ γδ T cells, but not the IFN-γ+ ones, preferentially expressed IL-7Rα (CD127) on their surface, which coincided with an elevation of hepatocyte-derived IL-7 at 12 h post-infection. IL-7Rα blockade in vivo severely impeded the expansion of IL-17-producing cells following viral infection. In vitro, IL-7 synergized with IL-23 and stimulated IL-17 production from γδ T cells in response to TCRγδ stimulation. Collectively, this study showed that the IL-7/IL-17 cascade played a critical role in initiating the adaptive anti-viral responses in viral hepatitis. The cross talk among hepatocytes, innate and adaptive immune cells may be critical to mount the successful adaptive anti-viral immune response in the liver.
Differential T cell responses to lingering viral antigen following the resolution of infection. (P6044)
Authors: Misumi Ichiro , Whitton Lindsay , Whitmire Jason 
Published: January 2023

Abstract
The contraction phase after acute virus infection is a poorly understood period when virus-specific T cells decline in number but continue to evolve functionally until achieving memory cell homeostasis. After lymphocytic choriomeningitis virus (LCMV) infection, virus-specific CD4+ T cells decline in number gradually over a 1-2 month period before establishing stable levels of memory. We find that naïve virus-specific CD4+ T cells continue to divide in response to residual LCMV antigen for several weeks after the eradication of infectious virus. The cell division of CD4+ T cells during this time is slower than that seen during the acute phase of infection when there is live replicating virus, but faster than the homeostatic cell division observed in long-term immune mice. The viral antigen driving this cell-division has a half-life of about 40 days. The presentation of this antigen does not depend on B cells, FcγRII, or antigen-antibody complexes. In contrast to CD4+ T cells, CD8+ T cells halt cell division within days after acute LCMV infection is resolved, indicating that CD8+ T cell responses are tightly linked to endogenous processing of de novo synthesized virus protein and the cross-presentation of residual viral antigen to CD8+ T cells is inefficient during the contraction phase. Our results indicate that the longevity of residual viral antigen governs the duration of the helper T cell contraction phase, serving to sustain T cell numbers and influence the memory pool.
Recirculation and retention contribute to a population of mature Tregs in the thymus. (P1153)
Authors: Yang EnJun , Zou Tao , Zhang Shirley , Hall Maclean , Kambayashi Taku 
Published: January 2023

Abstract
Regulatory T cells (Tregs) are a subset of T cells that are important in maintaining immunological tolerance. After final development in the thymus, T cells are thought to quickly leave the thymus to populate peripheral secondary lymphoid organs. However, in contrast to conventional T cells, we hereby report that a large fraction of Tregs reside in the thymus for a prolonged period of time. Based on studies with RAG2.GFP reporter mice and bone marrow chimeras, we found that newly developed Tregs represent &lt;50% of the thymic Treg population in adult mice. The fraction of Tregs occupied by newly developed Tregs decreases further with age. We next tested whether the pool of mature Tregs (not newly developed) in the thymus was formed by recirculation of Tregs from the periphery and/or by Tregs that never leave the thymus. Adoptive transfer studies suggested that Tregs can recirculate back to the thymus. In addition, we found that Tregs are retained in the thymus for a prolonged period of time, as Tregs of donor origin were detected in donor thymi 6 weeks after thymic transplantation. Finally, using a combination of adoptive transfer and bone marrow transplantation, we estimate that compared to recirculation, thymic retention of Tregs contributes twice as much to the establishment of the mature Treg pool in the thymus. The thymic resident Treg population may affect the development of new Tregs in order to control antigen specificity or numbers of newly developing Tregs.
TCR signaling by conventional T cells is required for peripheral maintenance of Foxp3+ regulatory T cells (P1155)
Authors: Leichner Theresa , Satake Atsushi , Kambayashi Taku 
Published: January 2023

Abstract
Regulatory T cells (Tregs) control inappropriate conventional T cell (Tconv) activation. Treg maintenance is highly dependent on their IL-2-mediated proliferation in the periphery. Since Tconvs but not Tregs produce IL-2, IL-2 could serve as a gauge for Tregs to adjust their numbers accordingly to the size of the Tconv pool in the periphery. As the production of IL-2 by Tconvs is dependent on TCR signaling, we hypothesized that TCR signaling by Tconvs in response to self MHC class II (MHC-II) molecules is required for Treg maintenance. Indeed, when TCR signaling was specifically ablated in Tconvs but not in Tregs, an abrupt drop in Treg numbers was observed. Moreover, Treg numbers could not be maintained when T cells were adoptively transferred into MHC-II-deficient bone marrow chimeric mice, where MHC-II molecules were missing on radiosensitive antigen presenting cells. The drop in Treg numbers was associated with Tconv activation and the development of severe dermatitis, gut pathology, and weight loss. The decrease in Treg numbers and the onset of this disease was delayed by the injection of IL-2. These results suggest that IL-2 produced through TCR signaling by Tconvs in response to self MHC-II molecules is critical for the maintenance of Tregs and for subsequent prevention of a widespread inflammatory syndrome. This points to a potential role of positive selection in producing Tconvs bearing TCRs with adequate affinity to self MHC-II complexes in Treg maintenance.
Differential selection of Th1 effector and memory pools depends on the qualitative nature of TCR-driven CD4+ T cell activation (P1175)
Authors: Williams Matthew , Kim Chulwoo 
Published: January 2023

Abstract
Not all clones that participate in the effector Th1 response to intracellular viral or bacterial infection can give rise to memory T cells. In this study, we generated a mouse with a fixed TCRα chain of known specificity in order to characterize the evolution of effector and memory TCRβ repertoires following LCMV or Listeria infection by deep sequencing. The repertoire diversity of antigen-specific Th1 cells significantly decreased in the transition from effector to memory, and hierarchical cluster analysis revealed that the TCR repertoires of memory T cells, regardless of infectious model, were more similar to each other than to the effector Th1 populations from which they arose. We further generated a large panel of TCRs derived from our sequencing results that were cloned into retroviral expression vectors and used to generate TCR “retrogenic” bone marrow chimeras. Individual clonal T cell populations harvested from these mice were analyzed in adoptive transfer experiments to allow for their longitudinal analysis following infection. We found that the individual memory potential of each clone did not correspond to overall TCR avidity, as measured in tetramer equilibrium binding assays, but to the acquisition of high functional avidity and slow off-rates, as determined by tetramer decay. Our findings support a model in which differential selection of the Th1 effector and memory pools depends at least in part on the qualitative nature of TCR-driven CD4+ T cell activation.
The role of Foxp3 in augmented homeostatic proliferation of Treg (P1187)
Authors: Lin Yan , Camirand Geoffrey , Rothstein David 
Published: January 2023

Abstract
Natural CD4+Foxp3+ Tregs (nTreg) exhibit a high rate of constitutive homeostatic proliferation (HP) compared to CD4+Foxp3- conventional T (Tconv) cells (~50% vs. &lt;10% proliferate over 10d, respectively, as determined by transfer of CFSE-labeled CD4 subsets into replete recipients). nTreg from Filig mutant mice (~10% wt Foxp3 level), exhibited low HP (&lt;10%). Transducing Filig nTreg with Foxp3 retrovirus (rv), restored high HP (40%) vs. vector control (12% HP). However, when Foxp3 rv was expressed in Tconv cells, the resulting induced Tregs (iTregs) exhibited similar HP to Tconv cells transduced with vector control (17% vs. 16%, respectively). Thus, Foxp3 is necessary but not sufficient for high HP. Next, we examined the role of Foxp3 in HP of endogenously selected T cells (using EdU labeling). Filig bone marrow transduced with Foxp3 rv vs. control rv was transplanted into irradiated congenic hosts. Surprisingly, in this setting, Tconv cells transduced to constitutively express Foxp3 (iTregs), underwent high HP (32%) compared to control rv (or nontransduced) Foxp3- counterparts (12% HP). Thus, forced Foxp3 expression during thymic ontogeny augments HP capacity, not seen when Foxp3 is acquired by mature Tconv in the periphery. These findings provide new insight into the regulation of HP in Treg, the role of Foxp3 expression in Treg homeostasis, and highlight additional distinctions between induced and natural Treg.
CLEC16A: A novel C-type lectin domain-containing protein pertaining to systemic lupus erythematosus (P4092)
Authors: Tam Rachel CY , Yan Sheng , Lau Chak Sing , Chan Vera SF 
Published: January 2023

Abstract
Several independent genome wide association studies have shown associations of single nucleotide polymorphisms within the C-type lectin domain family 16 member A gene, CLEC16A, with various autoimmune diseases, including systemic lupus erythematosus (SLE). However, the expression and function of this novel C-type lectin domain-containing protein is still largely unknown. This study aims to identify the expressed forms of CLEC16A in peripheral mononuclear cells (PBMCs) and to relate its expression level with SLE disease activity. Two expressed isoforms, isoforms 1 and 2, of CLEC16A have been cloned and sequenced from PBMCs from healthy Chinese, each bearing differences in sequence when compared to the reference sequences derived from Caucasians. In isoform 1, exon 5 was consistently absent in Chinese PBMCs. However, in isoform 2, the stretch of 48-bp in-frame deletion within exon 11 in the reference sequence was found present in Chinese PBMCs. The mRNA expression levels of both isoforms were found significantly lower in SLE patients than in normal individuals. Yet, their expression levels did not correlate with disease activity as measured by SLE disease activity index. The molecular and cellular functions of CLEC16A are currently under investigation, which should shed light on the importance of this protein in SLE pathogenesis.