bluuebunny/crossref_metadata_2025_split huggingface dataset:
Pre-Ligand Assembly Domain (PLAD) is an important therapeutic target in inflammatory arthritis (131.21)
Authors: Deng Guo-Min , Lenardo Michael 
Published: January 2023

Abstract
Tumor necrosis factor (TNF)-α is an important proinflammatory cytokine and plays a crucial role in pathogenesis of inflammatory arthritis such as rheumatoid and septic arthritis. TNF-α exerts its effect by binding to the extracellular domain of TNF receptor (TNFR) 1 and 2. The amino-terminal portion of the TNFR extracellular domain, known as cysteine rich domain 1 (CRD1), or the pre-ligand binding assembly domain (PLAD), plays an important role in the TNFR signaling pathway. Our studies demonstrate that soluble TNFR1 and TNFR2 PLAD proteins could block the actions of TNF-α, the TNFR1 PLAD protein could significantly inhibit inflammatory arthritis such as arthritis induced by TNF-α, bacterial DNA, lipopolysaccharide and collagen. The TNFR1 PLAD protein could inhibit nuclear factor (NF)-κB activation and osteoclastogenesis triggered by TNF-α. The TNFR1 PLAD protein also could suppress expression of receptor activator of NF-κB (RANK), RANK ligand and TNFR1 in arthritic joints. To understand the mechanism of the PLAD protein inhibited inflammatory arthritis, we further studied how TNFR PLAD protein blocks the effect of TNF-α and inflammatory arthritis. Our data demonstrate that the TNFR PLAD may directly bind to TNFR and block relationship of TNF-α binding to TNFR. The TNFR1 PLAD protein could suppress the expression of matrix metalloproteinase and inducible nitric oxide synthase in collagen-induced arthritis. Our research findings indicate the TNFR1 PLAD protein may offer a new approach to inhibit the pathogenic effects of TNF-α, and may be a good prototype for a new generation of pharmaceuticals against TNFRs that could be useful in treatment of inflammatory arthritis.
Sphingosine 1 phosphate (S1P) -Type 1 Sphingosine 1-Phosphate G Protein-Coupled Receptor (S1P1) axis promotes peripheral generation of CD4+CD25+ regulatory T cells (131.5)
Authors: Wang Wengang , Goetzl Edward J 
Published: January 2023

Abstract
Regulatory T cells (Tregs) are a CD4+CD25+ subset of T cells that control normal immunity and autoimmunity by suppressing diverse functions of other immune cells. S1P regulates many immune functions via S1P1 and occasionally through another of its five receptors. The generation of Tregs from CD4+CD25− T cells in non-thymic lymphoid organs may be evoked by T cell activation and the actions of certain immune regulators, such as TGF-beta and IL-4. Using flow cytometry, we found there were1.4–1.8 fold more Tregs in CD4+ T cells of spleen, lymph node, and blood from S1P1 TG mice than in wild-type (WT) mice. In contrast, the numbers of Tregs in the CD4+ T cells from thymus of WT and S1P1 TG mice were the same. Cell proliferation assays using CD4+CD25− effector T cells as targets indicated that overexpressed Tregs in S1P1 TG mice had similar suppressive effects per T cell as Tregs in WT mice. In vitro assays showed around 5% of isolated CD4+CD25− T cells from S1P1 TG mice were converted to CD4+CD25+ Tregs by TCR stimulation or antigen activation in the presence of 10−7 M S1P. Real time PCR and flow cytometry assay showed the induction of FoxP3 mRNA and protein expression in these peripherally generated Tregs. Increased peripheral generation of Tregs by the S1P-S1P1 axis may alter the level of immune and autoimmune reactions sufficiently to make this system a therapeutic target for immunomodulatory drugs.
Tolerogenic Vaccination Using IL-12 Gene-Silenced Dendritic Cells for Autoimmune Arthritis (131.9)
Authors: Min Wei-Ping , Zheng Xiufen , Popov Igor , Li Mu , Zhang Xusheng , Sun Hongtao , Vladau Costin , Suzuki Motohiko , Inman Robert D , Garcia Bertha 
Published: January 2023

Abstract
We have recently demonstrated that dendritic cell (DC)-mediated immune modulation and deviation can be accomplished through RNA interference (RNAi), highlighting the therapeutic potential of RNAi-modified DC as antigen-specific tolerogenic vaccines. To date, an RNAi-based vaccine has not been reported. The current study was designed to develop siRNA-modified DC as antigen-specific, tolerogenic vaccines for prevention and intervention of autoimmune arthritis. Using small interfering RNA (siRNA) that specifically targets IL-12p35 gene (IL-12 siRNA), we have generated a type of DC that exhibits multiple tolerogenic characteristics. Immunization with type II collagen (CII)-pulsed and IL-12 gene-silenced DC (CII-pulsed/gene-silenced DC) resulted in antigen-specific nonresponsiveness in T cell responses. Vaccination with CII-pulsed/gene-silenced DC prevented collagen-induced arthritis (CIA) onset in a murine rheumatoid arthritis model. Furthermore, administration of CII-pulsed/gene-silenced DC was sufficient to inhibit progression of CIA. The therapeutic effects were further evidenced by decreased clinical scores, inhibited inflammatory infiltrates, and suppressed T cell and B cell responses to CII. In conclusion, this study is the first to demonstrate the therapeutic utilization of RNAi-modified DC as antigen-specific tolerogenic vaccines for autoimmune arthritis.
Simultaneous Detection and Quantitation of Multiple Antigen Expression in Tissue by Fluorescence Microscopy and Hyper-spectral Imaging (132.4)
Authors: Carpenter Anne , Chatigny Julie , LeFever Mark , Ashworth-Sharpe Julia , Haubein Ned , Vanpatten Kristie , Kosmeder Jerry , Lett Scott , Grogan Thomas , Bieniarz Chris , Miller Philip , Pestano Gary 
Published: January 2023

Abstract
Analyses of biomarkers in tissues represent a critical component in the development of therapeutics. In drug development, entire signaling pathways are profiled with a variety of high-throughput techniques. However, in the field of histology, tissue-based diagnostics are generally dominated by single- or dual-analyte tests. In addition, manual methods for slide processing promote inconsistency. We have developed automated instrumentation for conducting high-throughput IHC and ISH in widely available formalin-fixed paraffin-embedded samples. In this report we describe the utilization of Quantum dots (Qdots®) in advanced antigen detection in intact tissues. Specifically, we developed a novel set of hapten and anti-hapten monoclonal antibodies for use in multi-analyte assays. Spectral imaging software was utilized to deconvolute images generated by fluorescence microscopy on tissue samples. In model systems, we demonstrate simultaneous detection, and quantitation of expression levels and cell numbers, for markers including CD20, CD34, CD45, IgGκ, IgGλ, and Ki67. These reagents provide a growing set of tools that could complement early marker validation. The examples in this study demonstrate the utility of coupling high-throughput, automated histochemistry with quantifiable, multiplexed analyte detection. These findings are reported for individual cells in their anatomic context.
KIR/HLA Compound Genotypes Determine Early Antiviral Response of Natural Killer Cells (35.17)
Authors: Ahlenstiel Golo , Martin Maureen P , Gao Xiaojiang , Carrington Mary , Rehermann Barbara 
Published: January 2023

Abstract
A growing number of genetic epidemiological studies have suggested an influence of the extraordinarily polymorphic KIR/HLA compound genotypes in determining outcome to viral infections, but functional data to explain these genetic data have not been reported to date. Using an in vitro influenza A virus infection model, we here attribute functional differences in human NK cell activity to distinct KIR/HLA genotypes. Using cytokine EIAs and multicolor flow cytometry, we show that NK cells from KIR2DL3+/KIR2DL1+ and HLA-C1 homozygous subjects display a faster response kinetics and release more IFN-γ and TNF-α faster to influenza A virus infection than NK cells from KIR2DL3+/KIR2DL1+ and HLA-C2 homozygous subjects. Subanalysis revealed that the HLA-C-regulated, i.e. KIR2DL3+ NK cell subpopulation of HLA-C1 homozygous subjects responded more rapidly than their HLA-C-regulated, i.e. KIR2DL1+ counterpart in HLA-C2 homozygous subjects. This differential functional response was independent of Bw4/KIR3DL1 interactions. It was also not due to intrinsic difference between NK cell of HLA-C1 and HLA-C2 homozygous subjects, as demonstrated by equal cytotoxicity in the absence of HLA-C. In summary, these results provide first functional evidence for the epidemiological associations between specific KIR/HLA compound genotypes and protection against viral infection observed at the population level.
IFN-γ up-regulates expression of the MHC class II transactivator (CIITA) type IV promoter in multiple myeloma cells (35.20)
Authors: Piskurich Janet F. , Zhao Mojun , Flynt Frederick L. , Hong Mei , Chen Han , Gilbert Carolyn A , Briley Nicole T 
Published: January 2023

Abstract
MHC Class II transactivator (CIITA) is the master transcriptional regulator of MHC class II (MHC II) genes. MHC II expression is central to antigen presentation and the immune recognition of tumor cells. The human CIITA gene, MHC2TA, is regulated in a cell-type specific manner by multiple promoters. Regulation by IFN-γ is generally via the type IV promoter (pIV) and involves the binding of IFN regulatory factor (IRF)-1 and IRF-2. The response of pIV to IFN-γ was examined in myeloma cells. Western blot and RT-PCR show that STAT1 is activated by phosphorylation and IRF-1 RNA expression increases in response to IFN-γ. RT-PCR and functional promoter analyses show that IFN-γ up-regulates the activity of MHC2TA pIV as does ectopic expression of IRF-1 or IRF-2, and the IRF-E site is required. Binding studies confirm IRF-1 and IRF-2 binding to MHC2TA pIV. Myeloma cells express positive regulatory domain I-binding factor 1 (PRDI-BF1), also called B lymphocyte-induced maturation protein-1 (Blimp-1), which represses both MHC2TA pIII and pIV. Importantly, these cells retain the ability to up-regulate CIITA pIV and MHC II in response to IFN-γ. Although PRDI-BF1/Blimp-1 diminishes the ability of myeloma cells to present antigen by limiting CIITA and MHC II expression, these studies indicate that expression can be restored by the use of cytokines like IFN-γ.
Research support: RO1CA10220, RO1CA114504, MEDCEN Foundation of Central GA
The role of the 19S ATPase S6a in the transcriptional regulation of major histocompatibility class II (MHC II) genes (35.26)
Authors: Truax Agnieszka Dorota , Greer Susanna F 
Published: January 2023

Abstract
MHC II molecules are glycoproteins that present intracellular antigens to CD4+ T cells and play an important role in induction and regulation of adaptive immune responses. MHC II molecules are regulated at the level of transcription by a master regulator, the class II transcriptional activator, CIITA, whose association with the MHC promoter is necessary for initiation of transcription. It is well established that one mechanism of regulating transcription is through degradation of factors by the 26S proteasome. The proteasome is composed of a 19S regulatory particle that recognizes ubiquitinated proteins and a 20S proteolytic core where the tagged proteins are degraded. Previous studies in yeast have demonstrated that the S6a ATPase of the 19S associates with actively transcribed genes, suggesting a non-degradative role for the ubiquitin-proteasome machinery. To further understand these roles in mammalian cells, we have investigated the role of S6a in regulating CIITA mediated MHC II transcription. Our research indicates that S6a is recruited to actively transcribing MHC II and CIITA genes and that RNAi mediated S6a knockdown correlates with decreased expression of MHC II and CIITA. Our study demonstrates a novel mechanism of regulating MHC II expression and will enhance our understanding of mammalian transcription.
Research supported by the NMSS, the Georgia Cancer Coalition and GSU.
Genomic analysis of RAE-1 genes (35.30)
Authors: OGASAWARA KOUETSU , Ishizaki Kazusa , Fujiwara Naruyoshi , Sasazuki Takehiko 
Published: January 2023

Abstract
NKG2D is an activating receptor on NK cells and activated CD8 T cells that has been implicated in tumor immunity, viral immunity and autoimmunity. The ligands of NKG2D are tightly regulated stress-inducible molecules, whose expression is triggered by transformation or infection. In mice, these NKG2D ligands include the retinoic acid early inducible-1 (RAE-1) proteins. However, the mechanisms of transcriptional regulation in RAE-1 genes are poorly understood. We examined what transcription factors regulate RAE-1 expression. We found that RAE-1 delta has six exons and RAE-1 epsilon has seven exons. We isolated genomic DNAs of upstream from exon one which are thought of RAE-1 promoters in RAE-1 delta or RAE-1 epsilon. We found that consensus sequences of several transcription factors are present in RAE promoters. Therefore, these transcription factors may regulate RAE-1 expressions.
This work is supported by Grants-in Aid for Science Research from the Ministries of Education, Culture, Sports, Science, and Technology of Japan, Grants-in Aid for Science Research from the Ministriesof Health, Labor, and Welfare of Japan, Human Frontier Science Program CDA, The Mitsubishi Foundation, The Sumitomo Foundation, Takeda Science Foundation and Uehara Memorial Foundation.
Loading and phosphorylation of RNA polymerase II influence production of secretory-specific Ig heavy chain mRNA in plasma cells. (35.33)
Authors: Milcarek Christine , Shell Scott 
Published: January 2023

Abstract
A crucial step in the development of B cells into plasma cells is the increased production of secreted immunoglobulin which occurs by differential selection of the secretory-specific polyA site. Factors involved in polyadenylation can be coupled with transcription through phosphorylation of the carboxyl-terminal domain (CTD) of RNA pol II. We analyzed loading and phosphorylation patterns of RNA Polymerase II along an identical immunoglobulin heavy chain gene expressed either in B- or plasma cells by chromatin immunoprecipitation using various antibodies. Data indicate that the switch to secreted immunoglobulin heavy chain mRNA production is influenced by increased RNA Polymerase II accumulation and CTD phosphorylation at the promoter in plasma cells. Loading of transcription factors is increased in the plasma cells. This allows for increased access of specific processing factors to the polymerase and nascent RNA.
Supported by NIH-CA-86433 to C.M.
FOXP3 COOPERATES WITH LINKER HISTONE H1b TO REPRESS IL-2 (35.46)
Authors: Mackey-Cushman Stephanie , Su Lishan 
Published: January 2023

Abstract
Regulatory T cells (Treg) are a subset of T cells indispensable for regulating self tolerance by actively suppressing autoreactive T cells. Treg cells are characterized as CD4+CD25hi, which preferentially express the transcription factor FoxP3. FoxP3 is required for Treg lineage development. Ectopic expression of FoxP3 converts CD4+CD25− T cells to Treg cells with suppressed IL2 expression, and mutations within the FoxP3 gene lead to autoimmune diseases in humans and mice. In order to understand the molecular mechanism by which FoxP3 functions in Tregs, we performed a large scale co-immunoprecipitation of FoxP3 followed by mass spectrometry identification of interacting proteins. We identified a physical interaction between FoxP3 and H1b, a specific isoform of histone H1. The leucine zipper region of FoxP3 is required for the interaction with H1b. This interaction is specific to H1b, while other H1 isoforms, H1a and H1e, do not interact with FoxP3. Furthermore, ectopic expression of FoxP3 and H1b in Jurkat T cells synergistically represses IL-2 expression following activation. These findings support a role for H1b in FoxP3-mediated repression of IL-2 expression in Treg cells.
Regulation of the Perforin Locus by IL-2 During CD8+ T-cell Differentiation (35.49)
Authors: Pipkin Matthew Eugene , Cruz-Guilloty Fernando , Lichtenheld Mathias Georg , Rao Anjana 
Published: January 2023

Abstract
The molecular basis of memory and effector CD8+ T cell differentiation is not well understood. Here, we show that the strength of IL-2 signaling determines the transcriptional profile of primary mouse cytotoxic T lymphocytes (CTL) during in vitro differentiation. CD8+ T cells were activated and cultured in either high or low dose IL-2 for 1 week; both groups proliferated equivalently and expressed similar amounts of IFNγ upon restimulation. However, cells cultured in high IL-2 maintained phosphorylated Stat5 throughout differentiation, induced the transcription factor Eomesodermin, expressed robust amounts of perforin and granzyme B and displayed a surface phenotype characteristic of effector cells. In contrast, cells cultured in low IL-2 expressed low amounts of granzyme B, reverted to a surface phenotype characteristic of memory cells and gradually silenced expression of perforin mRNA. DNase I hypersensitivity mapping showed no major differences in chromatin structure in &gt;200 kb surrounding the perforin gene, whereas chromatin immunoprecipitation assays showed that cells cultured in high IL-2 displayed increased density of RNA polymerase II at the perforin promoter. Based on these studies in vitro, we propose that the strength of signaling through the IL-2 receptor regulates commitment to the memory cell lineage in vivo.
Single Nucleotide Variations Between the I gamma 1 and I gamma 4 Promoters Result in Biased Recruitment of Transcription Factors in Response to CD40 Signaling (35.7)
Authors: Sinquett Frank L , Dryer Rebecca L , Covey Lori R 
Published: January 2023

Abstract
IgG subclass expression is highly biased in human IgM+ B cells activated with CD40L and IL-4. This bias is clearly reflected at the level of germline (Iγ) transcription despite extensive homology throughout the four Iγ promoters. Our current aim is to define the underlying mechanism by which Iγ1 transcripts are preferentially expressed relative to Iγ4 transcripts in order to extend our understanding of CD40 mediated humoral responses. Characterization of both promoters using defined regions of the Iγ1 and Iγ4 promoters revealed a marked decrease in Iγ4 transcriptional activity when compared to Iγ1 activity despite similar regulation. Expression studies have further shown that exchanging a previously defined 36 bp region of the Iγ1 promoter that is essential for Iγ1 transcription and binds NF-κB and various co-activators, can elevate Iγ4 transcription. This increase can be attributed to single nucleotide changes in either a putative CREB site or a putative NF-κB site within this region that renders this element CD40 inducible, and increases recruitment of p50, p65, c-Rel, CREB, and p300. Together, these data indicate that a single base pair change in a key regulatory region can contribute to differences in promoter strength. Competition assays and site-directed mutagenesis are currently in progress to further our understanding of the effect of these variances on Iγ transcription targeting.
Supported by AI37081
Role of aryl hydrocarbon receptor activation in antigen-presenting cell functions that result in diminished clonal expansion and differentiation of influenza virus-specific CD8+ T cells (36.27)
Authors: Jin Guangbi , Neumiller Joshua J. , Lawrence B. Paige 
Published: January 2023

Abstract
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is expressed in cells of the immune system. The AhR binds many different types of exogenous compounds, including pollutants and phytochemicals. The pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) is the most potent AhR agonist known, and is a well known immune suppressant. However, the precise molecular mechanism remains unclear. We have previously shown that AhR activation by dioxin suppresses the proliferation and differentiation of influenza virus-specific CD8+ T cells by disrupting events within bone marrow-derived cells important for the activation of naïve CD8+ T cells. However, TCDD does not affect CD8+ T cell directly, suggesting that defects in other cells essential for activating CD8+ T cells are affected by AhR activation. In this study, we show that the suppression of influenza virus-specific CD8+ T cell responses by TCDD appears to be independent of CD4+ cells, suggesting that deregulated APC function underlies TCDD-mediated decreasing the clonal expansion of virus-specific CD8+ T cells. To test this, we characterized the phenotype of DC present within lung, and examined their migration and function in secondary lymphoid tissue.
Funded by NIEHS Grants R01ES10619 and K02-ES012409 to BPL.
Fetal/neonatal ethanol exposure irreversibly alters immune parameters in young mice (36.28)
Authors: Fan Ji , Young Betty M , Cook Robert T , Edsen Michelle R , Legge Kevin L , Waldschmidt Thomas J , Tygrett Lorraine T , Schlueter Annette J 
Published: January 2023

Abstract
Chronic ethanol ingestion has been shown to impair immune defensive function of adult hosts to invading organisms. Children born to alcoholic mothers are known to have increased incidence and severity of infections, but EtOHÕs effect on the immune system in the fetus/neonate has not been well studied. We have developed a model of fetal/neonatal EtOH exposure in which female mice are maintained on 10% EtOH in the drinking water during breeding and gestation, and then on 12% EtOH during lactation. Pups have successfully been weaned and showed no significant difference in weights from controls through 42 days of age. These pups have been studied at 20 d (weaning) and 42 d (adolescence) for immunological cell numbers and function. Our data showed decreased splenic T and B cells in EtOH pups, but increased pulmonary leukocyte and dendritic cell (DC) numbers. Splenic DC and epidermal Langerhans cells numbers did not change. However, EtOH decreased baseline costimulatory molecule expression on splenic DC and the ability of these DC to stimulate allogeneic T cells, changes that persisted following cessation of EtOH exposure. These results indicate that mice born to EtOH-consuming mothers have immune deficiencies that are persistent following cessation of EtOH exposure. The dissection of T, B cell and DC dysfunction in EtOH pups is in progress.
Apoptosis-induced inhibition of CD1d-mediated antigen presentation: Different roles for caspases and p38 MAPK (36.4)
Authors: Khan Masood Alam , Sriram Venkataraman , Gourpura Renukaradhya J , Nagala Manjula , Brutkiewicz Randy R 
Published: January 2023

Abstract
The cross presentation of antigens derived from apoptotic cells enhances MHC class I or II-mediated antigen presentation. In the current study, we show that presentation of lipid antigens by CD1d molecules is inhibited by the induction of apoptosis in a p38 MAPK- or caspase-dependent manner. Treatment of murine CD1d1–expressing fibroblasts with PPMP or camptothecin (both of which can induce apoptosis) resulted in the activation of p38, and a p38-specific inhibitor (but not ERK1/2 pathway inhibitor) rescued CD1d-mediated antigen presentation in PPMP- or camptothecin-treated cells. The pan-caspase inhibitor Z-VAD-fmk rescued antigen presentation by CD1d in camptothecin (but not PPMP)-treated cells. Confocal microscopic analysis showed an altered intracellular distribution of CD1d in PPMP- or camptothecin-treated cells, which was corrected by the addition of p38 or caspase inhibitors. These results suggest that the induction of apoptosis compromises with CD1d-mediated antigen presentation.
This work was supported by NIH grant PO1 AI056097 to RRB.