bluuebunny/crossref_metadata_2025_split huggingface dataset:
CD8+ T-cells are required for the therapeutic effects of glatiramer acetate (Copaxone) on autoimmune demyelination (131.7)
Authors: Mendoza Jason P , York Nathan R. , Benagh Andrew , Karandikar Nitin J. 
Published: January 2023

Abstract
Murine experimental autoimmune encephalomyelitis (EAE) is an induced, autoimmune, demyelinating disease model for human multiple sclerosis (MS). Copaxone is an approved immunomodulatory therapy for MS that was discovered through studies in EAE. It is thought that the main mode of action of Copaxone is the induction of a Th1 to Th2 shift in CD4+ T-cells. We have shown that, in human MS, Copaxone therapy induces an upregulation of cytotoxic, suppressor CD8+ T-cell responses, suggesting that these cells are integrally involved in mediating the immune effects of this drug. In this study, we show that Copaxone induces robust CD8+ T-cell responses even in mice. Whereas Copaxone immunization ameliorates the clinical and histologic severity of EAE in wild type C57/BL6 mice, such protection is not observed in CD8(−/−) mice. Moreover, adoptive transfer of CD8+ T-cells from Copaxone-immunized (but not OVA-immunized) mice results in amelioration of EAE in CD8(−/−) mice, suggesting therapeutic potential for Copaxone-specific CD8+ T-cells. In conclusion, our studies provide strong evidence for the in vivo relevance of CD8+ T-cell responses during Copaxone therapy and support the hypothesis that the induction of immune modulatory CD8+ T-cells is an essential step in mediating therapeutic protection during autoimmune demyelination.
This research was supported, in part, by grants from the National Institutes of Health and the National MS Society.
Role of ATM in V(D)J recombination and cell cycle control (35.10)
Authors: Dujka Melanie E , Puebla-Osorio Nahum , Zhu Chengming 
Published: January 2023

Abstract
V(D)J recombination is a process specific to developing B- and T- lymphocytes in which double strand breaks (DSBs) are introduced into germline DNA and upon repair generate rearranged gene segments. This recombination event is cell cycle regulated and is confined to the noncycling G0/G1 phase of cell cycle. The protein kinase Ataxia Telangiectasia Mutated (ATM) is a vital mediator in both the DNA damage response and cell cycle. ATM was found localized at V(D)J mediated break sites, suggesting an additional function for ATM in recombination. In the absence of ATM, mice develop thymic lymphomas with chromosomal translocations. Therefore, we hypothesize a role for ATM in the cell cycle control of V(D)J recombination. Thymic lymphomas from Atm null mice revealed cells harbored multiple chromosomal translocations near sites of recombination. Wildtype and Atm null thymocytes were separated into cell cycle fractions of G0/G1 and S/G2/M. V(D)J mediated DSBs, indicative of recombination, were observed at the T cell receptor α, δ, and β loci in both cell cycle fractions of Atm null thymocytes but were found only in the G0/G1 fraction of wildtype thymocytes. Together, these results indicate the involvement of ATM in the cell cycle control of V(D)J recombination. Loss of this control can lead to sustained DNA DSBs and translocations, which could contribute to tumorigenesis.
Analysis of transcriptional regulation of mouse FoxP3 gene expression in CD4+ CD25+ regulatory T cells (35.21)
Authors: FURUUCHI KEIJI , TONE YUKIKO , KOJIMA YOSHITSUGU , TYKOCINSKI MARK L , GREENE MARK I , TONE MASAHIDE 
Published: January 2023

Abstract
FoxP3 is expressed at high level in CD4+CD25+ regulatory T cells (Treg) and functions as a key transcriptional factor that determines cell phenotype. Despite of the inherent interest, little is known concerning process of transcriptional regulation of FoxP3 due to lack of experimentally feasible to analyze promoter activity using transfection-based methods in natural Treg cells. We have, therefore, identified a model cell line that expresses FoxP3 at high levels upon stimulation. To identify the promoter region of the mouse FoxP3 gene, we determined the transcription initiation site by 5′RACE, using RNAs from CD4+CD25+ Treg cells and the cell line. The major start site mapped to the same position in the both type of cells, indicating that this cell line can be used as a model system. Luciferase reporter assays were performed using a 1.7kb 5′ franking fragment of the FoxpP3 gene as promoter. Although the cell line expressed FoxP3 mRNA at high levels, we could not detect any promoter activity in this region, suggesting that enhancers might be essential for this promoter to function. Indeed, we elicited that FoxP3 promoter activity by coupling it to the SV40 enhancer. We have now identified an enhancer region in the vicinity of the FoxP3 gene, that upon analysis have revealed important and unexpected insights into the transcriptional control of the FoxP3 gene.
Negative regulation of RORgt by direct binding to Egr3 (35.41)
Authors: Kersh Gilbert J , Xi Hongkang 
Published: January 2023

Abstract
The orphan hormone receptor RORgt is a transcription factor that is expressed in lymphoid tissue inducer cells, double positive thymocytes, and Th17 cells. Analysis of RORg-deficient mice has revealed that this factor is critical for normal function of all three of these cell types. It is therefore of interest to understand how RORgt function is regulated. We have found that RORgt function can be negatively regulated by Egr3, a zinc finger transcription factor that is induced by a variety of stimuli, including TCR signaling. Ectopic expression of both RORgt and Egr3 in a thymocyte cell line prevents RORgt from binding to target gene promoters, and blocks target gene expression. We have employed multiple strategies to demonstrate that the inhibition of RORgt function is due to direct binding of RORgt by Egr3. Immunoprecipitation of either Egr3 or RORgt brings down the other protein, even when both proteins are obtained by in vitro translation. A mammalian two-hybrid system also indicates that the two proteins physically interact. Mutational analysis of RORgt indicates that the domain responsible for steroid receptor coactivator recruitment is required for binding to Egr3. These studies demonstrate a novel function for Egr3, and suggest a mechanism for regulation of thymocyte and Th17 cell differentiation.
This work is funded by a grant from the American Cancer Society.
Identification of RNA binding proteins involved in norepinephrine mediated regulation of Thy1 mRNA (35.45)
Authors: Koduvayur Sujatha P. , LaJaevic Melissa Dawn , Graf Lloyd H , Cohen Rhonna L , Chambers Donald A 
Published: January 2023

Abstract
Our laboratory has shown that norepinephrine (NE) increases Thy1 mRNA decay in S49 T lymphoma cells through a ß 2 adrenergic receptor/cAMP/protein kinase A pathway. Thy1 message, like many cytokine messages, contains an adenine and uridine rich element (ARE) in its 3′untranslated region (3′UTR) that has been associated with mRNA destabilization. In this study we identify RNA binding proteins from S49 wild type (wt) and a kinase deficient mutant (kin−) and determine if NE (150uM, 30 min) treatment of the cells alters binding. Photo affinity binding assay with cytoplasmic extracts from both cell types and 32P labeled, 170 nt long Thy1 ARE probe revealed the binding of twelve proteins of 30–105 kDa. Of these AUF1, HuR, TIAR, KSRP, and Hsc70 were identified by crosslinking followed by immunoprecipitation (IP). Crosslink IP also identified protein –protein interactions between TIAR and Hsc70. A pull down assay using a biotinylated RNA probe identified PAI-1binding protein and Hsp70, in addition to the others mentioned above. NE treatment of wt and kin− cells did not change the binding profile or affect the apparent ability of these proteins to bind. These results suggest that although the identified proteins may form a part of the mRNA decay machinery, modifications of these proteins or other factors may be involved in the NE-mediated regulation of Thy1 mRNA stability.
Supported by NIH.
Are our immunoglobulin V genes pre-adapted for recognizing pathogens: a 3-dimensional analysis (35.53)
Authors: Thomson Christy , Bryson Steve , McLean Gary , Pai Emil , Schrader John 
Published: January 2023

Abstract
There is strong phylogenetic evidence that the immunoglobulin V genes that make a major contribution to the generation of diversity of our primary repertoire, are under selective pressure, perhaps by pathogens themselves. Here we report the X-ray crystallographic structural analysis and the consequent kinetic and thermodynamic characteristics of a representative of a subclass of human neutralizing antibodies that originate from a particular pair of Vh and Vk genes and that target a short linear epitope on a human pathogen, HCMV. By comparison of its characteristics with its putative germline ancestors, these studies demonstrate that conserved germline Vh and Vk genes encode key residues in CDR-1 and CDR-2 that are pre-adapted to provide the foundation for the generation of a high-affinity hypermutated antibody. They show that although somatic mutation results in improvements in complimentarity of the binding site, the antibody combining site of the neutralizing antibody does not conform to a lock and key model and retains considerable mobility that is stabilized by interaction with antigen. We speculate that this flexibility may enhance its effectiveness at neutralization of infectivity.
The Effect of Vitamin C on the phenotype and function of Dendritic cells during activation with LPS (36.24)
Authors: Jeong Young Joo , Woo Ah Mi , Maeng Hyung Gun , Kang Jae Seung , Shin Dong-Hoon , Lee Wang Jae , Hwang Young-il 
Published: January 2023

Abstract
Activating DCs produce reactive oxygen species (ROS), which are necessary during antigen processing. In this study, we hypothesized that vitamin C (VC), as an antioxidant, may affect phenotype and function of DCs following activation. Bone marrow cells isolated from mice were cultured and differentiated into DCs in the presence of GM-CSF for 7 days. DCs were pre-treated with various concentrations of VC, and activated with LPS for 24 hrs. The supernatants were collected for measurements of cytokine secretion, and the changes of several markers of DC activation were analyzed. VC in the culture media reduced intracellular ROS levels, suggesting anti-oxidant effects on DCs. The expression of CD40 and CD80 were not influenced, but those of CD86 and MHC class II were slightly reduced. The reduction of phagocytic functions of DCs with activation was suppressed by the presence of VC, dose-dependently. In the supernatants, dose-dependent reduction of IL-6 and increased secretion of IL-12p70 and IL-10 were observed with increasing concentration of VC. IFN-¥ã was not detected at all. Because VC affected some of surface marker expression and phagocytic functions of activated DCs and, most noticeably, changed cytokine secretion profiles, we also investigated the results of T cell activation by those DCs treated with VC, with respect to T cell proliferation and cytokine secretion by activated T cells.
SIRNA KNOCKDOWN OF SPHK1 PROTECTS MICE FROM IGE-MEDIATED ANAPHYLAXIS (37.6)
Authors: Melendez Alirio J. , Jayapal Manikandan , Pushparaj Peter Natesan 
Published: January 2023

Abstract
Sphingosine kinases are emerging a key enzymes in the intracellular signaling cascades, that mediate inflammatory responses, triggered by various inflammatory stimuli including: anaphylatoxins (C5a); bacterial-derived peptides (fMLP); and IgG or IgE receptor stimulation (FcγRI and FcεRI), in various immune cells, including in human mast cells. Moreover, we have recently reported a study on genome-wide gene expression profile of human mast cells sensitized with IgE alone, or stimulated by FcεRI aggregation, and found that sphingosine kinase 1 (SphK1) is one of the early genes to be activated. This, coupled to our previous studies indicating that SphK1 is pivotal in the signaling pathways initiated by FcεRI in human mast cells, prompted us to investigate the functional role of SphK1 in IgE-triggered inflammation/anaphylaxis in vivo. The systemic anaphylaxis reaction in mice has been considered to be a typical immediate hypersensitivity response determined by the activation of immune cells via antigen-induced aggregation of an IgE-sensitized FcεRI. To this end, we silenced the SPHK1 gene in vivo (using a specific siRNA), and investigated whether SphK1 knockdown influenced the outcome of the acute inflammatory responses, triggered during IgE-mediated systemic anaphylaxis in mice. We show here that, the siRNA for SphK1 protected mice from the IgE-mediated allergic reactions, including inhibition on: temperature changes; vascular permeability; immune-cell infiltration into organs; cytokine production; and vascular cell adhesion molecule expression. Thus, our findings strongly support a role for SpkK1 IgE-mediated anaphylaxis in vivo.
Thymic stromal lymphopoietin (TSLP) is a potent activator of human mast cells (38.4)
Authors: Jessup Heidi K. , Allakhverdi Zoulfia , Comeau Michael R. , Park-Yoon Bo-Rin , Ziegler Steven F. , Delespesse Guy 
Published: January 2023

Abstract
Compelling evidence suggests that the epithelial cell-derived cytokine thymic stromal lymphopoietin (TSLP) may initiate asthma or atopic dermatitis through a dendritic cell-mediated Th2 response. Here we describe how TSLP might initiate and aggravate allergic inflammation via the innate immune system. We show that TSLP stimulates the production of Th2 cytokines by progenitor-derived human mast cells (MC). MC expressed each chain of the TSLP receptor complex, i.e. the TSLP-binding chain (TSLP-R) and the IL-7R-α chain at the mRNA and protein levels. TSLP-R was also detected on MC infiltrating the bronchial mucosa of asthmatic patients as revealed by staining biopsy specimen with TSLP-R specific mAb. TSLP dose-dependently induced the release of IL-5 which was detectable after 6 h of culture and reached plateau at 24 h. The response of MC was specifically suppressed by neutralizing mAb to TSLP or TSLP-R. TSLP did not induce MC degranulation or the release of lipid mediators. In contrast, high levels of IL-5, IL-13, IL-6 and GM-CSF were released after 24 h of MC stimulation. TSLP exerted an opposing effect on the production of anti-inflammatory cytokines, i.e. it suppressed TGF-β and enhanced IL-10. Among several chemokines examined, TSLP enhanced the production of only IL-8 and I-309. Finally, we show that primary airway and skin epithelial cells release functionally sufficient levels of TSLP to stimulate human MC.
Studies of IgE receptor (FceRI) processing in human basophils (HB) (38.7)
Authors: MacGlashan Donald 
Published: January 2023

Abstract
Studies HB in the 1980s suggested that FceRI was not down-regulated following aggregation of the receptor. We now show that receptor, after strong stimulation, is lost from the surface and degraded, but with a very slow time course; the T1/2 for loss from the surface or degradation was 12 hours and 24 hours, respectively. FceRI degradation is inhibited with Bafilomycin A suggesting a lysosomal route of loss. Under strong stimulation there was modest loss of un-involved receptors, 13±5% loss @ 18 hrs. Loss of receptor was only partially sensitive to inhibition with a src-family kinase inhibitor, PP1, and insensitive to presence of a third generation syk inhibitor, NVP-QAB205. Previous studies have noted that there is both immature receptor (p46) and mature receptor (p60) expressed and in the current studies we noted that p46 receptor expression increased following stimulation with both IgE-dependent and IgE-independent stimuli. Resting levels were inhibited by cycloheximide, actinomycin D, MG-132, PP1 and NVP-QAB205 and enhanced with Bafilomycin A. However, these studies found that the lysosomal processing was sensitive to apoptosis and suggested that these changes were not due to changes in synthesis of p46 but to changes in degradation rate. Studies of unoccupied receptor loss also indicated endocytosis and lysosomal degradation (and some recycling), i.e., both forms of FceRI loss depend on lysosomal processing.
Supported by NIH-AI20253
Perinatal Exposure to Environmental Tobacco Smoke and Immunological Consequences in Primate Infants (40.11)
Authors: Hood Carol , Peak Janice , Spinner Abby , Datta Seema , Fiehn Oliver , Pinkerton Kent 
Published: January 2023

Abstract
Mechanisms of environment tobacco smoke (ETS) to adverse health in children are unknown. To understand how ETS may alter the developing immune system of nonhuman primates, rhesus macaque infants were exposed to ETS during perinatal life at 1mg/m3 total suspended particulates for 6 hours per day, 5 days per week. Four treatment groups were involved:
ETS from gestation day 40 to 3 months,ETS exposure from gestation day 40 to 13 months of age,ETS exposure starting 6 months post natal until 13 months of age,sham-control / filtered air throughout gestation and postnatal age to 13 months.
At three months of age, all ETS infants demonstrated a significant reduction in plasma protein (P = .03). Nicotine levels were three times higher in younger infants (3 months) than infants at 13 months of age (P = .002 Fisher) suggesting age-based differences in metabolism. Morphometric methods were used to examine the spleen. There was a significant increase in the relative proportion and volume of the white pulp found in the spleen following exposure to ETS at 3 months of age as well as in animals with exposure to ETS beginning at 6 months of age, compared to their control groups (P=&lt;.0001, P =.02, P = .01, P=.04 respectively) . These findings suggest that there are critical windows of susceptibility in infants to ETS. Changes in the proportion and volume of the white pulp of the spleen by ETS may play a key role to alter the immune status of the neonate.
Differential regulation of mucosal immunity by Bacillus anthracis edema toxin and cholera toxin as adjuvants for transcutaneous vaccines (41.3)
Authors: Duverger Alexandra , Tang Wei-Jen , Leppla Stephen H , Boyaka Prosper N 
Published: January 2023

Abstract
Our previous studies have shown that nasal co-administration of an anthrax edema toxin (EdTx) derivative [i.e., protective antigen (PA) plus an edema factor (EF) mutant (EF N447S)], enhances mucosal and systemic immunity to both PA and co-administered unrelated protein antigen. Unlike the ganglioside targeting adjuvant, cholera toxin (CT), nasal EdTx N447S did not target CNS tissues or induce inflammatory responses in the NALT. Here, we examined whether the difference in the cell targeting of these two adjuvants would lead to different mucosal and systemic Ab responses when used in transcutaneous vaccines. We found that both CT and EdTx N447S promoted IgG Ab responses in the serum. However, transcutaneous application of EdTx N447S, or even native EdTx, did not induce IgA Ab responses in the blood or in mucosal secretions. We next analyzed APC (i.e., B cells, macrophages and DCs) in the skin, inguinal lymph nodes, mesenteric lymph nodes (MLN) and Peyer’s patches (PP) of mice 18 hrs after transcutaneous application of OVA alone, OVA plus CT or OVA plus EdTx. Transcutaneous application of CT or EdTx affected the distribution of APCs and their expression of MHC class II and co-stimulatory molecules (i.e., CD80 and CD86) in the skin and draining lymph nodes. However, only CT had a significant effect on these parameters in the MLN and intestinal PP. Ongoing studies will determine the molecular signaling (i.e., cytokines and homing receptors) triggered by the transcutaneous application of CT vs. EdTx to affect systemic vs. mucosal tissues.
(Supported by NIH Grant: AI 43197)
Head and Neck Lymph Node (HNLN) B Cells from L-Selectin−/− (L-Sel−/−) Mice Show αE Expression Independent of Cholera Toxin (CT) Exposure (41.9)
Authors: Riccardi Carol , Pascual David W. 
Published: January 2023

Abstract
Oral immunization with soluble protein plus adjuvant stimulates excellent regional, but not distal Ab responses. Although low level expression of distal immune B cells is observed, these wane with time. Studies examining homing receptor expression should provide insight into understanding how distal mucosal immunity is maintained. Studies showed that B lymphocyte trafficking in the HNLN and NALT is L-Sel-dependent. However, the absence of L-Sel enhanced distal B cell maintenance following oral CT immunization of L-Sel−/− mice and was partly compensated by increased αEβ7+ B cells, as evident by the significantly increased IgA antibody-forming cells (AFC) in L-Sel−/− nasal passages (NP) when compared to B6 NP. These IgA responses were supported by elevated HNLN IgA AFC responses (10- to 100-fold greater), and these AFC endured beyond 42 days, but not in B6 mice. To discern whether αEβ7 expression was influenced by CT directly, naive HNLN, mesenteric LN (MLN), and Peyer’s patches (PP) were isolated and stimulated in vitro with LPS, LPS + CT, Con A, or media. The mitogen-stimulated L-Sel−/− HNLN, MLN, and PP B cells showed early expression of αEβ7 at 48 hrs, but by 96 hrs, the L-Sel−/− HNLN clearly showed the greatest levels (30–35%) of αEβ7+ B cells. Both unstimulated and stimulated HNLN B cells showed similar levels of αEβ7 expression, and CT did not enhance αEβ7 expression. MLN and PP showed fewer (~10%) αEβ7+ B cells, and B6 B cells from each tissue showed &lt;10% αEβ7+ B cells. These studies show that αEβ7 compensates for the absence of L-Sel on B cells, and these B cells are mostly derived from HNLN, and not from the gut.
Supported by NIH AI-55563.
Epithelial cell-intrinsic IKKβ expression regulates innate and adaptive immunity in the gut (42.4)
Authors: Zaph Colby , Troy Amy E. , Taylor Betsy C. , Berman-Booty Lisa D. , Guild Katherine J. , Du Yurong , Yost Evan A. , Gruber Achim D. , May Michael J. , Greten Florian R. , Eckmann Lars , Karin Michael , Artis David 
Published: January 2023

Abstract
Intestinal epithelial cells (IECs) provide a primary physical barrier against commensal and pathogenic microorganisms in the gastrointestinal (GI) tract, but the influence of IECs on the regulation of immunity is unknown. Here we show that IEC-intrinsic IKKβ-dependent gene expression is a critical regulator of dendritic cell (DC) and CD4+ T cell responses in the gut. Following infection with the parasite Trichuris, littermate control mice develop pathogen-specific TH2 cytokine responses and eradicate infection while mice with an IEC-specific deletion of IKKβ (IkkbΔIEC mice) fail to do so. Further, IkkbΔIEC mice exhibit exacerbated production of DC-derived IL-12/23p40 and TNF-α, heightened levels of CD4+ T cell-derived IFN-γ and IL-17, and develop severe intestinal inflammation. Blockade of proinflammatory cytokines during Trichuris infection ablates the requirement for IKKβ in IECs to promote CD4+ TH2 cell-dependent immunity, identifying a critical role for IECs in limiting type 1 cytokine production in the GI tract. These results suggest that the balance of IKKβ-dependent gene expression is critical for intestinal immune homeostasis by promoting mucosal immunity and limiting chronic inflammation.
This work is funded by the NIH, CCFA and the Irvington Institute.