bluuebunny/crossref_metadata_2025_split huggingface dataset:
Signals derived from intestinal microbial communities augment anti-viral immunity (46.11)
Authors: Abt Michael , Beiting Dan , Kobuley Dmytro , Wherry John , Artis David 
Published: January 2023

Abstract
Alterations in intestinal microbial communities in humans are associated with enhanced susceptibility to inflammatory diseases suggesting that signals derived from commensal bacteria may influence the development and/or function of the immune system. Supporting this, germ-free (GF) mice exhibit reduced numbers of lymphocytes in the periphery and intestinal intraepithelial compartment. However, whether alterations in the composition of commensal bacteria influence immunity to infection remains poorly defined. To test this, mice housed under conventional (CNV) or GF conditions were infected with acute LCMV and the development of antigen-specific CD8+ T cell responses were analyzed. At day 7 p.i., GF mice exhibited a significant reduction in the frequency and number of LCMV-specific CD8+ T cells in multiple tissues including the spleen and intestinal intraepithelial compartment. The diminished CD8+ T cell response was not the result of inherent developmental defects in GF mice as depletion of intestinal bacteria in CNV mice via antibiotics also resulted in diminished LCMV-specific CD8+ T cell responses and impaired viral clearance. Further, adoptive transfer of CFSE-labeled LCMV-specific CD8+ T cells revealed a delay in the activation and expansion of LCMV-specific CD8+ T cells in antibiotic-treated mice. Collectively, these studies indicate that signals derived from intestinal bacteria aid in the induction of a rapid immune response required for immunity to viral infection.
Evaluation of the botulinum neurotoxin neutralizing efficiency in immunoglobulin purified from clinical volunteers vaccinated with recombinant botulinum vaccine (46.5)
Authors: Shearer Jeffry , Vassar Michelle , Niemuth Nancy , Swiderski William , Holley H. , Metcalfe Karen , Hart Mary 
Published: January 2023

Abstract
Efficacy of the Recombinant Botulinum Vaccine A/B (rBV A/B) against lethal intoxication with botulinum neurotoxin complex (BoNT) cannot be evaluated directly in humans; therefore, efficacy will be based on meeting the requirements of the FDA Animal Rule. Serum BoNT neutralizing antibody concentration (NAC) is being proposed as a correlate of protection to evaluate efficacy in nonclinical models. Under this approach, NAC levels in animals vaccinated either with rBV A/B or passively immunized with BoNT neutralizing antibody will be used to estimate the potential level of protection in humans with similar NAC levels. Estimating the level of protection based on NAC requires establishing NAC neutralizing efficiency (the amount of BoNT neutralized per NAC unit). The neutralizing efficiency of human BoNT neutralizing antibody was evaluated using immunoglobulin purified from pooled sera of clinical volunteers vaccinated with rBV A/B. Solutions of immunoglobulin were prepared at three different NAC levels, mixed with serial dilutions of BoNT serotype A or serotype B and injected into mice. At each NAC level, 50% and 99% neutralization of BoNT was determined by probit analysis of mortality dose-response curves. A simple linearly proportional relationship does not exist between NAC level and the amount of BoNT neutralized. Extrapolations of level of protection must consider variability in BoNT neutralizing efficiency at different NAC levels.
A novel subset of lung cells responds to IL-33 and mediates innate Th2-type immune responses (48.4)
Authors: Bartemes Kathleen , Iijima Koji , Kobayashi Takao , Kephart Gail , Kita Hirohito 
Published: January 2023

Abstract
Interleukin (IL)-1 family member cytokines are likely pivotal as interfaces between innate and adaptive immune responses. IL-33, an IL-1 family member, is implicated in airway Th2-type responses, including eosinophilia, increased mucus production and hyperresponsiveness. Our goal was to elucidate the immunological mechanisms involved in IL-33-mediated Th2-type responses in the lungs. Airway exposure of naïve BALB/c mice to IL-33 resulted in bronchoalveolar lavage (BAL) eosinophilia and increased lung levels of IL-5 and IL-13, but not IL-4 or IFNγ. Naïve Rag1-/- mice showed similar airway eosinophilia and cytokine responses as wild-type mice, suggesting a role for innate immune cells. By FACS analysis, IL-5-producing lung cells lacked CD3, B220 and other lineage markers (i.e. Lin-), but they expressed CD44 and CD25; markers for NK cells, mast cells, basophils and eosinophils were undetectable. FACS-sorted Lin-CD25+CD44+ lung cells expressed GATA-3 and the IL-33 receptor, T1/ST2, and responded to IL-33 by producing IL-5 and IL-13; morphologically, these lung cells resembled lymphocytes. In vitro blockade of CD25+ cells with anti-CD25 mAb (PC61) abrogated IL-33-induced Th2 cytokine production. Thus, mouse lungs contain a novel subset of innate immune cells that respond to IL-33. These airway cells may initiate early Th2-type immune responses and induce asthma and other allergic diseases.
Expression of co-stimulatory TNFR2 enhances resistance of CD4+FoxP3- conventional T cells to suppression by CD4+FoxP3+ regulatory T cells (49.13)
Authors: Chen Xin , Hamano Ryoko , Subleski Jeffrey , Hurwitz Arthur , Howard O.M.Zack , Oppenheim Joost 
Published: January 2023

Abstract
Our previous study showed that TNFR2 is preferentially expressed by CD4+FoxP3+ regulatory T (Treg) cells and expression of this molecule identified the maximally suppressive Treg cells. TNFR2 is also expressed by a small fraction of CD4+FoxP3- conventional T (Tconv) cells in normal mice and its expression is up-regulated by T cell activation. Our data indicate that TNFR2 also co-stimulates Tconv cells. In this study, by using FoxP3/gfp KI mice, we showed that TNFR2 signaling did not induce FoxP3- CD4 cells to become suppressive. TNFR2 is associated with greater suppressive functions when expressed by Treg cells and is associated with greater resistance to suppression when expressed by Tconv cells, because TNFR2+ Tconv cells were potently suppressed by FoxP3+TNFR2+ Treg cells, while resisting suppression by FoxP3+TNFR2- Treg cells. Furthermore, in mice with 4T1 breast tumor and LLC, intratumoral Tconv cells expressing elevated levels of TNFR2 acquired the capacity to resist the suppression mediated by LN-derived Treg cells. However, they remained susceptible to inhibition by more suppressive tumor infiltrating Treg cells which expressed higher levels of TNFR2. Intratumoral Treg cells express more TNFR2 and they are able to overcome the inhibition-resistance of intratumoral Tconv cells, resulting in a dominant immunosuppressive tumor environment. (Funded by NCI Contract N01-CO-12400)
Re-programming of IL-10 signal transduction: an inflammation-induced mechanism to control T cell susceptibility to suppression (49.18)
Authors: Raimondi Giorgio , Kowalkowski Frank , Kane Lawrence , Thomson Angus 
Published: January 2023

Abstract
Modulation of regulatory T cell (Treg) suppressive function may play an important role in the development of autoimmune diseases, tumor growth and transplant outcome. Currently, little is known about the molecular mechanisms that render T cells refractory to suppression. Since IL-10 is an important mediator of Treg activity, we investigated the influence of pro-inflammatory mediators on the IL-10 signaling pathway in T cells. The mouse CD4 T cell line D10 was incubated for various periods with the supernatant of bone marrow-derived dendritic cells stimulated overnight with the TLR4 ligand LPS (MATSup). We then measured the integrity of the IL-10 signaling pathway by assessing phosphorylation of the transcription factor STAT3 in response to titrated amounts of IL-10. Even after 24h incubation with MATSup, phosphorylated STAT3 was readily detected in T cells in response to IL-10. Surprisingly, measurement of cytoplasmic levels of total STAT3 and of pro-inflammatory STAT1 revealed short- term (3 to 6 hours) reduction of both transcription factors, followed by upregulation with an altered ratio between STAT3 and STAT1 in favor of the latter. This altered ratio appeared within 12 hours and was preserved for at least 24 hours. These results suggest that pro-inflammatory mediators re-program the IL-10 (and probably other) signaling pathway in T cells, reducing their susceptibility to suppression. Interference with this process may represent an effective immunomodulatory strategy.
Strong CD28 co-stimulation Limits Generation of Regulatory T cells from Naïve T cells through Lck and PI3K signaling (49.4)
Authors: Semple Kenrick , Nguyen Antony , Yu Xue-Zhong 
Published: January 2023

Abstract
CD28 co-stimulation is required for the generation of regulatory T cells (Tregs) in the thymus as well as in the periphery through Lck-signaling. However, high level of CD28 limits the generation of Tregs from naïve CD4 T cells, although underneath mechanism(s) has not been defined. Here we investigated the role of CD28-mediated signaling pathways in the down-regulation of Treg generation. To address the question, we used a series of transgenic (Tg) mice on CD28-deficient background that bears WT CD28 or mutated CD28 in its cytosolic tail incapable of binding to Lck, PI3K or Itk. Under the culture condition with TGFβ that favors Treg generation, the generation of Tregs (CD25+Foxp3+) from naïve CD4 T cells with WT CD28 Tg (CD25-Foxp3-) was inversely related with the level of CD28 co-stimulation mediated by anti-CD28 or B7. Regardless of exogenous IL-2, the suppression of Treg generation caused by strong CD28 signals was intact in the T cells with CD28 Tg incapable of binding Itk, compromised in those incapable of binding PI3K, and essentially abrogated in those incapable of binding Lck. The essential role of Lck and contributory role of PI3K in CD28-mediated down-regulation of Treg generation were confirmed by using Lck- or PI3K-specific inhibitors. These results reveal novel mechanistic insight how CD28 can negatively regulate Treg generation, which may provide rationale for promoting T-cell immunity or tolerance by regulating Tregs through targeting CD28-signaling.
TORC1 and TORC2 selectively regulate CD4+ T helper cell lineage differentiation (50.35)
Authors: Delgoffe Greg , Waickman Adam , Pollizzi Kristen , Xiao Bo , Worley Paul , Powell Jonathan 
Published: January 2023

Abstract
The mammalian target of rapamycin (mTOR) is a highly conserved serine-threonine kinase that regulates cell division, survival, and translation. Downstream mTOR signals via two functionally distinct complexes, TORC1 and TORC2. We have previously shown T cells deficient in mTOR fail to differentiate into Th1, Th2, or Th17 cells and instead become Foxp3+ regulatory cells. We further dissected mTOR signaling in T cell differentiation by deleting Rheb (the activator of TORC1) and rictor (an essential TORC2 subunit). T cells lacking Rheb selectively lack TORC1 activity, and, like mTOR mutants, fail to differentiate into Th1 or Th17 cells when skewed. However, Rheb-deficient T cells retain their ability to become Th2 cells. This is due in part to diminished STAT4 and STAT3 activation, but elevated STAT6 activation. Alternatively, rictor-deficient cells specifically lack TORC2 activity, and fail to become Th2 cells when skewed. However, they retain the ability to become Th1 and Th17 cells, with a pattern of STAT activation that inversely correlates with TORC1 mutants. Interestingly, neither TORC1 nor TORC2 deficiency alone is sufficient to induce Treg differentiation. In vivo, TORC1 deficiency protects against EAE while TORC2 deficiency leads to accelerated disease. Our data support a novel paradigm in which TCR engagement in the absence of mTOR defaults to a regulatory fate, and TORC1 and TORC2 activation selectively regulate T cell effector lineage differentiation.
Studying the cooperativity at the SLP-76 signaling complex critical for the immune response (50.7)
Authors: Barda-Saad Mira , Shirasu Naoto , Pauker Maor , Hasan Nirit , Perl Orly , Balbo Andrea , Yamaguchi Hiroshi , Houtman Jon , Appella Ettore , Schuck Peter , Samelson Lawrence 
Published: January 2023

Abstract
Following T-cell antigen receptor (TCR) engagement, multiprotein signaling complexes essential for regulating T-cell functions are formed. A multi-molecular complex consisting of SLP-76, Nck and VAV is recruited to the T-cell/antigen presenting cell (APC) site during initial T cell activation and is crucial for regulation of the actin machinery. Most of the studies performed to date have concentrated on the role of the signaling molecules that comprise this molecular complex. However, the regulation of the complex formation and the cooperation and competition among its components during formation have not yet been explored. In this study, a multidisciplinary approach including molecular imaging, biochemical and biophysical techniques was utilized in order to define the composition, stoichiometry, and specificity of the SLP-76:Nck:VAV complex. High-resolution imaging techniques, such as fluorescence resonance energy transfer (FRET) analysis, were performed in live activated T cells in order to focus on the binding properties within the trimolecular complex. Our data reveal a direct interaction between Nck and VAV which is mediated by the binding of the C-terminal SH3 domain of Nck to VAV’s N-terminal SH3 domain. Disruption of the VAV:Nck interaction significantly impaired actin polymerization. This research provides novel insights into the regulation of a signaling complex formation critical for immune response.
Identification of a class of small molecules that up-regulate production of anti-inflammatory cytokine IL10 via inhibition of GSK3β (51.10)
Authors: Gururaja Tarikere , Ding Rongxian , Yung Stephanie , Yi Sothy , Zhao Haoran , Singh Rajinder , Kinoshita Taisei , Masuda Esteban 
Published: January 2023

Abstract
Interleukin-10 (IL-10), also known as human cytokine synthesis inhibitory factor (CSIF), is an anti-inflammatory cytokine produced primarily by monocytes and to a lesser extent by lymphocytes. This cytokine exhibits pleiotropic effects in immunoregulation and inflammation and is capable of inhibiting synthesis of various pro-inflammatory cytokines such as IFNγ, IL-2, IL-3, IL-6, TNFα and GM-CSF made by cells such as macrophages and Th-1 cells. During the course of our investigation to find small molecules that are capable of inhibiting the production of a pro-inflammatory cytokine, IL-23, we came across a sub-set of small molecules that exhibited an interesting property of up-regulating expression of IL-10 besides inhibiting IL-23 production. Such observation prompted us to further investigate the mechanism of action and target identification for this class of compounds. Preliminary experiments revealed that enhanced IL-10 production upon LPS stimulation of human THP-1 monocytic cells in the presence of compounds appears to be due to inhibition of GSK3β kinase. Results from these studies were further substantiated by screening the lead candidate against a panel of kinases in-vitro including GSK3β and a cell-based β-cateinin accumulation assay that directly measures GSK3β kinase on-target activity.
Modulation of CD40L expression by polypyrimidine tract binding protein (PTB) (51.8)
Authors: Matus Nicodemos Rodrigo , Vavassori Stefano , Laughlin Jennifer , Covey Lori 
Published: January 2023

Abstract
The expression of CD40L is tightly regulated at the posttranscriptional level throughout a time course of T cell activation, which corresponds to a significant increase in message stability at late times of activation (24-48 hr). Previous work revealed a cytoplasmic polypyrimidine tract binding protein (PTB)-containing-complex binds to the CD40L 3’UTR at late times of T cell activation. We used RNA interference to downregulate PTB in D1.1 Jurkat T cell line to address the functional roles of PTB in CD154 mRNA stability and surface expression. Real-time qPCR measurement of the CD154 mRNA decay revealed that downregulating PTB caused a &gt; 2-fold decrease in the CD40L mRNA half-life. The downregulation of PTB also caused an approximate 2-fold decrease in the mean fluorescence (MFI) of CD40L. Analysis of PTB cellular distribution during a time course of CD4+ T cell activation revealed cytoplasmic and nuclear localization in all resting and activated cells. However, there was an increase in expression at late times of activation. Binding studies revealed that CD40L mRNA is bound by nuclear PTB at all times of activation indicating that the requirements for binding of CD40L message by nuclear versus cytoplasmic PTB is highly distinct. Finally, the binding of CD40L message corresponded to a change in phosphorylation status of cytoplasmic PTB. These findings support a model whereby activation-induced phosphorylation of PTB is required for cytoplasmic binding to CD40L mRNA.
Lymphocyte type determines the effect of single nucleotide polymorphisms on Lymphotoxin Alpha expression. (51.9)
Authors: Yokley Brian , Posch Phillip 
Published: January 2023

Abstract
The lymphotoxin alpha (LTA) gene contains many single nucleotide polymorphisms (SNPs); some influence expression levels and are associated with disease. The impact of SNPs on LTA expression primarily has been analyzed in B cells, yet T cells contribute the most to LTA levels. Thus, we investigated the cell type-specific effects of the LTA +81C/A and +369G/C SNPs on expression. The LTA +81A SNP significantly increased expression relative to +81C in T cells; opposite of its impact in B cells. Conversely, the LTA +369G SNP was found to significantly decrease expression only in T cells under basal conditions. The impact of this SNP was partly due to stronger binding to and repression of LTA +369G by Sp1. ChIP assays showed Sp1 binding to this region of the LTA locus in vivo, but the two Sp1 sites in close proximity could not be distinguished. However, mutation of the LTA +369 SNP-associated Sp1 site did result in an expected increase in expression further implicating that Sp1 binding to this site suppresses expression. Numerous studies associate SNPs with disease and mechanisms are proposed to explain these associations based on the known impact of the SNP on expression. We show that LTA SNPs can differentially impact expression levels depending on stimulation status and cell type. Thus, it is critical to consider the cell type-specific impact of a SNP and the pertinence of different cell types to disease pathogenesis when proposing a mechanism behind a disease association.
Towards the development of a universal T cell influenza (FLU) vaccine (52.10)
Authors: Xu Dong , Popmihajlov Zoran , Arnold Katherine , Sumner Jonathan , Hussell Tracy , Feldmann Marc , Smith Kendall 
Published: January 2023

Abstract
Present day FLU vaccines have not changed since WWII, and depend upon the generation of neutralizing antibodies, which the virus readily escapes via mutations. Therefore, we focused on immunizing T cells with serologically non-cross reactive viral strains in a murine model. C57Bl/6 mice infected intra-nasally (i.n.) with H3N2-FLU were protected against a lethal challenge of H1N1-FLU. Cellular composition of the Broncho-Alveolar Lavage (BAL), lungs and mediastinal lymph nodes monitored by flow-cytometry 8 days after infection with H1N1-FLU yielded a vigorous cellular response, with a 2:1 ratio of CD8+ vs. CD4+ T cells in the BAL, and a high proportion of immunodominant nucleoprotein (NP) peptide-specific tetramer+ cells (~ 40%). By comparison, mice immunized i.n. with replication-incompetent adenovirus containing NP (10e7 pfu) were also protected against lethal challenge. Although the H3N2-FLU-infected control animals mobilized greater numbers of CD8+ T cells in the BAL 14d post H1N1 challenge, the vaccinated animals had 3X as many NP-specific tetramer+ T cells. Therefore, a non-replicating adenovirus vaccine can protect against a lethal challenge with a non-serologically cross-reactive flu strain, so that a “T cell vaccine” is effective in preventing symptomatic disease.
Differential lymphocyte activation induced by mammalian-derived Ebola virus-like particles in a post-exposure murine model. (52.23)
Authors: Anthony Scott , Bradfute Steven , Bavari Sina 
Published: January 2023

Abstract
Ebola virus is a member of filoviradae, a family of negative single stranded RNA viruses which cause hemorrhagic fever with mortality rates of up to 90%. To date there is no approved treatment to combat this deadly disease. During studies in a murine model, it was found that Ebola virus-like particles (eVLP) provided sufficient protection when given 24 hours after Ebola virus exposure. However, a study determining the specific, in vivo eVLP activation of immune cell subsets during the course of infection has not been performed. Utilizing flow cytometry, lymphocytes within the draining lymph node of the eVLP vaccination site were compared to corresponding cells in unvaccinated mice. In preliminary studies, eVLP induced significantly different levels of B cell activation on day 7 of the infection. In day 7 eVLP vaccinated mice, activation was found to be significantly higher in the draining inguinal lymph node compared to the contra-lateral inguinal node. Future studies will attempt to expand upon the functional significance of this lymphocyte response, while also incorporating the role of macrophages and dendritic cells. This study better characterizes the local, in vivo, adaptive immune responses induced by eVLP in this post-exposure model of protection. By understanding the proper adaptive immune response to therapeutically protect against this virus, it may be possible to modulate future therapies to target these specific successful responses for further up regulation.
Immunological evaluation of Recombinant Mycobacterium vaccae expressing Mycobacterium tuberculosis-specific antigens in mice (52.5)
Authors: Amoudy Hanady 
Published: January 2023

Abstract
Tuberculosis (TB) is considered to be one of the leading causes of death from infectious diseases all over the world. The danger is coupled with the emergence of multi-drug resistant strains which renders treatment a real challenge. The Vaccine-BCG- used against TB is very disputable giving wide ranges of protection reaching as low as nil, in some parts of the world. Several M. tuberculosis-specific DNA regions including RD1 were found to be deleted from the vaccine strains. Deleted region RD1 encode proteins that are major targets for T cell recognition in humans and animals infected with M. tuberculosis and M. bovis, respectively. The aim of this study is to express open reading frames (ORFs) from RD1 region in M. vaccae and evaluate their ability to induce specific immune responses in mice. The appropriate ORFs were cloned in M. vaccae using the shuttle vectors pSMT3, and pDE22. Expression of the corresponding proteins was detected by western blotting using anti-hemagglutinin (HA) and/ or antigens- specific antibodies. Recombinant mycobacteria were injected in Balb/C mice, spleens were removed, and specific immunity to the recombinant antigens was assessed using standard spleenocytes culture and cytokines quantitation assays. This system of expression using shuttle vectors proved to be a promising system to identify candidate antigens to be used in diagnosis and to develop improved vaccine strains of BCG.