bluuebunny/crossref_metadata_2025_split huggingface dataset:
CD4+ T cell regulation of CD25 expression controls the development of effector CD8+ T cells in acute and chronic virus infections. (137.14)
Authors: Molloy Michael , Zhang Weijun , Usherwood Edward 
Published: January 2023

Abstract
Both CD4+ T cell help and IL-2 have been postulated to ‘program’ activated CD8+ T cells for memory cell development. However, the link between these two signals has not been well elucidated. We have studied effector and memory CD8+ T cell differentiation following infection with vaccinia virus and murine gammaherpes virus in the absence of both CD4+ T cells and CD25. Expression of CD25 on antigen-specific CD8+ T cells peaked 3-4 days after infection and was dependent on CD4+ T cell help. CD4+ T cell or CD25-deficiency led to normal early effector CD8+ T cell differentiation, but a subsequent lack of accumulation of CD8+ T cells resulting in overall decreased memory cell generation. Interestingly, KLRG1high CD127low short-lived effector cells were diminished in the absence of IL-2 signaling, while memory precursors remained intact. Interestingly, the mononucleosis-like expansion of Vβ4+ T cells seen in mice infected with MHV-68 was impaired in a similar fashion to antigen specific responses. Upon secondary antigen encounter, CD25-deficient CD8+ T cells were capable of undergoing robust expansion, but short-lived effector development was again impaired. These results demonstrate that CD4+ help and IL-2 signaling are linked via CD25 upregulation, which controls the expansion and differentiation of antigen-specific effector CD8+ T cells.
Cytokine production patterns identify novel CD8 effector subsets (137.18)
Authors: Rudenga Benjamin , Khanna Kamal , Lefrançois Leo 
Published: January 2023

Abstract
Underlying the continuing rational development of vaccines to infectious agents is a deeper understanding of the immunological determinants affecting the outcomes to various strategies of vaccination. For example, different forms of Listeria monocytogenes (LM) based vaccines elicit distinct immune responses. Thus, heat killed LM is less effective than irradiated LM which is in turn less effective than live LM in inducing protective CD8 T cell immunity. To characterize the underlying reasons for these differences we analyzed the genetic signature of CD8 T cells responding to the different immunogens. Interestingly, cytokine and chemokine genes were induced very early after activation and their levels correlated with the “strength” of the immunogen. We have begun to examine the expression of some of these cytokines at the protein level using multiplex bead assays and intracellular staining. Several cytokines and chemokines have been identified as potential targets; however, IL-3 was found to be expressed early in the immune response to live LM and was also induced after vesicular stomatitis virus (VSV) infection. Only a subset of CD8 T cells produced IL-3 and we are now determining the relationship between this population, other effector subsets, and memory development. Ongoing studies are focused on deciphering the complexity of cytokine production by CD8 T cells responding to infection in the hope of designating effector and memory lineages based on the identified patterns.
TULA-family proteins are key to the regulation of T-cell-driven inflammatory responses. (138.13)
Authors: Newman Tiffanny , Carpino Nick , Ganea Doina , Safadi Fayez , Yen Jimmy , Tsygankov Alexander 
Published: January 2023

Abstract
The novel TULA family consists of two proteins recently implicated in cellular regulation. TULA-1 is expressed only in lymphoid cells and has been shown to exert a pro-apoptotic effect in T-cells, possibly through its interaction with AIF. TULA-2 is expressed ubiquitously and is an active phosphatase capable of dephosphorylating proteins involved in receptor-mediated signaling. CD4+ T-cells from mice lacking both family members (dKO) are hypersensitive to TCR stimulation. Initial characterization T-cell subpopulations in TULA dKO mice revealed an atypical CD45RB distribution among dKO CD4+ T-cells. Further analysis of memory and regulatory markers indicate that both memory and regulatory T-cell populations are expanded in dKO mice. To determine in vivo consequences of these changes, we utilized two mouse models of inflammation in which T-cells play a key role: trinitrobenzene sulfonic acid (TNBS)-induced colitis and colitis induced by the adoptive transfer of CD45RB-high CD4+ T-cells. Studies utilizing TNBS indicated that TULA dKO mice are more sensitive to colitis as compared to WT mice and that the increased sensitivity is dependent on T-cells. Consistent with these findings, dKO CD45RB-high CD4+ T-cells were significantly more colitogenic than the in WT mice in the transfer colitis model. Taken together, our data suggests that TULA-family proteins are key to the physiological regulation of T-cell reactivity that drives inflammatory responses in the intestine.
Licensing of naïve T cells for rapid TNF production (138.14)
Authors: Priyadharshini Bhavana , Welsh Raymond , Greiner Dale , Gerstein Rachel , Brehm Michael 
Published: January 2023

Abstract
TNF is a potent inflammatory cytokine with numerous immunological functions, including the ability to limit the magnitude of virus-specific T cell responses during infection. Naïve T cells in secondary lymphoid organs (SLOs) exhibit a unique ability to produce TNF rapidly after activation and prior to acquiring other effector functions. Peripheral T cells are a heterogeneous population of cells at various stages of post-thymic development. To determine how maturation influences the licensing of T cells to produce TNF, we compared cytokine profiles of single positive thymic T cells, recent thymic emigrants (RTEs) and matured-naïve T cells during TCR activation. Thymic T cells exhibited a poor ability to produce TNF when compared to splenic T cells, despite possessing a similar potential for activation. Moreover, thymic T cells had significantly reduced levels of TNF message and phosphorylated MAPK (Erk1/2) when compared to their splenic counterparts. Stimulation of thymic T cells in the presence of splenic B cells and APCs partially licensed the T cells to produce TNF but not to the same extent as peripheral T cells. RTEs had a significantly enhanced ability to produce TNF as compared to thymic T cells, but fully matured naïve T cells were the most efficient TNF producers. Together, these findings suggest that TNF expression by naïve T cells is regulated via a gradual licensing process that requires functional maturation in peripheral lymphoid organs.
Mechanism of exaggerated inflammation in A2A adenosine receptor-deficient mice. (138.28)
Authors: Subramanian Meenakshi , Kini Radhika , Madasu Manasa , Ohta Akiko , Sitkovsky Michail , Ohta Akio 
Published: January 2023

Abstract
Extracellular adenosine regulates inflammatory responses via A2A adenosine receptor (A2AR). Indeed, A2AR-deficiency results in much exaggerated inflammatory tissue damage, indicating non-redundancy of adenosine-A2AR pathway in the inhibitory mechanism of immune activation. In this study, we found that intravenous injection of concanavalin A into A2AR-deficient mice induced not only stronger hepatic damage than in wild-type mice but also higher levels of cytokines, i.e. TNF-α, IL-4 and IFN-γ. Since NKT cells are responsible for the early production of these cytokines after concanavalin A injection, NKT cells were suspected to be under physiological control of adenosine-A2AR pathway. To directly examine the role of A2AR in NKT cell activation, we specifically stimulated NKT cells with α-galactosylceramide in the presence of dendritic cells. As a result, cytokine production from NKT cells decreased in the presence of A2AR agonists, suggesting that A2AR signaling can down-regulate NKT cell activation. We further injected α-galactosylceramide into mice and observed higher cytokines production in A2AR-deficient mice comparing to wild-type mice. Correspondingly, α-galactosylceramide induced stronger liver damage in A2AR-deficient mice. These results strongly suggest that NKT cell activation is controlled by endogenous adenosine via A2AR, and this physiological regulatory mechanism of NKT cells is critical in the control of tissue-damaging inflammation.
High-throughput multi-parameter analysis of human immune responses to the seasonal Influenza vaccine in the elderly and young subjects: defining metrics of immune health and immune senescence (138.4)
Authors: Furman David , Shen-Orr Shai , Kidd Brian , Dekker Corry , Maecker Holden , Davis Mark 
Published: January 2023

Abstract
Aging is a complex process in which the immune function is severely affected. Increased prevalence of infectious diseases and poor vaccine responses are characteristic in the elderly. Here we seek to define the immune response to the seasonal inactivated Influenza vaccine in young subjects and in an elderly population with the aim of finding signatures of immune health and immune senescence. Ninety-one subjects, age-ranges 20-30, 60-79 and 80-96, were drawn prior to the vaccine, 7 and 21 days after vaccination. PBMC were isolated to phenotype 18 immune cell subsets and for high-throughput analysis of signaling phosphoproteins in multiple cell-subsets, after various stimulations. Serum samples were used to determine cytokine levels and antibody titers to the vaccine strains and to non-circulating Influenza strains. Dramatic age-related changes in STAT signaling were observed in CD8 and CD4 T cells when stimulated with IL-6, IFN-α or IFN-γ. The level of PLCγ phosphorylation in B cells from older subjects was significantly lower than the observed in the younger group, independently of the reduced B cell frequency. Weak antibody response against the vaccine strains while high level of seroprotection against older non-circulating vaccines was seen in the elderly and strongly associated to the IL-6 or IFN-α pathways. A better comprehension of human immune aging is necessary for designing age-specific vaccines with higher efficacy.
House dust mite extract directly regulates the alternative activation of macrophages by a TLR4-independent mechanism. (141.3)
Authors: Keegan Achsah , Qi Xiulan , Vogel Stefanie , Shirey Kari , Heller Nicola 
Published: January 2023

Abstract
Asthma is a clinical disorder characterized by episodes of airway narrowing, inflammation, and mucus production. House dust mites (HDM) are a major source of allergens that contribute to allergy and asthma. HDM contains a complex mixture of multiple agonists of pattern recognition receptors including bacterial-derived LPS, and multiple allergen groups. A recent report demonstrated a role for epithelial-cell expressed TLR4 in the Th2 response to nasal challenge with HDM. We and others have shown that macrophages activated by IL-4/IL-13 differentiate into alternatively activated macrophages (AAM) and participate in the recruitment of eosinophils during Type 2 inflammation. To determine whether HDM has a direct effect on macrophages, we treated bone marrow-derived macrophages (BMM) with HDM in the presence or absence of IL-4. HDM extract induced expression of YM1/2 (2-fold) and Mac 3, but not arginase I. IL-4 induced expression of AAM proteins including arginase I, YM1/2 (8-fold), and Mac3. Interestingly, HDM extract strongly induced the expression of a 39 kDa protein (9-fold) also recognized by the anti-YM1 antibody; this protein was modestly induced by IL-4. Together, HDM and IL-4 strongly induced YM1/2 and the 39 kDa protein. The HDM-induced responses were TLR4-independent. These results suggest that molecular patterns found in HDM extract regulate alternative macrophage activation by a TLR4-independent mechanism. Supported by AI059775 (ADK) and AI18797 (SNV)
The novel role of downstream of kinase-1 in the ovalbumin-induced murine model of asthma (141.4)
Authors: Park Yeong-Min , Lee Chang-Min , Park Jin Wook , Lee Chun Geun 
Published: January 2023

Abstract
The downstream of kinase (DOK)-1, a 62 kDa adaptor protein negatively regulates the protein tyrosine kinase (PTK) pathway in mast cells, but the role of DOK-1 in a murine model of asthma has not been defined. This study provides experimental evidence demonstrating that DOK-1 is a key regulator for aiway inflammation in a murine model of asthma. Here, we have found that the markable reduction of pulmonary CD4+ T cell population and Th2 cytokines leads to the inhibition of pulmonary CD4+ T cell differentiation into Th2 cells in DOK-1 overexpression mice. DOK-1 overexpression in STAT-6 knockout (KO) mice increased the expression of STAT-4 and T-bet, which is transcription factor for Th1 differentiation. Conversely, Th2 regulatory molecules, STAT-6 and GATA-3 are impaired by DOK-1 overexpression in STAT-4 KO mice. Taken together, DOK-1 regulates Th1/Th2 immune balance through STAT4- and STAT6-dependent pathways, our findings provide additional insight into the immunological function of DOK-1 in Th2 inflammation and suggest a new potential therapeutic target for the treatment of asthmatic patients.
Early expression of IL-19 in asthmatic airways promotes Th2 response (141.5)
Authors: Hung Li Yin , Nguyen Tiffany , Huang Fei , Chen Yin , Wu Reen 
Published: January 2023

Abstract
IL-19 is a proinflammatory cytokine that belongs to the IL-10 cytokine family. In addition to its known connection with psoriasis, previous work in our lab along with others, have also shown its association with asthma. Using cultured primary mouse airway cells, we demonstrated that Th2 cytokines such as IL-4 and IL-13 can induce the expression of IL-19 mRNA. In a mouse OVA-induced asthma model, we showed that IL-19 expression is increased in lung tissue during early disease stages. Additionally, we observed that other IL-10 family members such as IL-20, IL-22, or IL-24, were not induced. We further identified that airway epithelial cells, rather than the alveolar macrophage, were the source of IL-19 in the asthma model. Although we show that IL-19 can promote Th2 cytokine production by activated T cells, restimulation with IL-19 does not further increase Th2 cytokine production on memory cells. This phenomenon correlates with a down-regulation of IL-20R2 when cells are differentiated in the presence of IL-19, indicating a negative feedback regulatory mechanism. These results demonstrate that IL-19 is produced by airway epithelial cells during the early stage of the asthmatic response, and may further amplify the Th2 response by acting on activated T cells recruited to the lung during later disease stages by inducing Th2 cytokine production.
Invariant NKT cells in pig lung: a potential asthma model (141.8)
Authors: Gourapura Renukaradhya , Manickam Cordelia , Dwivedi Varun 
Published: January 2023

Abstract
Biomedical research in pigs is critical for better human health, as they are immunologically and physiologically more similar to humans, and are an excellent animal model to alleviate human asthma. Natural Killer T (NKT) cells with an invariant TCR are called "invariant NKT" (iNKT). iNKT cells are activated by CD1d-bound glycolipids with a unique α-anomerically linked sugar (e.g. α-GalCer). For the first time in pigs, we detected iNKT cells and CD1d expression. The pig iNKT cells proliferated and secreted both Th1 and Th2 cytokines when cultured in the presence of α-GalCer. In particular, iNKT cells from lung, and lung draining lymph nodes proliferated to higher numbers (&lt;1% to &gt;30%), with the secretion of more cytokines, compared to iNKT cells from other tissues. Recently, “many pivotal roles” played by iNKT cells in asthma pathogenesis have been reported. Direct activation of pig lung iNKT cells using α-GalCer resulted in airway hyperreactivity (AHR), and clinically those pigs had dyspnea with elevated body temperature. The AHR induced pig lung had petechial hemorrhages with increased mucus secretion in the airway. The α-GalCer received pigs had three-fold increase in cytokines, IL-6, IL-12 and IL-10, and two-fold increase in CD1d expression on the dendritic cells and macrophages at post-24 of inoculation. A moderate increase in iNKT cells in lungs of AHR induced pigs was also detected, compared to controls. This work is supported by USDA and NPB grants to RJG.
Helios is a Marker for Human Thymic-derived FOXP3+ Regulatory T Cells (143.1)
Authors: Tran Dat , Thornton Angela , Shevach Ethan 
Published: January 2023

Abstract
RATIONALE: FOXP3+ regulatory T cells (Tregs) are central to the maintenance of self-tolerance and the prevention of autoimmunity. Since they are generated both in the thymus and the periphery, the ability to distinguish these two populations would be critical to understand their development, particularly in autoimmune and inflammatory conditions. METHODS: Peripheral blood of healthy adult donors and patients with systemic lupus erythematosus as well as cord blood were analyzed for FOXP3 and Helios expression within the CD4+ T cell population by flow cytometry. Cytokine secretion and suppressive function were studied in the Tregs and correlated with Helios expression. RESULTS: Helios is expressed selectively in the FOXP3+ Tregs and is decreased with age. Helios identifies thymic-derived, cytokine non-producing FOXP3+ Tregs and not peripheral or TGFbeta-induced FOXP3+ T cells. Knockdown of Helios with siRNA does not alter the cytokine profile and suppressor function of Tregs, suggesting that Helios is not critical for their anergic and suppressive properties. CONCLUSIONS: Helios is expressed uniquely in human FOXP3+ Tregs and distinguishes the thymic over the peripheral derived Tregs. While it is expressed predominately in the Tregs, it does not appear to play a critical role in their anergic and suppressive functions.
Exclusion of cytoplasmic fragments in flow cytometric analysis of lymphoma samples using DyeCycle Violet (144.1)
Authors: Modiano Jaime , Ito Daisuke , O’Brien Timothy 
Published: January 2023

Abstract
Cytoplasmic fragments that originate from fragile lymphoid cells and are commonly seen in lymphoma fine needle aspirate samples. These cytoplasmic fragments can interfere with accurate gating of target cells and quantification protocols used for flow cytometry because of their variable size and expression of lymphoid cell surface antigens on their membrane. The aim of this study was to develop a method to efficiently exclude cytoplasmic fragments in lymphoma samples analyzed by flow cytometry. Tumor cells prepared from lymph node biopsy and fine needle aspirate samples of dogs with lymphoma were stained using DyeCycle Violet (DCV, a membrane-permeable DNA-binding fluorescent dye), antibodies against cell surface antigens, and 7-amino-actinomycin D (7-AAD, a membrane impermeable, DNA-binding fluorescent dye). Multi-parameter flow cytometry was used to exclude antigen-positive cytoplasmic fragments for analysis based on the selective uptake and fluorescence of these dyes. Both cytoplasmic fragments, which are DCV-negative and CD45-positive, and dead cells, which are 7-AAD-positive, could be efficiently separated from tumor cells in lymphoma samples. DCV staining is a useful method to improve the gate setting and quantitative analyses of lymphoma samples by flow cytometry. DI and JFM contributed equally to this work
An automatable device for high content chemotaxis assays in the presence of a stable gradient (144.13)
Authors: Vu Elizabeth , Meyvantsson Ivar , Worzella Tracy , Skoien Allyson , Echeverria Victoria , Lamers Casey , Echeverria Daniella , Hayes Steve 
Published: January 2023

Abstract
Current approaches to study chemotaxis are limited by gradient instability, low throughput, cumbersome manual manipulations, high cell number requirements, inability to acquire high-content data, and/or poor data quality and reproducibility. We present here a new microfluidics-based approach to study leukocyte chemotaxis. The device features strong miniaturization of the use of cells (&lt;12,000 cells per data point). The microfluidic channels in the device are arrayed in 96 well format and can be operated by standard liquid handling robotics or by handheld pipettors. Cells migrate along a surface in the presence of a stable gradient of chemoattractant that is maintained for at least three hours. We show the ability to obtain consistent dose response curves to chemoattractants as well as chemotactic inhibitors with the device. Because cells are microscopically accessible, the information content is high. Not only can the number of cells migrated be assessed, but also migration distances of individual cells, and even scoring of cell activation and polarization.
Bio-inspired controlled release for regulatory T cell recruitment in vivo (144.3)
Authors: Jhunjhunwala Siddharth , Raimondi Giorgio , Hall Sherri , Thorne Stephen , Thomson Angus , Little Steven 
Published: January 2023

Abstract
Increased frequency of regulatory T cells (Treg) in organs affected by autoimmunity and in transplants is associated with suppression of autoimmune reactions and graft rejection, respectively. However, to the best of our knowledge very few (if any) effective therapeutic strategies exist to increase local numbers of Treg. In this context, it is noteworthy that tumors evade the immune system through recruitment of Treg using the chemokine CCL22. Our hypothesis was that, if we can mimic this tumor evasion strategy by creating a biological gradient of CCL22 at a local site using controlled release vehicles, we will be able to recruit Treg to that particular site. Here we report the fabrication of CCL22 containing microparticles using a degradable, biocompatible, FDA approved polymer. We programmed these CCL22 particles to release the chemokine continuously over a 3 week period, and designed them such that they do not move from the site of injection in vivo. Importantly, in a murine model for chemotaxis we are able to demonstrate recruitment of adoptively transferred Treg specifically towards the CCL22 microparticles using non-invasive live animal imaging. We envisage that this technology could potentially be useful for the suppression of adverse inflammatory reactions associated with autoimmunity and transplant rejection; a therapeutic application that we are currently testing.
Macrophages Fascilate Chronic Cardiac Rejection through a Monocyte-Induced by IFNγ (MIG) Mediated Process (145.14)
Authors: Moffatt-Bruce Susan , Bobek Daniel , Meyerson Joseph , Wang Jiao Jing , Hadley Gregg 
Published: January 2023

Abstract
Introduction Chronic rejection (CR) in cardiac transplantation leads to allograft dysfunction and transplant related heart failure. We investigated the role of macrophages-associated chemokines, monocyte-induced by IFNγ (MIG) and IFN-inducible protein 10 (IP-10) and their receptor CXCR3, in chronic cardiac allograft rejection. Methods BALB/c (H-2d) hearts were transplanted into C57BL6 (H-2b) recipients (n=6). Group AR had no treatment. Group CR had 3 mg i.p. of GK1.5 (antiCD4). The hearts were analyzed using qRTPCR and IHC. The spleens underwent FACS analysis. CXCR3 transgenic(Tg) and CXCR3KO mice were transplanted with BALB/c hearts. Results In group AR, hearts rejected at day 7. In group CR, the functioning hearts were harvested at day 30. Using ICH group CR allografts had significantly more macrophages compared to both AR allografts and isografts. FACS analysis of splenocytes in group CR detected macrophages to be more abdundant compared to group AR (p&lt;0.05). qRTPCR analysis detected a significant increase in allograft MIG and IP-10 in the group AR,(p&lt;0.05)but only MIG was significantly upregulated in group CR relative to both group AR and isografts(p&lt;0.05). CXCR3Tg and CXCR3KO transplants rejected at day 7. Conclusions Chronic cardiac allograft rejection may be a macrophage dependent process, potentially mediated through MIG. However, our results suggest that MIG and its receptor are likely to only be one component of a highly complicated process.