bluuebunny/crossref_metadata_2025_split huggingface dataset:
Granzyme B Is Critical for T-Cell Receptor Activation Induced Cell Death of Type 2 Helper T Cells (87.34)
Authors: Devadas Satish , Das Jyoti , Liu Catherine , Zhang Liying , Roberts Arthur I. , Pan Zui , Moore Paul A. , Das Gobardhan , Shi Yufang 
Published: January 2023

Abstract
Although CD95L is required for T cell receptor activation induced cell death (AICD) in T helper 1 (Th1) cells, the molecular mechanisms mediating AICD in Th2 cells are unknown. We found that death receptor medicated apoptosis was not involved in AICD of Th2 cells because blocking their cognate ligands with recombinant soluble receptors of specific antibodies had no effect on apoptosis of activated Th2 cells. Furthermore, we showed that the major caspases were not actively involved in AICD of Th2 cells. However, inhibition of granzyme B (Gzmb) specifically abolished AICD in Th2 cells but not Th1 cells. Likewise, Th2 cells derived from Gzmb-deficient mice were resistant to AICD, and Gzmb deficiency or inhibition of Gzmb activity consequently enhanced the production of Th2 cytokines. Most interestingly, Gzmb-deficient mice exhibited increased susceptibility to allergen-induced asthma. Thus, Gzmb plays a critical role in the AICD of Th2 cells. This finding provides a novel mechanism for the balance between Th1 and Th2 cells during immune responses.
LAT Plays Important Role in Thymocyte Survival and Mature T Cell Activation (87.35)
Authors: Shen Shudan , Zhu Minghua , Zhang Weiguo 
Published: January 2023

Abstract
LAT (Linker for Activation of T-cells) is a membrane-associated adaptor protein that plays a critical role in linking T-cell receptor engagement to activation of its downstream signaling events. Upon tyrosine phosphorylation, LAT binds Grb2, Gads, and PLC-gamma1. Previous studies show that LAT is essential in thymocyte development as LAT-deficient mice completely lack mature T cells. In addition, LAT-deficient Jurkat cells have a severe defect in TCR-mediated signaling. In order to investigate the role of LAT in T cell activation, survival, and homeostasis, mice in which the LAT gene can be inducibly deleted from thymocytes and mature T cells by injection of tamoxifen were generated. Our data showed that tamoxifen treatment of these mice led to a dramatic loss of thymocytes, especially DP thymocytes. Deletion of LAT in mature T cells affected TCR-mediated signaling, IL-2 production, and proliferation. These data suggested that LAT plays an important role in thymocyte survival and TCR-mediated signaling in mature T cells.
Participation of nitric oxide synthase (NOS) and atypical protein kinase C (PKC) ζ in thyroid hormone-mediated apoptosis of murine T lymphocytes (87.39)
Authors: Cremaschi Graciela Alicia , Klecha Alicia Juana , Vercelli Claudia , Franchi Ana María , Genaro Ana María , Arcos María Laura Barreiro 
Published: January 2023

Abstract
Thyroid hormones (TH) regulate immune responses and are able to influence proliferation and differentiation in several cell types. However, their direct actions on lymphocytes have not been elucidated. We have demonstrated that short time culture (up to 72 h) of quiescent BW5147 (BW) T cells in the presence of TH induced cell division. Here, prolonged actions of TH on BW cells and the intracellular signals involved were studied. After 5 days of culture, TH inhibited BW growth inducing apoptosis, as shown by Hoescht staining and DNA ladder assay. These effects were accompanied by an increase in NOS activity measured by [14C]-citrulline formation. Also, an increase in iNOS and a decrease in the PKC ζ isoform, both at the protein (by western blot analysis) and mRNA levels (by RT-PCR evaluation) were seen An increase in reactive oxygen species (determined by conversion of dichlorofluorescin-diacetate to the fluorescent DCF) was found. Additionally, TH induced an increment in protein nitrosylation.
These results show that prolonged TH exposition lead to an exacerbate increase in iNOS activity, that would in turn result in PKC ζ decrease, probably through protein nitrosylation. These intracellular signals would trigger apoptotic mechanisms involved in TH regulation of lymphocyte activity.
Importance of siRNA negative control selection: Evaluation of non-specific protein knock-down by negative control siRNA (87.42)
Authors: Sullivan Yvonne B. , Hughes Douglas E. , Narahari Janaki , Webb Brian L. 
Published: January 2023

Abstract
Small interfering RNA (siRNA) are double-stranded RNA consisting of 19–22 base-pairs that direct the cleavage of complementary mRNA targets. The utility of siRNA as a research tool depends on the specificity of the gene silencing, as knockdown of unintended gene targets (off-target effects) can potentially complicate the interpretation of siRNA data. To investigate the extent of off-target effects at the protein level, we evaluated RNAi negative control products from three commercial vendors for non-specific protein knockdown utilizing the SuperSignal® siRNA/Antibody Modules and SuperSignal siRNA Chemiluminescent Detection Module. Using Western blotting we found that the expression level of a number of different proteins was reduced when A549 cells were transfected with negative control siRNAs from two different vendors. Minimal non-specific protein knockdown was detected with the ON-TARGETplus™ siCONTROL® Non-Targeting Pool. These data suggest that the selection of an optimal negative control is critical to correct interpretation of siRNA experiments and that non-specific knockdown of protein should be evaluated.
Synergy Between Molecular targets of Costimulation in Promoting Persistence of CD8 T cells (87.5)
Authors: Song Jianxun , Song Ava , Croft Michael 
Published: January 2023

Abstract
Costimulation regulates multiple cellular processes including proliferation, expansion and survival of T cells, and molecular targets of costimulation might be useful to augment T cell activities. Two defined targets of costimulatory signals in CD4 T cells are the anti-apoptotic Bcl-2 family molecule Bcl-xL, and survivin, an Inhibitor of Apoptosis family member that might regulate cell division and survival. However, the relative importance and relationship between these molecules in CD8 T cells is not clear. To understand whether they have overlapping or synergistic functions, we used retroviral transduction to introduce Bcl-xL and survivin separately, or together linked by a 2A picornavirus self-cleaving peptide, into antigen-responding CD8 cells. In vitro, we found that CD8 effector T cells expressing both Bcl-xL and survivin strongly expanded at an early stage and had a long-term survival advantage. In vivo, with response to tumor-expressed Ag following adoptive T cell transfer, antigen-reactive CD8 T cells expressing both Bcl-xL and survivin displayed a greatly enhanced tumor protective activity compared CD8 cells expressing either molecule introduced separately. These results indicate that Bcl-xL and survivin can critically contribute in a synergistic, non-redundant, manner to augment the accumulation and persistence of CD8 T cells following encounter with antigen. The data provide new insights into why costimulatory signals might need to be sustained over time or provided periodically via ligation of multiple receptors.
Counting agonist in functional T cell receptor clusters using nanopatterned corrals (87.51)
Authors: Rossenova Boryana , Jackson Bryan , Dustin Michael , Groves Jay T 
Published: January 2023

Abstract
T lymphocytes, the initiators of the adaptive immune response, are remarkable for their ability to recognize very few agonist peptides presented on APC, using their T cell receptors (TCR). Recent studies emphasize the role of spatial organization of TCR at the interface between the two cells in regulating T cell activation - a central cluster and peripheral microclusters. The microclusters are thought to initiate, amplify and maintain signaling, however their mechanism of function is largely unknown. Using nanopatterned square corrals, 1 micrometer in length, we can regulate the maximum number of agonist-MHC molecules per cluster presented by a fluid supported lipid bilayer containing MHC and ICAM-1 molecules. We find that at activating mean concentrations of agonist, T lymphocytes are unable to initiate signaling if the maximal number of agonist molecules per corral is 1–2. Surprisingly, agonist number as little as twice this amount per corral, results in full cell activation, as measured by the calcium flux inside the cell. These observations suggest that microclusters are not necessary to amplify signaling beyond what 3–4 agonist molecules are sufficient to do.
Protracted Contraction and Memory T Cell Differentiation in Mice Deficient in TNF and Perforin (87.6)
Authors: Schmidt Nathan W. , Harty John T. 
Published: January 2023

Abstract
Following infection, pathogen-specific T cells undergo an expansion in numbers followed by contraction to relatively stable memory numbers. Work from others suggests that CD8 T cell responses to infection are altered in mice lacking TNF receptors or perforin. To understand how TNF regulates T cell responses we infected perforin/TNF-double deficient (DKO) mice, perforin-deficient (PKO) mice, and TNF-deficient (TKO) mice with Listeria monocytogenes (LM) and analyzed the MHC Ia-, MHC Ib-, and MHC II-restricted T cell responses. DKO mice exhibited a delay in contraction of MHC Ia- and MHC II-restricted T cells compared to WT mice. All three T cell responses exhibited delayed acquisition of CD62L, a central memory marker, in both DKO and TKO mice. DKO mice have a prolonged infection of LM, however, this did not account for the delay in contraction of the T cell populations, which still occurred in mice treated with antibiotics to eliminate the prolonged infection. These data suggest that TNF and perforin regulate antigen-specific T cell homeostasis, while TNF regulates the phenotype of antigen-specific T cells. Ongoing studies are addressing the mechanisms underlying delayed contraction and whether additional memory characteristics are altered by TNF deficiency.
Pretreatment of CD138− B Lymphocytes with a Covalent Reactive Antigen Analog to Factor VIII Reduces Antibody Production in response to FVIII (88.14)
Authors: Smith Keri Csencsits , Planque Stephanie , Nishiyama Yasuhiro , Taguchi Hiroaki , Escobar Miguel , Paul Sudhir 
Published: January 2023

Abstract
B lymphocytes producing high titer antibodies to factor VIII (FVIII) are of major concern in the treatment of bleeding episodes in a subpopulation of patients with hemophilia A. We hypothesize that an electrophilic, covalent reactive analog of FVIII (FVIII-CRA) can induce antigen-specific tolerance by permanent engagement of B cell receptors. FVIII-CRA was prepared by derivitizing lysine side chains with phosphonate diester groups. These groups are predicted to bind enzyme-like nucleophilic residues located within the BCR. To test the effects of FVIII-CRA binding on B lymphocytes, we isolated CD138− splenocytes from FVIII-deficient mice (B6;129S4-F8tm1Kaz/J) immunized with FVIII. These putative memory B lymphocytes were then re-stimulated for 6 days with identical doses of FVIII or FVIII-CRA. The number of antibody secreting cells (ASC) produced in response to FVIII-CRA re-stimulation was significantly lower than FVIII re-stimulation (avg of 36 ASC vs 111 ASC, p = 0.03). In addition, cell viability following low dose FVIII-CRA treatment was decreased by ~ 22% when compared with identical doses of FVIII. These results suggested that FVIII-CRA may tolerize memory B lymphocytes. To confirm this, we pretreated CD138− splenocytes with FVIII-CRA and then added a stimulatory dose of FVIII to the cultures. Pretreatment with FVIII-CRA resulted in a significant decrease in the number of ASC produced in response to FVIII (by 17-fold compared to control pretreatment). These results suggest that FVIII-CRA may represent a viable means to induce antigen-specific B cell tolerance in hemophilia.
Costimulatory molecule-targeted immunotherapy of chronic graft-versus-host disease (88.17)
Authors: Kim Juyang , Kim Hye J. , Park Keunhee , Kim Jiyoung , Choi Hye Jeong , Yagita Hideo , Nam Seok H. , Cho Hong R. , Kwon Byungsuk 
Published: January 2023

Abstract
Chronic graft-versus-host disease (cGVHD) is an increasingly frequent complication of allogeneic stem cell transplantation. Current therapies for cGVHD reduce symptoms but are not cures. Using the B10.D2„³Balb/c (H-2d) minor histocompatibility antigen-mismatched model which reflects clinical and pathological symptoms of human cGVHD, we show that a single injection of an agonistic monoclonal antibody (mAb) to CD137, a member of tumor necrosis factor receptor superfamily, reverses skin fibrosis, ulceration and alopecia, ultimately improving general health conditions. The reversal is associated with markedly reduced CD4+ T cell cytokines and increased apoptosis of donor CD4+ T cells. The Fas pathway is required for the therapeutic effect of anti-CD137 mAb on cGVHD. Taken together, these data indicate that anti-CD137 mAb has a therapeutic effect on cGVHD by removing donor CD4+ T cells mediating cGVHD. Our study demonstrates an agonistic mAb specific for a costimulatory molecule as a possible target for therapeutic intervention in cGVHD.
Bovine apolipoprotein B-100 is a dominant immunogen in therapeutic cell populations cultured in FCS in mice and humans (88.29)
Authors: Sakamoto Norihisa , Tuji Kuzuhide , Muul Linda M , Lawler Ann M , Candotti Fabio , Metcalf Julia A , Tavel Jorge A , Lane H.Clifford , Urba Walter J , Fox Bernard A , Varki Ajit P , Lunney Joan K , Rosenberg Amy S 
Published: January 2023

Abstract
Recent studies have demonstrated that cell populations intended for therapeutic purposes that are cultured in heterologous animal products can acquire xenoantigens, potentially limiting their utility. In investigations of the immune response to murine ES cells, we found that a strong antibody response was generated after the second infusion. Both polyclonal and monoclonal antibody responses, derived from immunized mice, were found to be specific for bovine apolipoprotein B-100 which binds to abundant low density lipoprotein receptor on the cell surface and is internalized. Critically, we have shown that in the majority of patients administered three different types of cell-based therapies utilizing cells grown in FCS containing media, an antibody response to bovine apoB-100 develops following the second infusion and is the dominant specificity. The low level of engraftment of genetically modified autologous cells in an ADA-SCID patient with a strong bovine apoB response suggests that such antibodies may influence cell engraftment.
Regulation of Airway Tolerance and Antigen-specific Regulatory T cells by the Costimulatory Molecule OX40 (88.39)
Authors: Duan Wei , Croft Michael 
Published: January 2023

Abstract
Respiratory exposure to soluble antigen leads to airway tolerance and antigen-specific suppression of airway inflammation. There is increasing evidence that this involves inducible regulatory T cells (Treg) that suppress pathogenic Th2 cells. What controls Treg activity is not clear. We investigated the role of the costimulatory molecule OX40 in the generation and function of Foxp3+ Treg induced by soluble OVA, given either intravenously or intranasally. By tracking adoptively transferred OVA-specific CD4 cells, we observed that markedly fewer Foxp3+ cells were induced in the absence of OX40, suggesting OX40 can control their expansion and/or survival. Also, already generated OVA-specific CD4 Foxp3+ cells when adoptively transferred into sensitized mice, suppressed lung inflammation. This suppression was impaired if the Treg lacked OX40, further demonstrating that OX40 can positively control the function of this Treg subset. In contrast, we found that an OX40 agonist antibody reversed established airway tolerance when given with antigen immunization, implying that anti-OX40 either augmented Th2 cell expansion or inhibited Treg. Thus, OX40 signals can positively affect inducible Treg but they might equally control pathogenic T cell activity. These studies further substantiate the idea that OX40 is central to the activities of several subsets of T cells that control airway inflammation.
T cell response after TCR antagonism: A unique unresponsive CD4+ T cell phenotype (88.41)
Authors: Edwards Lindsay J , Evavold Brian D 
Published: January 2023

Abstract
The outcomes of T cell interaction with ligands of varying potency include the induction of clonal unresponsiveness (anergy) and inhibition of T cell responses (antagonism), among others. The phenotype of peptide-induced anergy has been extensively described, and a number of groups have demonstrated antagonism. However, the phenotype of CD4+ T cells after encountering an antagonistic stimulus has not been previously characterized, despite the fact that several pathogens, including HIV, HCV and malaria, include epitopes that have been defined as T cell antagonists. We demonstrate that post-antagonism, T cells fail to proliferate. However, these cells produce large quantities of IL-2 and express CD25 levels similar to wild type stimulated cells. Our data indicate that antagonized CD4+ T cells fail to respond to IL-2 stimulation, despite the production of this cytokine, as well as expression of its receptor. This response represents a novel phenotype, distinct from all previously described forms of anergy.
This research was supported by NIAID (R01 AI056017).
Class II MHC Deficient Mice Have Contact Hypersensitivity Responses that Are Regulated by CD4+CD25+ T Cells (88.5)
Authors: Kish Danielle D. , Gorbachev Anton V , Fairchild Robert L 
Published: January 2023

Abstract
CD4+CD25+ T cells limit the expansion of effector CD8 T cells in skin draining lymph nodes (LN) during hapten sensitization for contact hypersensitivity (CHS) responses. This regulatory function results in decreased swelling responses to hapten challenge of the skin. Depletion of CD4 T cells prior to hapten sensitization results in increased numbers of hapten-primed effector CD8 T cells and CHS responses that are elevated in magnitude and extended in duration. These increased CHS responses are similar to those observed in sensitized and challenged class II MHC−/− mice. However, depletion of CD4 T cells in class II MHC−/− mice increased CHS responses to hapten sensitization and challenge when compared to untreated class II MHC−/− mice. ELISPOT analyses indicated increased hapten-specific CD8 T cell priming in LN of class II MHC−/− mice compared to the wild-type and these numbers increased significantly with anti-CD4 or anti-CD25 mAb treatment prior to sensitization. Flow cytometry analyses of LN cells from class II MHC−/− mice revealed a small but distinct population of CD4T cells and ≥50% expressed CD25. qRT-PCR analyses of RNA prepared from LN cells of class II MHC−/− and wild-type mice indicated the presence of FoxP3 expression that was absent when the animals were depleted of CD4 or CD25, but not NK1.1, cells. Purified CD4+ CD25+ T cells from class II MHC−/− and wild-type mice were equivalent in the ability to limit CD4+CD25− T cell proliferation in vitro. These results identify a CD4+ CD25+ regulatory T cell population in class II MHC−/− mice.
CD11c is Critical in Modulating Cytokine Production by Activated T cells (88.6)
Authors: Adams Jillian E , Barnum Scott 
Published: January 2023

Abstract
It is well established that CD11a (a.k.a. LFA-1) functions in T cell activation by providing a co-stimulatory signal and by participating in the formation of the immunological synapse. The role of other β2-integrins in T cell activation remains unknown. Recent EAE studies have suggested a role for CD11b and CD11c in T cell biology. These findings prompted us to simultaneously stimulate naïve T cells in vitro, using anti-CD3ε and anti-CD11c antibodies. We found that treatment of T cells with anti-CD3ε and anti-CD11c (clones N418 or HL3) antibodies substantially suppressed IFN-γ production at 72 hours. T cells treated with anti-CD3ε and anti-CD11c antibodies do not produce detectable amounts of IL-4 and IL-10 at 12, 24, and 48 hours. In these same cultures, IFN-γ production was only detected at 12h post-stimulation, but at a concentration 3 logs lower than that observed with anti-CD3ε treatment alone. Interestingly, TGF-β1 production was increased 3 fold on treatment with anti-CD11c and anti-CD3ε compared to anti-CD3ε treatment alone. The suppression of IL-4, IL-10, and IFN-γ production could not be rescued by co-stimulation with anti-CD28 antibodies. Our data suggest that CD11c may be a novel negative regulator in T cell activation.
Supported by NH46032 from the NIH.
SHPS-1 positively regulates T cell priming and cytokine production by dendritic cells (DCs). (89.15)
Authors: Tomizawa Takeshi , Kaneko Yoriaki , Okajo Jun , Okuzawa Chie , Saito Yasuyuki , Sekigami Tomomi , Kaneko Yuka , Matozaki Takashi , Nojima Yoshihisa 
Published: January 2023

Abstract
[Background] SHPS-1, a transmembrane protein that binds protein tyrosine phosphatases in its cytoplasmic region, is predominantly expressed on DCs, but the role of SHPS-1 signals during T cell priming by DCs is largely unknown. Here, we investigated the physiological roles of SHPS-1 in the functions of DCs by the use of SHPS-1 mutant mice, which lack most of the cytoplasmic region of this protein.
[Methods] DCs were prepared from spleens by Auto MACS. The expression of SHPS-1 or costimulatory molecules on DCs were examined by FCM. IL-12-induced IFN-γ production by DCs was determined by ELISA. The ability of DCs for antigen presentation was evaluated by the incubation of OT-II cells with OVA-loaded DCs.
[Results] FCM analysis showed that SHPS-1 was predominantily expressed on CD8α− DCs. The expression level of SHPS-1 was reduced in DCs from SHPS-1 mutant mice, but the expression levels of costimulatory molecules on DCs from SHPS-1 mutant mice were almost comparable to that on WT DCs. On the contrary, IL-12-induced production of IFN-γ was reduced in DCs from SHPS-1 mutant mice. The ability of DCs to stimulate proliferation of OVA-specific CD4+ T cells was also impaired in DCs from SHPS-1 mutant mice.
[Conclusion] SHPS-1 positively regulates DC functions including CD4+ T cell priming and cytokine production.
Cell Specific Inactivation of Multiple TLRs via Conditional Deletion of Their Master Chaperone gp96/grp94 (89.16)
Authors: Yang Yi , Li Zihai 
Published: January 2023

Abstract
gp96/grp94 is the endoplasmic reticulum (ER) resident protein chaperone of the HSP90 family. In contrast to its cytosolic counterpart HSP90, very few client proteins and yet no cochaperones of gp96 have been uncovered. To study the function of gp96 in vivo, we generated a gp96 conditional knockout mouse model. By crossing with a macrophage-specific Cre recombinase transgenic mouse, we successfully deleted gp96 in macrophages, but not in any other cell types, including dendritic cells (DCs). We found that the mutant macrophages failed to respond to multiple Toll-like receptor (TLR) ligands, including TLR2, TLR4, TLR5, TLR7 and TLR9. Since gp96 is indispensable for TLRs, the gp96 conditional knock-out mouse model provides a unique system to examine the contribution by TLR in inflammation and immune response in vivo. We found that macrophage-specific gp96 knockout mice were resistant to endotoxin shock, but highly susceptible to Listeria monocytogenes infection, demonstrating that macrophages but not other myeloid cells are the dominant source of proinflammatory cytokines in these conditions. Our finding is significant in that it not only confirmed a previous report that gp96 is required for cell surface expression of certain TLRs, but also for the first time revealed that gp96 is required by those TLRs that normally reside in the endocytic compartments. We conclude that the Toll-like receptor family members are delicately regulated by gp96 as their master chaperone.