bluuebunny/crossref_metadata_2025_split huggingface dataset:
Full-length IL-33 acts differently from mature IL-33 in vivo partially in an ST2-independent fashion (120.30)
Authors: Atamas Sergei , Pickering Edward , Kopach Pavel , Kang Phillip , Lockatell Virginia , Papadimitriou John , McKenzie Andrew , Luzina Irina 
Published: January 2023

Abstract
IL-33 is a key regulator of inflammation and immunity. It remains controversial whether protease-mediated activation of IL-33 is needed for functional effects. We constructed and validated recombinant adenoviruses for gene delivery of full-length mouse (flm) and mature mouse (mm) (aa 109-266) IL-33 to mouse lungs in vivo. Gene expression was confirmed by RT-Q-PCR and ELISA. Analyses of BAL samples and lung tissues revealed substantial differences between flmIL-33 and mmIL-33. Both isoforms caused pulmonary infiltration and BAL influx of T and B lymphocytes and neutrophils, whereas mmIL-33 also caused significant pulmonary eosinophilia (~47-55% of BAL cell count) and hyperplasia of mucus-producing goblet cells. Multiplex analyses showed that mmIL-33 caused significant increases in IL-4, IL-5, IL-13, IL-17, and KC; flmIL-33 tended to stimulate IFN-γ; and both isoforms induced MCP-1 with a greater increase induced by mmIL-33. Subsequent analyses were performed in mice deficient of ST2 (IL-33 receptor). ST2 deficiency completely abrogated mmIL-33-induced pulmonary eosinophilia, goblet cell hyperplasia, and elevations in IL-4 and IL-5. However, lymphocytic infiltration induced by flmIL-33 or mmIL-33 was attenuated but persistent in the absence of ST2. These data suggest that full-length IL-33 is independently functionally active in vivo, in part in an ST2-independent fashion, and that Th2 effects but not lymphocytosis induced by mature IL-33 are ST2-dependent.
Resolvins attenuate cigarette smoke-induced lung inflammation and injury (120.31)
Authors: Phipps Richard , Hsiao HsiMin , Thatcher Thomas , Vickery Thadeus , Serhan Charles 
Published: January 2023

Abstract
Cigarette smoke induced chronic obstructive pulmonary disease (COPD) is a major global health issue with an increasing incidence worldwide. Inflammation plays an integral role in the development of chronic lung diseases such as COPD. Unresolved inflammatory responses in COPD can lead to tissue damage and dysfunction. Current therapies for COPD only treat symptoms and do not alter the course of the disease. Resolution of inflammatory responses was once believed to be passive, however, recent studies have shown that resolution is a biosynthetically active process involving endogenous pro-resolving lipid mediators, derived from omega-3 and omega-6 polyunsaturated fatty acids that control and limit inflammatory responses. Resolvin D1 (RvD1) is an endogenous lipid mediator that displays potent anti-inflammatory and pro-resolving actions without being immunosuppressive. We hypothesized that RvD1 would be beneficial in a smoke-induced lung injury model. Cigarette smoke-exposed mice treated with RvD1 exhibited a greater than 50% reduction in neutrophil accumulation in the lung lavage fluid, coupled with significant reductions in the neutrophil chemoattractants KC and MIP-2. IL-6 and IL-1β were also reduced. RvD1 treated mice also had enhanced phagocytosis of apoptotic neutrophils by macrophages. Interestingly, RvD1 drives polarization of alternatively activated macrophages (M2) that may facilitate resolution of inflammation and attenuate tissue damage.
Activated Adventitial Pulmonary Artery Fibroblasts Drive Macrophages Toward a Distinct M2-like Phenotype through STAT3 Signaling in Pulmonary Hypertension (120.36)
Authors: El Kasmi Karim , Riddle Suzette , Anderson Aimee , Flockton Amanda , Noe Megan , McKeon Alexandre , Frid Maria , Sokol Ronald , Stenmark Kurt 
Published: January 2023

Abstract
We have previously documented that in the adventitia of distal pulmonary arteries (dPA) in calves with pulmonary hypertension (PH) macrophages that express CD163 (marker of alternative activation, AAM) accumulate in proximity to adventitial fibroblasts (adFibs) in areas of remodeling. AAMs have been implicated in promoting remodeling in PH in response to STAT6 signaling. Our recently published data suggest that in the dPA, adFibs express an epigenetically stable pro-inflammatory phenotype capable of activating macrophages. We hypothesized that adFibs activate AAMs through STAT3 but not STAT6. adFibs were isolated and cultured from the dPA of hypertensive (PH-fibs) and control (CO-fibs) calves and used to generate conditioned media (CM). Bovine, murine and rat bone marrow derived macrophages (BMDMs) were incubated in CM for 16hrs followed by qRT-PCR analysis. In response to PH-fibs CM, bovine BMDMs expressed Cd163, Cd206, and IL4ra consistent with AAMs. Furthermore, in response to PH-Fib but not CO-Fib CM both murine and rat BMDMs expressed Cd206, Cd204, Irf4, Klf, jmjd3; STAT3-dependent genes Socs3, Cebp/b, Pim1, Nfatc2, Il4ra, Arg1, but not STAT6-dependent genes Chi3I3 and Retnla1; phospho-STAT3 but not phospho-STAT6; and mRNA for Arg1 (previously thought to be STAT6 dependent) in murine Stat6-/- BMDMs. These results define the activated dPA adFibs as a key locus of paracrine signaling that triggers STAT3 in macrophages, which then express a distinct AAM phenotype.
Modulatory effects of the tumor suppressor p53 in T cell activation and function (121.13)
Authors: Guo Gang , Nguyen Doan , Zhang Shu-Zhong , Huang Yunping , Ramsay Alistair , Ochoa Augusto , Cui Yan 
Published: January 2023

Abstract
Compelling evidence demonstrates that the tumor suppressor p53 plays important roles in regulating physiological functions, such as stem cell state, development, tissue homeostasis, inflammation, metabolism, and aging. Early immunological studies showed that p53 is rapidly activated upon TCR stimulation. Nevertheless, the mechanism that underlies the TCR-mediated p53 activation is unclear. Recently, we reported that p53 negatively regulates Th17 differentiation. Here, we further examined the function of p53 during non-polarizing T cell activation. Upon TCR stimulation (α-CD3+ α-CD28), especially under suboptimal conditions (low level of α-CD3), p53null T cells proliferated more aggressively with a higher effector function than their WT counterparts. To investigate the molecular mechanism by which p53 modulates T helper differentiation, we examined their gene expression via microarray. Among the 45,000 probe sets hybridized, 4025 genes within the Gene Ontology (GO) pathways of immune responses (innate immunity, IL-1β, IL-6, and IL-17 pathways), DNA damage responses, Cell cycle regulation, apoptosis, and survival showed significant differences (p &lt; 0.05) between activated WT and p53null CD4 cells. We are in the process of confirming the differential expression among some of these crucial factors at the protein level and validating their cellular manifestation during T cell activation. Overall, these results support our hypothesis that p53 negatively modulates T cell responses.
The role of TNF receptor superfamily members in thymic Treg development (121.23)
Authors: Mahmud Shawn , Hogquist Kristin , Farrar Michael 
Published: January 2023

Abstract
CD4+FOXP3+ regulatory T cells (Tregs) express several TNF receptor superfamily (TNFRSF) members. However, it is unclear what role these molecules play in Treg development in the thymus. We found that CD4+CD25+FOXP3- Treg progenitors and CD4+CD25+FOXP3+ thymic Tregs, but not CD4SP conventional thymocytes, express high levels of OX40, GITR, and TNFR2. Utilizing a novel Nur77-GFP Bac reporter mouse in which thymocyte GFP level is proportional to TCR signal strength, we observed that OX40 and GITR expression by Tregs positively correlates with Nur77-GFP expression; this suggests that those thymocytes perceiving the strongest TCR signals during development are engendered with the highest level of expression of these TNFRSF members. Importantly, the ligands for these receptors are expressed on APCs in the thymus. To identify the functional role these TNFRSF members play in thymic Treg development, we made mixed bone marrow chimeras with WT and Gitr-/- bone marrow. Gitr-/- donor cells gave rise to normal numbers of CD4+CD25+FOXP3- Treg progenitors but exhibited an ~30% decrease in FOXP3+ mature thymic Tregs relative to WT donor cells. Furthermore, in vitro stimulation of Treg progenitors with GITRL or OX40L leads to a marked enhancement of Treg differentiation. We are presently investigating whether GITR, OX40 and TNFR2 play partially redundant roles in driving thymic Treg development by generating Ox40-/- x Gitr-/- mice in conjunction with neutralizing antibodies to TNFR2.
A Kinase/Phosphatase RNAi screen to identify genes required for PD-1-mediated inhibition of TCR activation (121.3)
Authors: Fonseca Catia , Imam Sabrina , Alexe Gabriela , Piccioni Federica , Root David , Haining William 
Published: January 2023

Abstract
Signaling via PD-1 negatively regulates T cell activation and plays a central role in the state of T cell dysfunction known as exhaustion. However the signaling mechanisms by which PD-1 inhibits TCR activation are not fully understood. Because protein phosphorylation/dephosphorylation regulates TCR signaling, we screened an shRNA library targeting all human kinases and phosphatases to determine which were required for PD-1 inhibition. To model PD-1 inhibition of TCR stimulation we used Jurkat cells engineered to expressed high levels of PD1; ligation of PD-1 by bead-bound antibodies inhibited CD3/CD28-induced IL2 secretion in a dose-dependent fashion. We screened a lentiviral-based arrayed shRNA library of 518 kinases and 150 phosphatases for genes that could rescue PD-1-mediated inhibition of IL2 expression (measured by multiplex qPCR). We identified 25 kinases or phosphatases for which knockdown by two or more shRNAs increased IL2 expression in PD-1-inhibited cells but not in unstimulated cells. Nine of 25 hits were validated in multiple cell lines and secondary assays.Hits included known negative regulators of T cell activation, suggesting our screen recovered expected gene targets. This study uncovered new potential negative regulators of TCR signaling mediated by PD-1 and can be adapted to identify signaling intermediates downstream of other inhibitory receptors.
Molecular signature of human CD8+ T cell with different polyfunctionality in response to different antigen dose (121.8)
Authors: Chiu Yen-Ling , Mathias Oelke , Schneck Jonathan 
Published: January 2023

Abstract
T cell polyfunctionality has been shown to be crucial in protective immunity. However, the molecular mechanism controlling CTL polyfunctionality have not yet been determined. Specifically the genetic control linking antigen doses to polyfunctionality has not been explored. In this study, we found that human antigen specific CTL expansion induced by high dose antigenic stimulation is associated with CTL with lower polyfunctionality and higher inhibitory receptors expression, while low does antigen stimulation results in higher T cell polyfunctionality and lower level of inhibitory receptors. Notably, we found that even in an artificial APC system with only signal 1 and signal 2, antigen dose still directly mediated T cell polyfunctionality independent of any inhibitory receptor signaling. The molecular signature associated with optimal CTL polyfunctionality is identified by genomic microarray and these findings are thus important framework for further studies on how to preferentially modulate T cell polyfunctionality.
Chemokine Receptor CCR7 Does Not Play an Appreciable Role in the Trafficking of Central Memory CD4 T Cells to Lymph Nodes. (122.1)
Authors: Campbell James , Vander Lugt Bryan , Tubo Noah , Boes Marianne , Malissen Bernard , Fuhlbrigge Robert , Kupper Thomas 
Published: January 2023

Abstract
CCR7(-/-) mice exhibit profound anomalies in the architecture of their secondary lymphoid organs, which complicates the study of CCR7-mediated T cell trafficking in these mice. To circumvent this confounding variable, we established in vivo models that force WT and CCR7(-/-) populations to compete for access to various tissues within mice possessing otherwise normal lymphoid organs. We found CCR7(-/-) naïve T cells to be profoundly reduced within lymph nodes with respect to their WT counterparts, a reduction unrelated to effects of CCR7 on thymic development. Strikingly, we found the entry of memory CD4 T cells into lymph nodes from the blood to be entirely autonomous of CCR7, calling into question the function of CCR7 molecules expressed on the surface of these cells. Contrary to previous studies, we found that CCR7 deficiency had no effect on the exit of CD4 T cells from inflamed skin. CCR7(-/-) CD4 T cells were inadequately imprinted for development into skin-selective E-lig(+) cells during primary cutaneous responses, most likely due to the inability of CCR7(-/-) naïve T cells to enter peripheral lymph nodes.
Human Th17 cells have stem cell-like features and promote long-term tumor immunity (122.10)
Authors: Kryczek Ilona , Zhao Ende , Liu Yan , Wang Yin , Vatan Linda , Szeliga Wojciech , Klimczak Aleksandra , Lange Andrzej , Zou Wiping 
Published: January 2023

Abstract
Th17 cells play a role in multiple pathological scenarios. However, their functional phenotype, self-renewal potential, survival pattern and the underlying controlling mechanisms are poorly investigated. Now we have studied Th17 cells in the pathological microenvironments of GVHD, ulcerative colitis and cancer in humans. Th17 cells are increased in the chronic phase of these diseases. Despite their phenotypic markers of terminal differentiation, in vivo adoptive transfusing experiments demonstrate that Th17 cells mediate long-term anti-tumor immunity. Th17 cells have stem cell-like features including high capacity of proliferative self-renewal, potent persistence and apoptotic resistance in vivo, and the generation of other types of T helper cells and express high levels of multiple core stem cell genes and anti-apoptotic genes. We demonstrate that their stem cell-like characters are regulated collaboratively by HIF-1α and Notch. Our data indicate that human Th17 cells may be a long-lived proliferating T cell population with stem cell characters.
Functional significance of Syntaxin 8 in Cytotoxic T Lymphocyte cytotoxicity (122.3)
Authors: Bhat Shruthi , Qu Bin , Kummerow Carsten , Rettig Jens , Schwarz Eva , Hoth Markus 
Published: January 2023

Abstract
Cytotoxic T Lymphocytes (CTLs) are activated CD8+ T cells which identify and kill pathogen infected, tumor and nonself cells by FAS ligand expression and by secretion of perforin and granzymes from lytic granules specifically at the immunological synapse (IS). This prevents killing of healthy bystander cells. Therefore at the time of killing these cytotoxic molecules are to be delivered to the IS. SNAREs are membrane associated proteins involved in vesicle transport, docking and fusion. Mutations of these proteins can cause diseases like Familial Hemophagocytic Lymphohistiocytosis. We have shown previously that Vti1b is required for docking of lytic granules at the IS. We also found that Vti1b and Syntaxin 8 (Stx8) colocalize with perforin in human CTLs. Further experiments showed that Stx8 partially colocalized with Vti1b. To elucidate the role of SNAREs in CTL cytotoxicity, siRNA were used to downregulate their expression. Stx8 downregulation did not impair the IS formation as CD3 accumulation at the IS was unaffected. We have previously shown that the killing capacity of CTLs is impaired by Vti1b downregulation. Here, Stx8 downregulated CTLs used in real time killing assay showed reduction in killing. Cycloheximide treatment of control and Stx8 downregulated CTLs showed that Stx8 is required for killing by perforin secretion. This conclusion is also supported by our preliminary ELISA data, which shows reduced perforin secretion in Stx8 downregulated CTLs.
The effects of age on CD4+ T cell TLR-2 expression and TLR-2 mediated direct co-stimulation (122.6)
Authors: Lancioni Christina , Sinnott Brian , Null Megan , Lewinsohn David , Lewinsohn Deborah 
Published: January 2023

Abstract
Background: Functional Toll like receptors (TLR) are found on human T cells. Neonatal antigen presenting cells (APC) have altered responses to TLR ligands; it is unknown if neonatal CD4+ T cell TLR responses are comparable to adult. Objective: Comparison of TLR-2 expression and responsiveness to TLR-2 direct co-stimulation among total and naïve CD4+ T cell subsets from neonates and adults. Methods: T cell subsets (CD3+CD4+ or CD3+CD4+CD45RA+) were isolated from cord and adult peripheral blood mononuclear cells by flow sorting ( &gt;99.5% purity) and cultured in an APC-free model with α-CD3 ± α -CD28 ± TLR-2 ligand Pam3CYS4. Quantitative PCR and ELISA determined TLR-2 expression and cytokine production. Results: Neonatal CD4+ T cells are predominately naïve (97.2% CD45RA+) but express 5-fold more TLR-2 mRNA than adult CD4+ T cells. Pam3CYS4 provides direct co-stimulation in the presence of α-CD3 to adult and neonatal total CD4+ T cells resulting in IL-2, TNF-α, and IFN-γ production; provision of additional co-stimulation (α-CD28) further enhances cytokine production. When comparing total CD4+ T cell populations, neonatal cells generate significantly less cytokines than adult. When naïve neonatal and adult T cells are compared, neonatal cells generate more cytokines in response to TLR-2 mediated co-stimulation. Conclusions: Naïve neonatal CD4+ T cells have enhanced TLR-2 expression and increased sensitivity to direct TLR-2 co-stimulation as compared to naïve adult cells.
Mechanisms of age-associated T cell defects: With age naïve CD4 T cells become Th1 prone, make increased OPN and acquire epigenetic changes in methylation of Type I and 2 cytokine loci consistent with that profile (122.8)
Authors: Zhang Wenliang , Kuang Yi , Browne Eva , Arcaro Kathleen , Swain Susan 
Published: January 2023

Abstract
As animals age, their T cells develop defects that compromise their immune responses and prevent effective vaccination to new pathogens. Our research has shown that naïve CD4 T cells become progressively more defective with age. When they respond, they produce less IL-2 and expand less. This pattern is intensified in the memory cells to which they give rise, which expand poorly and produce little if any IL-2, and more Th1, but less Th2-polarized cytokines. The molecule mechanisms that cause these shifts are largely unknown. We find that during their 10 response, aged naïve CD4 cells produce more IFNγ and more of the Th1-polarizing cytokine osteopontin (OPN). We find that the IFNγ promoter region of the aged naïve CD4 T cells, has become hypomethylated, while promoters for IL-4 and GATA3 are hypermethylated, suggesting that epigenetic changes that modify chromatin accessibility may be responsible for the altered cytokine production during initial response. We are investigating whether the increased initial levels of IFNγ and OPN are responsible for programming the aged cells to become memory cells with restricted cytokine profiles. Our hypothesis is that signals that accumulate during the increased persistence of naïve T cells that occurs during aging, cause epigenetic changes in cytokine loci that in turn contribute to the development of age-associated functional defects.
The role of ICOS co-stimulation on CD8 T cell differentiation during infection. (122.9)
Authors: Mouriès Juliette , Yagita Hideo , Lefrançois Leo 
Published: January 2023

Abstract
Co-stimulatory molecules give critical signals to CD8 T cells during their priming phase to allow the generation of protective cytotoxic T cells (CTLs). In this study, we explored the role of ICOS on CTL differentiation by blocking ICOSL with an antibody in vivo. Using a bacterial and a viral infection models, we showed that despite normal expansion during ICOSL blockade, the generated CD8 T cells had lower granzyme expression, they preferentially differentiated more into memory precursor effector cells (CD127hi KLRG1low) and their cytokine production (IFN-γ, TNF-α, IL-2) was enhanced. Finally, CTLs generated in the absence of ICOS signaling at initial priming accumulated more at memory stage. Interestingly, the observed phenotype is very similar to Blimp-1 deficiency, a transcription factor involved in terminal differentiation, suggesting a link between ICOS and the Blimp-1/Bcl-6 axis. In fact, after ICOSL blockade, Bcl-6 expression was increased in CTLs. Bcl6 and Blimp-1 are known reciprocal and antagonistic regulators, indicating that Blimp-1 might be down regulated following ICOSL blockade. The molecular basis of this regulation is still under investigation. Finally, using mixed bone marrow chimera with cells from WT and ICOS KO mice, we were able to recapitulate the phenotype observed with ICOSL blockade, indicating that the direct triggering of ICOS on CD8 T cells is responsible for tuning the response.
Lupus-prone (NZBxNZW)F1 mice exhibit decreased ability to induce iTregs due to defects in both antigen presenting cells and T cells (123.1)
Authors: Chhabra Anita , Parnell Sarah , Alard Pascale , Kosiewicz Michele 
Published: January 2023

Abstract
Quantitative and qualitative defects in regulatory T cells (CD4+CD25+Foxp3+; Tregs) are associated with many autoimmune diseases, including systemic lupus erythematosus (SLE). SLE-prone (NZBxNZW)F1 (BWF1) mice have very low frequencies of CD4+Foxp3+ in lymphoid organs compared to non-autoimmune strains, and adoptive transfer of Tregs prevents SLE presumably by restoring the “proper” Treg:effector T cell ratios. In this study, we investigated the mechanisms that could affect Treg frequencies in BWF1 mice. There were no differences in either thymic production or peripheral proliferation of Tregs between BWF1 and control mice. However, in vitro induction of Treg conversion (iTregs) was defective using either BWF1 antigen presenting cells, i.e., TGFβ-treated bone marrow-derived dendritic cells (DC) or untreated CD103+ DC (DC able to induce conversion constitutively), or BWF1 T cells. Furthermore, BWF1 mesenteric lymph nodes contained decreased CD103+ DC concomitant with decreased Helios-CD4+Foxp3+ cells (a potential marker of iTregs) frequencies compared to controls. Taken together, these data suggest that defects in both DC and T cells in BWF1 mice lead to defective conversion of iTregs and may contribute to the decreased Treg frequencies, and consequently, to the development of disease. Identifying the mechanisms that cause these defects could lead to the development of therapeutic strategies that restore an optimal Treg compartment, and thereby treat lupus.