bluuebunny/crossref_metadata_2025_split huggingface dataset:
RhoH is a key functional determinant of the “stop and go” nature of CD4+ T cells during activation (109.17)
Authors: Baker Christina , Comrie William , Fedorchuk Christine , Hyun Young-Min , Meier-Schellersheim Martin , Kim Minsoo 
Published: January 2023

Abstract
CD4+ T cells spend the majority of their time perusing lymphoid organs in search of cognate antigen presented by APCs, and then cells quickly recirculate through the bloodstream to another lymph node. Therefore regulation of a CD4+ T cell response is dependent upon the ability of cells to arrive in the correct location following chemokine gradients (“go” signals) as well as to receive appropriate TCR activation signals upon cognate antigen recognition (“stop” signals). The mechanism(s) by which a CD4+ T cell regulates these “go” and “stop” signals are not well defined. We find that over-expression of the hematopoietic specific RhoH protein in the presence of chemokine signals results in decreased Rap1-GTP and LFA-1 adhesiveness to ICAM-1, thus impairing T cell chemotaxis, while in the presence of TCR signals there is enhanced and sustained Rap1-GTP and LFA-1 activation as well as prolonged T:APC conjugates. RhoH-/- CD4+ T cells appear to circulate through the blood and lymphoid tissue more quickly corroborating our in vitro migration data. RT-PCR analyses of activated CD4+ T cells and live images of T cell migration and immunological synapse formation have revealed that functions of RhoH take place primarily at the levels of transcription and intracellular distribution. Thus, we hypothesize that RhoH expression may provide a single molecular determinant that allows a CD4+ T cell to switch between sensing chemokine mediated “go” signals and TCR dependent “stop” signals.
PIM kinase activity is required for B cell function in vitro and in vivo (109.5)
Authors: Jackson Leila , Pheneger Jed , Wright A. , Robinson John , Allen Shelley , Hicken Eric , Carter Laura 
Published: January 2023

Abstract
The PIM family of serine/threonine kinases has roles in proliferation, differentiation, and cell survival. In lymphocytes, their activity is regulated by inducible expression following cytokine or antigen receptor signaling. Using small molecule inhibitors, we have examined the role of PIM1/3 kinases in B cells in vitro and in vivo and in the MRL/LPR model of lupus. PIM kinases were expressed 1-3 hours after B cell activation with anti-IgM or anti-CD40 but not BAFF. Inhibition of PIM-1/3 kinase activity decreased anti-IgM- and anti-CD40-induced B cell proliferation without inducing apoptosis. AR-254 also prevented IL-4-, and IFNy-induced B cell survival, but had no affect on BAFF-induced B cell survival consistent with the inability of BAFF to upregulate PIM expression. AR-770 administration in the HEL immunization model resulted in a 90% reduction of HEL-specific IgG2a and IgG3, but had no effect on IgG1 suggesting a role for PIM kinases in IFNy-mediated class switching. In the MRL/MpJ-Fas&lt;lpr&gt; lupus model, mice treated with AR-770 had &gt;90% reduction in proteinuria AUC, normalized anti-dsDNA antibody titers and ~80% decrease in kidney glomerulo- and interstitial nephritis as well as vessel and protein cast formation. Under specific conditions, Taken together, the data suggests that inhibition of PIM kinases may be a useful strategy for patients with B cell-mediated diseases such as lupus.
Role of C-reactive protein in regulatory monocyte induction and activation in trauma patients (110.18)
Authors: Mold Carolyn , West Sonlee , Du Clos Terry 
Published: January 2023

Abstract
C-reactive protein (CRP) is an acute phase reactant. Its plasma concentration increases from &lt;5 to &gt;100 µg/ml within 48 h of injury due to increased synthesis by hepatocytes responding to IL-6. Highly elevated CRP levels are seen in trauma, and maintenance of these levels is associated with a poor prognosis, but the role of CRP in trauma is poorly understood. We measured a panel of monocyte markers and cytokines in 27 trauma patients for 3 days following admission and analyzed their correlation with CRP levels. We identified a subpopulation of monocytes expressing high levels of CD14 and CD16 that were not found in healthy volunteers. These cells also expressed CD163 and HLA-DR. The percentage of monocytes found in this subpopulation was highly correlated to plasma CRP levels (p=0.0001) and to intracellular heme oxygenase (HO-1) (p=0.0009). CRP levels also correlated with monocyte HO-1, and plasma anti-inflammatory cytokines, IL-10, IL-1RA, and TGF-β. We hypothesized that CRP activates monocytes to a regulatory phenotype as part of the compensatory anti-inflammatory response in trauma. Incubation of isolated monocytes with trauma plasma, but not CRP-depleted trauma plasma, increased CD16 and HO-1. Differentiation of monocytes with M-CSF also induced CD16 and HO-1, and M-CSF-differentiated monocytes responded to CRP by secretion of IL-10 and IL-1RA. These results suggest a role for CRP in the generation and activation of regulatory monocytes during the acute phase response.
Transgenic expression of MDA5 enhances interferon responses, CD8 activation, and viral clearance (110.19)
Authors: Crampton Steve , Deane Jonathan , Otubusin Owokunile , Hasty Karen , Bolland Silvia 
Published: January 2023

Abstract
MDA5 is a pattern recognition receptor responsible for detecting double stranded viral RNA in the cytoplasm. Ligation of MDA5 leads to the induction of inflammatory cytokines and type one interferons (IFN-I) that help coordinate the adaptive and innate immune response against the virus. Here, we describe the phenotype of mice overexpressing MDA5 under its endogenous promoter. MDA5 transgenic (MDA5 Tg) mice have normal numbers of B and T cells, yet expanded numbers of CD11b+ cells expressing Ly6C. Splenocytes isolated from MDA5 Tg display an interferon gene expression signature. MDA5 Tg CD8+ T cells have increased surface expression of CD44, CD69, Ly6C and Ly6A/E, indicative of an activated phenotype. In vitro, CD8+ T cells from MDA5 tg hyperrespond to stimulation. Surprisingly, MDA5 tg mice fail to mount a CD8+ T cell-driven antigen-specific immune response against Vesicular stomatitis virus (VSV). MDA5 tg mice also lacked VSV neutralizing antibodies 16 days after infection with the virus. Although MDA5 tg mice failed to mount a full immune response against the virus, they were resistant to a lethal dose of VSV. This result suggests that increased viral resistance of MDA5 Tg is independent of antibody-neutralization and CD8+ T cell-mediated clearance, and might be due to high expression levels of interferon inducible inhibitors of virus replication. MDA5 tg mice could serve as a model for diseases involving type one interferon and CD8+ T cells, such as Type One Diabetes.
The role of autophagy and amphisome formation in virus recognition and virus-stimulated IFN-α production by human plasmacytoid dendritic cells (111.30)
Authors: Deng Jing , Dai Jihong , Fitzgerald-Bocarsly Patricia 
Published: January 2023

Abstract
pDC secrete large quantities of IFN-α in response to viral nucleic acid stimulation via TLR7/9. However, little is known about virus trafficking in pDC after entry. To investigate the subcellular trafficking of viral particles, we examined intracellular events after viral stimulation. The formation of double-membrane autophagosomes was observed by EM within 2hr stimulation of pDC with HSV, Flu, HIV-1 or CpGA. Class I/III PI3K inhibitors 3-methyladenine, wortmannin and LY294002 significantly suppressed virus-induced IFN-α production at an early stage. However, using Amnis imaging flow cytometry, we found that uptake of HSV-GFP was not inhibited, suggesting virus-induced autophagy rather than uptake was specifically blocked by PI3K inhibitors. To further investigate events related to virus-induced autophagy, we studied the colocalization of HSV-GFP particles, early endosomal marker CD71 and autophagagosomal marker LC3B. CD71 co-localized with LC3B after pDC stimulation with virus or CpGA, indicating amphisomes, which are autophagic vacuoles formed from fusion of autophagosomes and endosomes, were induced. With chloroquine, an inhibitor of endosomal fusion and acidification, there was accumulation of the internalized HSV-GFP particles and amphisomes. Collectively, our studies demonstrate that amphisomes are induced by virus or TLR9 ligand stimulation in pDC. Further studies will be conducted to characterize the role of these amphisomes in virus-induced IFN-α production.
Caspase-1-induced pyroptosis is an innate immune effector mechanism against intracellular bacteria (111.33)
Authors: Miao Edward , Leaf Irina , Treuting Piper , Mao Dat , Sarkar Anasuya , Wewers Mark , Aderem Alan 
Published: January 2023

Abstract
Macrophages mediate crucial innate immune responses via caspase-1-dependent processing and secretion of IL-1β and IL-18. While wild type Salmonella typhimurium infection is lethal to mice, a strain that persistently expresses flagellin was cleared by the cytosolic flagellin detection pathway via NLRC4 activation of caspase-1; however, this clearance was independent of IL-1β and IL-18. Instead, caspase-1 induced pyroptotic cell death released bacteria from macrophages, exposing them to uptake and killing by reactive oxygen species in neutrophils. Similarly, caspase-1 cleared Legionella and Burkholderia by cytokine independent mechanisms. Our results show, for the first time, that caspase-1 can clear intracellular bacteria in vivo independent of IL-1β and IL-18, and establish pyroptosis as an efficient mechanism of bacterial clearance by the innate immune system.
RGS2 is a negative regulator in STAT3-mediated Nox-1 expression (111.5)
Authors: Baek Suk Hwan , Lee Hyung-Kyoung , Kim Jin-Sik , Park Dae-Weon , Chin Byung-Rho , Bae Yoe-Sik 
Published: January 2023

Abstract
RGS2 protein is well known as a negative regulator of GPCR signaling. In our previous research, we understood that RGS2 mRNA expression is decreased by TLR4, and thereby foam cell formation is induced. The present study proved how the decreased RGS2 regulates specific gene, Nox-1, expression. While TLR2 stimulation increased Nox-1 expression and decreased RGS2 expression in wild type mice macrophages, such phenomena did not occur with TLR2 KO mice macrophages. Nox-1 expression increase by TLR2 was intensified by RGS2 siRNA, and was inhibited by RGS2 overexpression. TLR2 stimulation caused phosphorylation of tyrosine 705 and serine 727 residues, and STAT3 siRNA inhibited Nox-1 expression by TLR2. RGS2 directly associated with STAT3 acting as a negative regulator by inhibiting Nox-1 expression by TLR2. PKC-eta and PLD2 also participated in Nox-1 regulation by controlling RGS2 expression. Overall, these results suggest that TLR2 stimulation decreases RGS2 expression mediating PKC-eta and PLD2, and thereby will increase Nox-1 expression by releasing the transcription ability of STAT3 restrained by RGS2.
Toll-like receptor3 overexpression alters responses to toll-like receptor7 ligand (111.6)
Authors: Zheng Wenjie , Crampton Steve , Bolland Silvia 
Published: January 2023

Abstract
Macrophages are key components of the inflammatory response. PolyIC induces type I IFN and inflammatory responses through TLR3 or MDA5 pathway, depending on the activating conditions and cell types. In our hands, bone marrow-derived macrophages (BMM) in vitro preferentially engage TLR3 in response to stimulation with naked poly IC. In contrast, the same macrophages activate TLR3-independent pathways in response to Fugene-complexed poly IC.These results suggest that lipid-complexed poly IC is more likely to induce inflammatory cytoplasmic pathways. To further investigate the role of TLR3 in macrophage activation, we have generated TLR3-overexpressing transgenic mice (TLR3 Tg). TLR3 Tg mice contain 20-30 copies of a BAC clone which spans the entire mouse tlr3 gene. As a consequence, TLR3 mRNA expression is greatly enhanced in BMM and splenoctes of TLR3 Tg mice. These mice had normal life span and no obvious changes in immunological characteristics or spontaneous pathology. When we stimulated TLR3 Tg BMM with naked poly IC, we detected a small increase in their hyperresponsiveness to poly IC compared to WT cells. Interestingly, TLR3 Tg BMM greatly enhanced the synergistic effect of TLR7 agonist pretreatment with naked poly IC in the inflammatory cytokine secretion, but did not enhance the TLR9 agonist pretreatment. In the future, we will further investigate the mechanisms involved in these effects and explore in vivo consequences of TLR receptor cross-regulation.
Inhibition of Th17 cells and promotion of Tregs in Fcγ chain-deficient mice contributes to the attenuated atherosclerotic lesions (112.1)
Authors: Ng Hangpong , Burris Ramona , Nagarajan Shanmugam 
Published: January 2023

Abstract
The presence of anti-oxLDL IgG is well documented in clinical and animal studies. However, the role for FcγRs to the progression of atherosclerosis has not been studied in detail. In the present study, we investigated the role for activating FcγR in the progression of atherosclerosis using apoE-Fcγ-chain double knockout (DKO) mice. Relative to apoE KO mice, arterial lesion formation was significantly decreased in apoE-Fcγ-chain DKO mice. Bone marrow chimera studies showed reduced lesions in apoE KO mice receiving the bone marrow of apoE-Fcγ-chain DKO mice. Cytokine responses by activated CD4+ cells showed higher levels of both IL-10 and IFN-γ secretion. Moreover, compared to apoE KO mice, anti-oxLDL IgG1 (Th2) and IgG2a (Th1) were increased in apoE-Fcγ-chain DKO mice. Interestingly, the number of Th17 cells and the secretion of IL-17 by activated CD4+ cells were decreased in apoE-Fcγ-chain DKO mice. Notably, the number of T-regulatory cells, expression of mRNA, and secretion of TGF-β and IL-10 were increased in apoE-Fc-chain DKO mice. Furthermore, secretions of IL-6 and STAT-3 phosphorylation essential for Th17 cell genesis were reduced in apoE-Fcγ-chain DKO mice. Collectively, our data suggest that activating FcγR promotes atherosclerosis by inducing Th17 response in the hyperlipidemic apoE KO mouse model. Our findings indicate that blocking activating FcγR may be a possible therapeutic approach to inhibit T cell responses contributing to the progression of atherosclerosis.
CD8 effector cell immunological synapse and cell division in the virally infected CNS (112.20)
Authors: Kim Jiyun , Stewart-Hutchinson Phillip , Kang Silvia , McGavern Dorian , Dustin Michael 
Published: January 2023

Abstract
Two-photon intravital imaging through thinned skull was used to visualize the dynamics of LCMV specific P14 effector T cells on day 6 LCMV meningitis. Most P14 were motile without forming stable synapses with infected stroma before the mice succumb to immunopathology. To determine if a subset of effector T cells can form more stable interactions with professional antigen presenting cells (APCs), we visualized the dynamics of CFP+ P14 T cells in CD11c-YFP transgenic or CX3CR1GFP/+ mice. p14 formed synapses with CD11c+ dendritic cells, found mostly on the meninges and perivascular spaces in the layer 1 cortex, and with activated CX3CR1+ microglia in the brain parenchyma and perivascular areas. Long contacts were rare with ramified CX3CR1+ microglia and small CD11c+ cells. P14 motility increased upon infusion of anti-MHC class I and anti-LFA-1 blocking antibodies. Fluo-Red dye infusion revealed Ca2+ flux in both P14 and tissue cells. Unexpectedly, during extensive imaging, we noted division of P14. P14 underwent divisions after contacting CD11c+ and away from these cells during the imaging period. We conclude that while the dominant behavior of P14 effector T cells during LCMV meningitis is continuous motility, a subset arrests with professional APCs. These prolonged interactions were sometimes terminated by cell division. Other instances of cell division suggest that there is significant division of anti-viral effector cells at the site of infection.
The transcription factor Klf2 regulates B cell trafficking in a lineage-specific manner (112.21)
Authors: Sebzda Eric , Hoek Kristen , Cendron Delphine , Gordy Laura , Collins Patrick , Parekh Vrajesh , Aune Thomas , Joyce Sebastian , Thomas James 
Published: January 2023

Abstract
Manipulation of homing receptors on effector lymphocytes may provide therapeutic relief from autoimmune disorders; however, few treatments are available to effectively limit B lymphocyte trafficking and prevent effector cell access to peripheral tissues. Understanding the molecular networks that regulate lymphocyte trafficking will be required to develop stable, efficacious drugs that work in a lineage-specific manner. In this regard, our preliminary findings indicate a regulatory link between Kruppel-like factor 2 (Klf2) and B cell homing receptors. Conditional excision of Klf2 leads to lineage-specific defects in mature B cell compartmentalization including displacement of marginal zone (MZ) B cells in the spleen, aberrant trafficking of follicular (FO) B cells, and an absence of peritoneal B1 lymphocytes. These phenotypes are quickly replicated in vivo following tamoxifen-induced Klf2 excision, consistent with a post-developmental defect in mature B cell homeostasis. Moreover, we demonstrate that Klf2 differentially regulates homing receptor expression in a B cell lineage-specific manner, resulting in functionally altered migration patterns. These results highlight Klf2 as a non-redundant transcription factor involved in naïve B cell homing. In turn this study may provide impetus for the rational design of therapeutics that target the molecular mechanisms governing B cell migration.
The role of β2 integrins in host immune responses to bacterial infection (112.26)
Authors: Bose Tina , Lefrancois Leo 
Published: January 2023

Abstract
β2 (CD18) integrins (CD11a,b,c,d) are important adhesion molecules necessary for leukocyte migration, homing, and cell to cell interactions. CD18-deficiency in humans is characterized by recurrent bacterial infections and poor wound healing, due to defects in leukocyte activation and migration. However, the in vivo role of each of the individual β2 family members is poorly understood. CD11a, CD11b and CD11c are all upregulated on activated CD8 T cells. In order to study the potential role of β2 integrins in host defense, CD11a, CD11b and CD11c deficient and wildtype mice were infected with Listeria monocytogenes (LM). Our results indicate that the absence of CD11b and CD11c had no effect on antigen specific CD8 T cells in response to infection with LM. On the contrary, after infection, antigen specific CD8 T cells in CD11a -/- mice exhibited diminished proliferation and activation resulting in significantly reduced numbers at the peak of the response. Moreover, CD11a KO mice exhibited a reduced differentiation of short lived effectors subset (KLRG1 hi CD127 lo) although cytokine and granzyme B production was unaffected. Interestingly, upon secondary infection with LM, antigen specific memory CD8 T cells in CD11a KO responded robustly. Overall, these results suggest that CD11a plays a crucial role in regulation of primary CD8 T cell effector differentiation, but may be dispensable for secondary responses to Listeria infection.
Thrombin mediates kinesis in human resting CD8 T cells from healthy controls and HIV infected individuals through specific interaction with thrombin receptor (PAR-1). (112.7)
Authors: Hurley Amanda , Smith Mindy , Karpova Tatiana , Nikel Erin , Packard Beverly , Imamichi HIromi , MacNally James , Higgins Jeanette , Sneller Michael , Lane Clifford , Catalfamo Marta 
Published: January 2023

Abstract
Chronic infections, such as HIV, are complicated with coagulation abnormalities and increased risk of cardiovascular events, however the mechanisms that link infection/inflammation and coagulation are not clearly defined. Thrombin is the main protease of the coagulation cascade and also participates in the host inflammatory response to tissue insult through activation of protease activated receptor-1 (PAR-1). PAR-1 is expressed in cells of the innate immune system. In this study we characterized the expression of PAR-1 on T cells from healthy controls. PAR-1 was expressed on CD4 and CD8 T cells with effector memory phenotype (23%±1.9 and 48%±3.5 respectively). Compared to healthy controls, patients with HIV infection had 1.6 times increased (p=0.0001) expression of PAR-1 on CD8 but not CD4 T cells. We hypothesized that thrombin could be involved in the recruitment of T cells to the place of tissue insult. Therefore, we imaged the effect of thrombin in individual cells over time using time-lapse microscopy. Thrombin was able to induce rapid kinesis after the addition of thrombin to human resting CD8 T cells in a PAR-1 dependent fashion. In addition, CD8 T cells from HIV infected patients were able to migrate in response to thrombin suggesting that during HIV infection CD8 T cells could respond to burst of thrombin leading to increased inflammation. This study provides new evidence of cross-talk between infection/coagulation adding a new player: T cells.
T cell expression of NOX2 regulates T helper differentiation (113.10)
Authors: Shatynski Kristen , Chen Haiyan , Williams Mark 
Published: January 2023

Abstract
Absence of phagocyte NADPH oxidase (NOX2) activity causes chronic granulomatous disease (CGD), a primary immunodeficiency characterized by recurrent bacterial infections. In seeming contradiction, immune responses to agents such as H. pylori and influenza imply an altered or augmented adaptive immune response in CGD. We have shown NOX2 expression in T cells, and oxidase deficient T cells demonstrate Th1 skewed cytokine secretion upon in vitro stimulation. The goal of this study was to determine if these differences are T cell inherent, or if other cells promote T helper polarization. Activation of purified naive CD4+ T cells from NOX2-deficient mice led to augmented IFN-γ and diminished IL-4 production, consistent with a Th1 skewing of naïve T cells. This was supported by enhanced expression of the transcription factor T-bet and lower levels of GATA-3. Similar to NOX2-deficient cells, Th2-biased BALB/c T cells treated with the antioxidant N-acetyl cysteine produced increased IFN-γ and decreased IL-4 as compared to untreated controls. Inhibition of TCR-induced STAT5 phosphorylation was identified as a potential mechanism for skewed T helper differentiation. Stimulation of TCR-transgenic, OT-II T cells in vitro with NOX2-deficient splenic APC produced similar amounts of IL-4, IL-17, and IFN-γ as compared to those incubated with wild type APC, supporting the conclusion that NOX2 can regulate the adaptive immune response in a T cell inherent fashion.
Twist1 regulates Th1 cell development by interfering with its transcriptional network (113.11)
Authors: Pham Duy , Kaplan Mark 
Published: January 2023

Abstract
Although T-bet and STAT4 are required for Th1 programming, detailed transcriptional networks in Th1 cells have not been clearly defined. Twist1 is a basic helix-loop-helix (bHLH) transcription factor that is regulated by STAT4 in Th1 cells. We demonstrated that knocking down Twist1 in Th1 cells using Twist1-targeting small hairpin RNA vector increased IFN-γ and TNF-α and that overexpression of Twist1 in Th1 cells results in the decrease in IFN-γ and TNF-α production, which correlates with a reduction of STAT4 activation, and the expression of number of Th1 genes including T-bet, Runx3, and Hlx1. However, ectopic expression of Runx3 but not IL-12Rβ2 or T-bet can rescue the inhibitory effect of Twist1. Twist1 functions through several mechanisms including the direct binding of Twist1 to the E-box containing regions of the Ifng, Il12rb2, Tbx21 and Runx3 promoters in activated Th1 cells. Moreover, there is a physical interaction between Twist1 and Runx3 in both cell line and primary T cell culture. This study provides new insight on how Twist1 negatively regulates the Th1 transcriptional network, and how modulating Twist1 expression could be an approach for gene-based therapy in autoimmune diseases mediated by Th1 cells.