bluuebunny/crossref_metadata_2025_split huggingface dataset:
mTORC2 negatively regulates DC PD-L1 and IL-10 through SOCS3 and STAT3 (172.34)
Authors: Rosborough Brian , Matta Ben , Turnquist Heth , Thomson Angus 
Published: January 2023

Abstract
The mammalian Target of Rapamycin (mTOR) is a central regulator of dendritic cells (DC) that functions in two independent complexes, mTOR complex (mTORC) 1 and 2. Rapamycin (RAPA) is a selective inhibitor of mTORC1, whereas novel ATP-competitive mTORC1/2 inhibitors (Torin1) inhibit both complexes. The immunoregulatory function of RAPA-resistant mTORC2 in DC is unknown. C57BL/6, PD-L1-/-, IL-6-/-, IL-10-/-, IL-12/23p40-/-, Ebi3-/-, interferon regulatory factor (IRF)1-/-, or FoxO1/3/4-/- bone marrow cells were cultured in GM-CSF and IL-4 to generate CD11c+ DC. RAPA or Torin1 was added on d2, and LPS added on d7. DC phenotype and signaling was assessed (d8) by flow cytometry and immunoblot, respectively. While RAPA reduced PD-L1, mTORC1/2 inhibition selectively upregulated PD-L1 expression on DC in a STAT3-dependent manner in unstimulated and LPS-stimulated DC. Whereas RAPA reduced IL-10 secretion, mTORC1/2 inhibition augmented IL-10 secretion following LPS stimulation. The PD-L1 regulator, IRF1, and FoxO, a STAT3 regulator, were not required for PD-L1 upregulation. PD-L1 upregulation on mTORC1/2-inhibited DC did not require autocrine IL-6, IL-10, IL-12/23, or IL-27. SOCS3, a negative regulator of STAT3, was reduced dramatically in mTORC1/2-inhibited, but not RAPA-exposed DC. mTORC2 negatively regulates STAT3 in DC through SOCS3. Blockade of mTORC2 signaling enhances PD-L1 expression and IL-10 secretion. These novel findings establish mTORC2 as a key regulator of DC function.
S4D-SRCRB, a soluble member of the group B scavenger receptor cysteine-rich superfamily, binds to both endogenous and exogenous structures (172.8)
Authors: Miró Cristina , Roselló Sandra , Padilla Olga , Yelamos José , Vila Jordi , Lozano Francisco 
Published: January 2023

Abstract
Human S4D-SRCRB is a soluble member of the Scavenger Receptor Cysteine-Rich Superfamily (SRCR-SF), and is composed of four group B SRCR domains separated by Pro-, Ser- and Thr-rich polypeptides, which are o-glycosilated. Northern blot analysis indicated that S4D-SRCRB is expressed as two major mRNA species: one of 2,8 kb, with a restricted tissue expression pattern (mainly kidney, placenta and HepG2 cell line), and another of 1,5 kb, with a broader distribution. To gain insight on the function of this molecule, we produced and affinity purified a recombinant form of this protein fused to a HA tag (rhS4D-HA) using an episomal expression system (pCep/Pu) in HEK 293-EBNA cells. Similarly to other members of the SRCR-SF (S5D-SRCRB, CD6 and DMBT1), rhS4D-HA binds to the surface of bacteria (both gram-positive and -negative) and fungi (both saprophytic and pathogenic), and this is mediated through recognition of lipotheicoic acid (LTA) or lipopolysaccharide (LPS), and lineal glucans, respectively. Interestingly, rhS4D-HA increased the secretion of IL-8 of HepG2 when exposed to bacterial cell wall components. Moreover, rhS4D-HA also binds to some (fibronectin and laminin) but not all (collagen) extracellular matrix proteins. Taken together the data suggest that S4D-SRCRB behaves as a dual-specific receptor for non-self and self-structures, likely playing a role in the innate defence and homeostasis of epithelial surfaces.
Mechanisms underlying T cell guided innate immunity after inhalation of Staphylococcus enterotoxin A (SEA) (173.1)
Authors: Kumar Sanjeev , Menoret Antoine , Vella Anthony 
Published: January 2023

Abstract
Inhalation of Staphylococcus aureus enterotoxin A (SEA) in mice induces strong pulmonary inflammation marked by vascular permeability and alveolitis, thereby modeling aspects of lung injury in humans. After SEA inhalation CD8+Vβ3+ T cells robustly expand but the early events of this response in the lung are unclear. Our new data show that Vβ3+ T cells are activated quickly after SEA inhalation since CD69 and the high affinity IL-2 receptor α-chain (CD25) were upregulated by 3 hours. This coincided with several changes in the innate immune system including recruitment of neutrophils, monocytes and NK cells into the lung, and imprinting an activation phenotype on macrophages. Importantly, in the absence of T cells all of these effects were abrogated. As a way to understand how T cells condition innate immunity, we analyzed cytokine production and found an unexpected role for TCRαβ T cells promoting IL-17 secretion by TCRγδ T cells. We demonstrated that TCRγδ cells did not produce IL-17 when tested in TCRβ-/- mice. Currently, we are defining how TCRγδ cells are induced to synthesize IL-17 by the TCRαβ cells. In sum, TCRαβ T cells mediate pro-inflammatory responses, not only as a direct source of cytokines, but also by guiding the recruitment and activation of innate cells. Thus, our data show role reversal between the adaptive immune and innate immune system.
Pathway Signature PCR Array: a novel method for studying inflammatory responses (173.11)
Authors: Wu Zhong , Schlumpberger Martin , Raza Sameen , Yu Jiaye , DiCarlo John , Wang Yexun , Devgan Vikram 
Published: January 2023

Abstract
Signaling pathways play central roles in cellular physiology, and assessing the state of pathways helps clarify molecular mechanisms of disease and inflammatory responses. Gene expression signatures of pathway activation status enable reliable measurement of pathway activity. Our group developed Pathway Signature PCR Arrays to analyze gene expression and determine pathway activity within a single real-time PCR experiment. This study identified a set of genes that provide a gene signature for evaluating IL6/STAT3 pathway activity. A set of 88 signature genes were derived from microarray profiling on HepG2 and MCF10A cells treated with IL-6 protein and STAT3-specific siRNA. Genes were selected on the basis of statistically significant expression changes in response to IL-6 and reversion of altered expression upon treatment with STAT3 siRNA. These 88 IL-6/STAT3 response genes were further verified by real-time PCR, using a panel of 14 different cell lines stimulated with IL-6 or inhibited with STAT3 siRNA. Using a mathematical classifier method, a subset of genes was identified as a gene expression signature for assessing IL-6/STAT3 pathway activity. These signature genes, along with IL-6 pathway-related genes identified by literature mining, constitute the IL-6 Pathway Signature PCR Array. This array provides a useful tool for studying regulation of pathway activity via gene expression changes during inflammatory responses. Array is for molecular biology applications only.
CD34 family proteins are key regulators of inflammatory cell migration and vascular integrity (173.17)
Authors: McNagny Kelly , Debruin Erin , Gold Matthew , Bennett Jami , Blanchet Marie-Renee , Maltby Steve , Hughes Michael 
Published: January 2023

Abstract
CD34 is a widely used marker of hematopoietic progenitors and vascular endothelia yet its function remains obscure. CD34 is the founding member of a family of three proteins (CD34, Podocalyxin and Endoglycan) with overlapping expression patterns and we have used gene targeting to investigate the function and redundancy of these proteins. We find that CD34 is uniquely expressed by mast cells, eosinophils and dendritic cells and that mice lacking CD34 are remarkably resistant to a variety of mucosal inflammatory diseases including allergic asthma, hypersensitivity pneumonitis and colitis. Resistance reflects defective inflammatory cell trafficking due to enhanced adhesion and impaired chemotaxis and these defects can be reversed through ectopic expression of a human CD34 transgene. Podocalyxin, in contrast, is uniquely expressed by a variety of embryonic tissues and inactivation of the Podxl gene leads to perinatal lethality due to a failure of tissues to undergo appropriate morphogenesis. To investigate the function of Podocalyxin (and CD34) on adult vasculature we have generated mice that selectively inactivate the encoding gene in endothelia. These exhibit excessive vascular leakage and develop an emphysema-like phenotype due to poor vessel patency in the lung. In summary, CD34 type proteins play critical roles in a variety of normal developmental processes and immunity. Mechanistically, they act as key regulators of cell adhesion, cell chemotaxis and morphogenesis.
Targeted PGE2 inhibition enhances anti-viral immunity through induction of Type I interferon in macrophages (P6180)
Authors: Divangahi Maziar , Coulombe François , Jaworska Joanna , Verway Mark , Tzalepis Fanny , Massoud Amir , Gillard Joshua , Wong Gary , Kobinger Gary , Joubert Philippe , Fritz Jörg , Xing Zhou , Powell William 
Published: January 2023

Abstract
The 1918 Spanish Influenza virus pandemic had a major impact on the rise in popularity of non-steroidal anti-inflammatory drugs (NSAIDs). Although these drugs are used extensively for the management of Influenza symptoms, their role in anti-viral immunity is poorly understood. NSAIDs act mainly by inhibiting cyclooxygenase enzymes involved in the synthesis of all prostaglandins, including PGE2. We now show that during Influenza infection, upregulated PGE2 inhibited macrophage type I interferon production and apoptosis via EP2 and EP4 receptors, thereby increasing virus replication. This inhibitory role of PGE2 was not limited to innate immunity since both antigen presentation and T cell mediated immunity were also suppressed. Inhibition of microsomal prostaglandin E-synthase 1 (mPGES-1), and thereby PGE2, in human Mϕ was protective against Flu infection. These data show that the mPGES-1/PGE2 pathway is specifically targeted by Influenza virus to evade host anti-viral immunity and propose that specific inhibition of PGE2 may serve as a new treatment for viral infection.
The length of the peptide in the MHC groove compartmentalizes the CD8+ T cell repertoire (P1186)
Authors: Miles John , van den Berg Hugo , Ekeruche-Makinde Julia , Skowera Ania , Cole David , Dolton Garry , Schauenburg Andrea , Tan Mai Ping , Pentier Johanne , Peakman Mark , Price David , Burrows Scott , Sewell Andrew , Wooldridge Linda 
Published: January 2023

Abstract
TCRs expressed at the CD8+ T-cell surface interact with short peptide fragments bound to MHC class I molecules (pMHCI). The TCR/pMHCI interaction is pivotal in all aspects of CD8+ T-cell immunity. However, the rules that govern the outcome of TCR/pMHCI engagement are not entirely understood, and this is a major barrier to understanding the requirements for both effective immunity and vaccination. Using large combinatorial peptide arrays, each containing between twenty billion to forty quadtrillion distinct peptides, we have discovered an unexpected feature of the TCR/pMHCI interaction by showing that all TCRs exhibit an explicit preference for an MHCI-bound peptide of defined length. Agonists of nonpreferred length were extremely rare, suboptimal, and often entirely distinct in sequence. Structural analysis indicated that alterations in peptide length have a major impact on antigenic complexity, to which individual TCRs are unable to adapt. This surprising finding, that the CD8+ T cell repertoire is rigorously "compartmentalized" by peptide length, has important consequences for cellular immune system research and vaccine development.
ROR1 directed delivery of FTY720 derivative OSU-2S mediates potent cytotoxic activity against chronic lymphocytic leukemia but not normal B cells in-vitro and in ROR1 expressing transgenic CLL mouse model in-vivo (P3275)
Authors: Muthusamy Natarajan , Mao Yicheng , Frissora Frank , Mani Rajeswaran , Wang Jiang , Wu Yun , Yu Bo , Baskar Sivasubramanian , Rader Christoph , Phelphs Mitch , Chen Ching-Shih , Lee Robert , Lee L. , Byrd John 
Published: January 2023

Abstract
ROR1, a member of the receptor tyrosine kinase-like orphan receptor family is expressed in chronic lymphocytic leukemia (CLL) but not normal B cells. We describe here a novel anti-human ROR1 antibody (2A2-IgG) based immunoliposomal delivery system for CLL therapy with improved specificity in vitro and in vivo. 2A2-IgG immobilized onto the surface of the liposomes (2A2-IgG-ILP), was evaluated for targeted delivery of a recently described novel phosphatase activator, OSU-2S. 2A2-IgG-ILP-OSU-2S demonstrated improved cytotoxicity in ROR1 positive CLL cells but not in normal B cells in-vitro. To validate the in-vivo efficacy of the ROR1 directed 2A2-IgG-ILP-OSU-2S, we developed transgenic mice expressing hROR1 on B cells (hROR1 Tg). Crossing the hROR1 Tg mice to the previously described Eμ-Tcl1 CLL mice resulted in double Tg mice with progressive CLL like disease characterized by accumulation of CD5+CD19+ROR1+ B cells. 2A2-IgG-ILP-OSU-2S exhibited selective binding and delivery of OSU-2S to ROR1+ but not ROR1- B cells in-vitro and elimination of circulating ROR1+ B cells in-vivo. Additionally, preliminary pharmacokinetic analysis of the targeted and non-targeted ILP-OSU-2S in the ROR1 transgenic mouse revealed altered pharmacokinetic profiles with 2A2-IgG targeted formulation. These studies validate the ROR1 targeted immunoliposomal formulations and the novel transgenic mouse models for evaluation of therapeutic agents directed to ROR1 positive B cell malignancies.
IL-1 and IFN-gamma differentially program human dendritic cells exposed to beta-glucan via IkappaB-zeta regulation (P6279)
Authors: Cardone Marco , Dzutsev Amiran , Li Hongchuan , Gerosa Franca , Winkler-Pickett Robin , Provezza Lisa , Riboldi Elena , Orr Selinda , Steinhagen Folkert , Wewers Mark , McVicar Daniel , Anderson Stephen , Goldszmid Romina , Lyakh Lyudmila , Trinchieri Giorgio 
Published: January 2023

Abstract
The molecular networks controlling the DC programming in response to C-type lectin receptor agonists including beta-glucan, a fungal and bacterial component and ligand for the Dectin-1 receptor, are still poorly characterized. We studied these immunological aspects by comparing the response of human mono-DCs to the Dectin-1 ligand, beta-glucan, and to the TLR4 ligand, LPS using a gene expression/perturbation approach. Gene expression analysis of mono-DCs stimulated by beta-glucan identified IL-1, TNF, and type I IFN encoded by “early/immediate” type genes as predicted regulators of the beta-glucan-induced “late” transcriptional response. A perturbation analysis revealed that endogenous IL-1, via the nuclear factor IkappaB-zeta, selectively promoted the expression of beta-glucan-induced “late” genes such as those encoding IL-6 and IL-23, thus programming DCs to initiate Th17 responses. TNF and type I IFN were found to modulate the response to both beta-glucan and LPS. IFN-gamma counteracted the activity of IL-1 and reprogrammed human mono-DCs to promote Th1-like cells rather than Th17 in response to beta-glucan. Our data provide evidence that IL-1 and IFN-gamma promote distinct adaptive immune responses against the fungal and bacterial component beta-glucan by differentially modulating the DC programming. They also suggest potential strategies to modulate the immunity against beta-glucan containing pathogens.
Cytotoxic effector CD4 T cells express NKG2C/E and are defined by a unique program dependent on Blimp-1. (IRC8P.482)
Authors: Vong Allen , Marshall Nikki , Kuang Yi , Berg Leslie , Swain Susan 
Published: January 2023

Abstract
CD4 T cells with cytotoxic activity can be generated during many immune responses to acute viral infections and they can contribute to CD4-mediated viral control. During the peak of the CD4 T cell response to an acute infection of influenza A/PR8 [IAV], cytotoxic CD4 T cells [ThCTL] are found in the lung. The cytotoxicity they mediate is MHC II-restricted and perforin-dependent. We find the lung ThCTL express NKG2C and E that are members of the killer cell lectin-like receptor family [KLR]. Although the ThCTL cytotoxicity is strictly antigen dependent, we show that NKG2C/E can costimulate the cytotoxic activity of ThCTL in vivo, since target cells lacking the ability to present peptide on Qa-1b, which binds to NKG2C/E, are significantly protected from lysis by ThCTL during IAV infection. The ThCTL express a unique pattern of molecules including high levels of granzyme B, PSGL-1, PD-1 and CD27 as well as transcription factors Eomes and Blimp-1. We find that Blimp-1 is critical for the generation and differentiation of this subset, since a CD4 T cell intrinsic defect in Blimp-1 leads to a broad reduction of most of the ThCTL program including cytotoxic activity and migration to the lungs as well as expression of NKG2C/E, granzyme B, PSGL-1 and CD27. These features help to define a distinct ThCTL cell subset generated under distinct conditions during an acute viral infection that has novel target specificity and thus likely occupies a unique functional niche.
Streptococcus pneumoniae specific Th17 memory immunity provides cross protection against invasive pneumococcal diseases in mice. (IRC9P.701)
Authors: Wang Yan , Tian Ying , Jiang Bin , Weiser Jeffery , Shen Hao 
Published: January 2023

Abstract
Pneumonia caused by Streptococcus pneumoniae (Sp) remains a leading cause of serious illness and numerous deaths in children and elderly worldwide. Current pneumococcal vaccine is effective in preventing colonization by inducing serotype-specific antibodies. However, there is an increasing prevalence of infection by serotype strains not included in the vaccine; this highlights the need for a universal vaccine that protects against all serotypes. In current study, we found that mice intranasal immunized with Sp were protected against challenge with a different serotype Sp strain and cleared bacteria from the lung by day 2 post-challenge, and exhibited less apoptosis and tissue damage in lung. Sp infection in lung resulted in a tremendous CD4+T cell expansion and activation that consisted of mostly IL-17 producing Th17 cells but also IFNγ producing Th1 cells. Adoptive transfer of T cells from Sp immunized mice provided cross protection with a dominant Th17 recall response in the lung from donor CD4+ memory T cells. Furthermore, adoptive transfer of Sp specific CD4+ memory T cells provided cross protection against pneumonia and bacteremia, which can be blocked by IL-17 neutralizing antibody and transfer Sp memory CD4+ T cells from IL-17 knockout mice. Our results suggest that Sp memory Th17 cells played a key role in providing broad protective immunity against invasive Sp infection in a serotype independent manner.
POU2AF1 arranges systemic distribution of follicular helper T cells (IRC10P.466)
Authors: Yamashita Keiji , Mitsuhashi Yukari , Nagaya Tomonori , Jistukawa Sumito , Kamekura Ryuta , Seki Nobuhiko , Kumagai Ayako , Kawata Koji , Himi Tetsuo , Ichimiya Singo 
Published: January 2023

Abstract
Follicular helper T (Tfh) cells are a distinctive subset of CD4+ T helper cells, which play a critical role to establish specific humoral immunity by helping the germinal center formation of B cells. Currently, it is known that Tfh cells are maintained by BCL-6, MAF, and BATF; however, a mechanism managing Tfh cells is not fully clarified yet. To further dissect a regulator of Tfh cells, we employed transcriptome analysis of human tonsillar Tfh cells and thymic naïve T cells as a control. Results demonstrated that, in addition to BCL-6, MAF, and BATF, Tfh cells highly expressed POU2AF1 (BOB.1 or OBF1) at transcript and protein levels. POU2AF1 is a transcriptional coactivator associated with either OCT1 or OCT2 through recognition of POU domains and assists B cells to produce specific antibodies. Analysis using Pou2af1-deficient mice showed that the expression levels of Pou2af1 did not seem to correlate the levels of Cxcr5,Icos,Tigit and Bcl-6 in Tfh cells. Mix cell culture experiments further revealed no functional differences in between Pou2af1-negative Tfh cells and control Tfh cells. But it was of our interest that, after stimulation with foreign antigens, the frequencies of Tfh cells in immune tissues and peripheral blood of Pou2af1-deficient mice were altered in comparison to those of wild-type mice. Taken together, these suggest that POU2AF1/Pou2af1 might have a role to regulate spreading manner of Tfh cells over the body from local sites of immune response.
Myeloid dendritic cells maintain Th17 lymphocyte responses during the obstructive phase of experimental biliary atresia (IRC10P.471)
Authors: Lages Celine , Simmons Julia , Maddox Avery , DiPasquale Betsy , Shanmukhappa Shiva Kumar , Chougnet Claire , Miethke Alexander 
Published: January 2023

Abstract
Biliary atresia (BA) is a fibro-inflammatory obstruction of the extrahepatic biliary tree in newborns. It leads to rapid biliary fibrosis and is the most common indication for pediatric liver transplantation. In a model of rhesus rotavirus (RRV)-induced murine BA, plasmacytoid dendritic cells (DC) initiate NK-cell mediated hepatobiliary injury at 3 days post infection (dpi). We hypothesize that myeloid (m)DC coordinate T-cell driven injury during the later obstructive phase of BA. Following RRV injection into CD11c-DTR GFP pups on day 1 of life, frequency of hepatic DC increased 6fold at 12dpi compared with non-infected controls. Expansion of mDCs was accompanied by accumulation of activated CD4 cells in the liver. Plasma IL17A/F levels were increased at 8 and 12dpi (p&lt;0.01) and hepatic IL17A mRNA increased 9fold between 8 and 12dpi. In vitro coculture of DC from 12dpi pups with naïve CD4 cells induced IL17A/6 production. DC-depletion following RRV infection with diphtheria toxin reduced the number of hepatic CD4 cells by 3fold which was associated with 2fold decrease in hepatic IL17A mRNA expression and plasma Th17-associated cytokine levels (IL-17F/6) at 12dpi. Importantly, diminished Th17 responses correlated with reduced plasma total bilirubin levels (6.1 vs 11.3mg/dL in control; p&lt;0.01), a biomarker for biliary obstruction. We conclude that mDC maintain hepatic Th17 responses and regulate cholestatic liver injury during the obstructive phase of experimental BA.
Gene targeting of RhoA reveals its essential role in thymocyte development and metabolism (LYM7P.718)
Authors: Guo Fukun , Zhang Shuangmin , Konstantinidis Diamantis , Yang Jun-Qi , Kalfa Theodosia , Zheng Yi 
Published: January 2023

Abstract
RhoA GTPase is an intracellular signal transducer capable of regulating a wide range of cell functions including cytoskeleton dynamics, proliferation, and survival. In T lymphocytes, studies by using dominant negative mutant or C3 transferase expressing transgenic mice suggest that RhoA is involved in TCR signaling and related T cell functions such as polarization, migration, survival, and proliferation. To date, the physiological role of RhoA in T cell development remains unclear. In this study, we have achieved T cell-specific deletion of RhoA and found that RhoA gene disruption caused (1) a drastic decrease in the numbers of all thymocyte subsets including CD4-CD8- double negative (DN), CD4+CD8+ double positive (DP), CD4+CD8- single positive (SP), and CD4-CD8+ SP T cells, (2) an impaired β-selection at DN stage and positive selection at DP stage associated with altered pre-TCR and TCR signaling transduction, respectively, (3) a diminished thymocyte proliferation which could be due to impaired cytokinesis, (4) an increased apoptosis, and (5) an enhanced mitochondrial function as manifested by increased oxidative phosphorylation, glycolysis, mitochondrial membrane potential, mitochondrial numbers, mitochondrial DNA and ROS production. Thus, RhoA critically regulates T cell development by impacting on β-selection, positive selection, cell proliferation and survival through mediating pre-TCR signaling, TCR signaling, cytokinesis, and metabolism.