bluuebunny/crossref_metadata_2025_split huggingface dataset:
Retargeting IgM Antibodies with Different Specificities to a Common Cell Type Reveals Shared Mechanisms Regulating Cellular Activation (91.12)
Authors: Keenan Thomas D , Radhakrishnan Suresh , Bell Michael , Barry Michael , Pease Larry R 
Published: January 2023

Abstract
IgM antibodies, as multivalent ligands, have the capacity to activate cells differentially by cross-linking targeted cell surface signaling molecules. For example, hIgM12 (B7DC Xab) activates dendritic cells (DCs), leading to potent immunomodulation, while hIgM22 binds glial cells and causes remyelination in mouse models of multiple sclerosis. Both antibodies activate common signaling intermediates in their respective cellular targets suggesting a common mechanism of action. Elucidation of these shared mechanisms became possible by targeting both antibodies to a common cell type. Our previous studies demonstrate that B7-DC expression is required to activate downstream signaling in DCs by hIgM12. We utilized a phage display system to identify 7mer peptides bound by each IgM and expressed each peptide attached to a surrogate cell surface carrier on retrovirally tranduced bone marrow-derived DCs from B7-DC -/- mice. Transduced cells were treated with hIgM12, hIgM22 or anti-carrier IgM, and subsequent activity was compared against B7-DC +/+ DCs or B7-DC-reconstituted B7-DC -/- DCs. DC activation by each IgM was specific to expression of their conjugate peptide construct. Activation by hIgM22 was indistinguishable from activation by hIgM12, while anti-carrier IgM caused no activation. This indicates that while their target specificity differs, once bound these IgMs activate cells via common mechanisms.
Modification of DC function via blockade of CD40:TRAF interactions (91.13)
Authors: Qu Xinyan , Stout Robert D , Suttles Jill 
Published: January 2023

Abstract
Ligation of CD40 on dendritic cells (DC) results in rapid protein kinase and transcription factor activation leading to enhanced expression of genes encoding a variety of cytokines and costimulatory molecules. CD40 does not contain cytoplasmic sequences with catalytic activity and transmits signals relying on the use of TNFR-associated factors (TRAFs), a family of adapter proteins that are involved in signaling by the TNF receptor family and the toll/interleukin-1 receptor (TIR) family members. Cell-permeable TRAF6 and TRAF2/3/5 blocking peptides (TRAFBPs) corresponding to the TRAF6 or TRAF2/3/5 binding region of CD40 were employed to better define the roles of TRAF2 and TRAF6 in CD40 signaling in DC. Both TRAF6BP and TRAF2BP inhibited CD40-stimulated DC production of TNF and IL-6, but did not impact CCL2 and CXCL8 production. ERK1/2 phosphorylation was inhibited by both TRAF6BP and TRAF2BPs, whereas JNK, p65 and AKT phosphorylation were abrogated by TRAF6, but not by TRAF2BP. Both TRAF6BP and TRAF2BP have no effect on p38 MAP kinase activation. During antigen presentation to T cells, pre-incubation of DC with TRAF6BP and TRAF2BP effectively inhibited Th1 and Th17 differentiation, but did not impact activation of IL-10-producing T cells. Thus, targeting TRAF:CD40 interactions in DC may provide a means to selectively modulate DC function.
Expression of a2V-ATPase following addition of ATP leads to cell death (91.18)
Authors: Levine Rita , Kwong Christina , Beaman Kenneth 
Published: January 2023

Abstract
Surface expression of a2V-ATPase (a2V) is congruent with cell destruction and regulates the immune response. Leukocytes isolated from whole blood display a2V on their surface and are highly sensitive to cell destruction following addition of ATP. Exogenous ATP enhances and alters surface expression of a2V. This augments the cell destruction of certain white cell populations.
Like the monocytic cell line studied previously, the epithelial cell lines, TOV112D and JEG3, treated with ATP significantly increased surface expression of a2V. Leukocyte populations incubated with ATP altered surface expression of a2V. Monocytes were strongly a2V positive and sensitive to ATP. The addition of ATP for 1hr resulted in total destruction of the monocyte population. Lymphocytes treated with ATP for 2hrs increased surface expression of a2V dramatically. In vitro addition of ATP led to an increase in granulocytic cell death. These data demonstrate that a2V is an important marker of cell destruction in vivo and in vitro.
Estrogen promotes pro-inflammatory response by regulating multiple transcription factors (91.7)
Authors: Dai Rujuan , Phillips Rebecca A , Karpuzoglu Ebru , Khan Deena , Ahmed S. Ansar 
Published: January 2023

Abstract
In vivo 17-β estradiol (estrogen) treatment has been shown to promote inflammatory responses in different types of immune cells including splenocytes. However, the molecular mechanism of estrogen mediated pro-inflammatory responses is still not clear, which impede the therapeutic intervention of estrogen related immune disorders. In the present study, we investigated the effects of estrogen on the signaling pathways, which involved in the regulation of inflammatory molecules, especially inducible nitric oxide synthase (iNOS). We found that although activation of IFNγ-STAT-1 signaling is essential for Con-A stimulated iNOS expression in mouse splenocytes, it is not sufficient. The induction of iNOS in splenocytes required synergistic activation of STAT-1 and NF-κB. Nevertheless, the induction of Monocyte chemotactic protein-1 (MCP-1) in splenocytes from estrogen-treated mice is not dependent on STAT-1 and NF-κB. We then demonstrated that estrogen increases the expression and DNA binding activity of c/EBPβ, which is also important for inflammatory regulation. Further, our data revealed that serine protease play important role in the regulation of IFNγ-STAT-1 and NF-κB signaling in splenocytes from in vivo estrogen treated mice. Together, our data indicate that estrogen promotes pro-inflammatory responses by modulating the activity of multiple transcription factors including STAT-1, NF-κB, and c/EBPβ.
Induction of IL-6 in mixed glial cultures – potential role of CGRP in CD40-dependent neuropathic pain (91.8)
Authors: Malon Jennifer , Cao Ling 
Published: January 2023

Abstract
We have previously demonstrated a critical role of spinal cord microglial CD40 in the development of spinal nerve L5 transection (L5Tx)-induced neuropathic pain. Cross-talk between pro-inflammatory mediators and pro-nociceptive factors is believed to play an essential role in the development of neuropathic pain. Here, we investigated the effects of the pro-nociceptive factors calcitonin gene-related peptide (CGRP) and CX3CL1 with/without CD40 on the production of several pro-inflammatory mediators by glial cells in vitro. Anti-CD40 induced low levels of nitric oxide, a known pro-nociceptive factor, while CX3CL1 enhanced CXCL10 production by mixed glia. Interestingly, while anti-CD40 and CX3CL1 had little effect on IL-6 production, CGRP induced a significant secretion of IL-6 by glia that is unaffected by CD40 co-stimulation. In vivo data indicated that spinal cord IL-6 was increased by day 1 and spinal cord microglial CD40 expression was increased in an IL-6 dependent fashion by day 3 post-L5Tx. Together, these data lead us to hypothesize that CGRP released from primary sensory nerve terminals following L5Tx induces spinal cord glial IL-6 production, which in turn enhances microglial CD40 expression, thus facilitating the development of neuropathic pain. Future studies with microglial-enriched and depleted populations will be used to further delineate this pathway.
Support of this study: 1K01DA023503-01A1 (Cao), NIDA.
CD34 mediates dendritic cell trafficking in hypersensitivity pneumonitis (92.1)
Authors: Bennett Jami L , Blanchet Marie Renee , Girard Melissa , Cormier Yvon , McNagny Kelly M 
Published: January 2023

Abstract
Hypersensitivity pneumonitis (HP) is an inflammatory lung disease characterized by a chronic T cell airway infiltration. Dendritic cells (DC) have been reported to play a crucial role in trafficking of antigen to the lymph nodes and antigen presentation in this disease. However, little is know about what triggers and mediates DC trafficking in HP. CD34 is a sialomucin best known for it's expression on hematopoietic progenitors. This protein, which was recently reported as a facilitator of cell trafficking, is also expressed on a few subsets of mature cells such as DCs. We therefore explored whether CD34 expression in DCs was involved in DC trafficking to and from the lung in a mouse model of HP, using wild type and Cd34-/- mice. Lung inflammation and DC recruitment was evaluated in both strains. Results show that lack of CD34 expression leads to a lower lung inflammation (as characterized by lower total cell counts in the broncho-alveolar lavage). DC recruitment was reduced in the alveoli of Cd34-/- mice, but was increased in the lung tissue. Using bone marrow chimeric mice, this effect was attributed to the Cd34-/- environment (including potentially radio-resistant DCs). Lastly, intraveinous injection of wild type DCs reconstituted disease in Cd34-/- mice. We conclude that CD34 could mediate DC trafficking in HP and possibly other inflammatory lung diseases.
Supported by the Canadian Institutes of Health Research (CIHR), AllerGen Network Centre of Excellence (KMM), Multiple Sclerosis Society of Canada (JLB), Michael Smith Foundation for Health Research (KMM, MRB), and FRSQ (MRB)
Curcumin stimulates cyclooxygenase-2 (COX-2), prostaglandin I2 synthase (PGI2S) and PGE2 synthase (PGE2S) gene expression in human coronary artery endothelial cells (93.19)
Authors: Tan Xiaoyu , Poulose Ellen M. , Stechschulte Daniel J. , Dileepan Kottarappat N. 
Published: January 2023

Abstract
Curcumin is reported to decrease atherosclerosis progression in Apo-E/LDL receptor knockout mice. PGI2 and PGE2 are generated from arachidonic acid (AA) by the COX-PGI2S/PGE2S pathway and are known to have important roles in cardiovascular function. In this study, we evaluated the in vitro effect of curcumin on the expression and function of COX-2, PGI2S and PGE2S in human coronary artery endothelial cells (HCAEC). HCAEC monolayers were incubated with curcumin (1.25-10 μM), and the mRNA and protein expressions were monitored by qRT-PCR and Western blots, respectively. PGI2 and PGE2 levels in the culture supernatants were quantified by EIA. Incubation of HCAEC with curcumin led to concentration-dependent increases in COX-2 mRNA and protein expression without change in COX-1 expression. Curcumin also induced 2-fold increases in PGI2S and PGE2S gene expression. Despite the increase in COX-2, PGI2S and PGE2S gene expression, curcumin failed to increase PGI2 or PGE2 production. Furthermore, curcumin inhibited LPS-induced PGE2 and PGI2 production by HCAEC and exogenous AA could not reverse the inhibitory effect. These results show that curcumin acts both as an inducer of COX-2, PGI2S and PGE2S gene expression and as an inhibitor of stimulated PGE2 and PGI2 synthesis. The ability of curcumin to regulate prostanoid biosynthesis in HCAEC indicates its therapeutic potential for cardiovascular disease. (Supported by NIH R01-HL070101 and the Carey Arthritis Fund).
IL17 stimulates granulopoiesis via IL6 induction (93.5)
Authors: Sun Honghong , Worthen Scott 
Published: January 2023

Abstract
IL-6 is an important inflammatory cytokine required for Th17 cell differentiation. IL-17, a proinflammatory cytokine involved in many inflammatory diseases, is produced by Th17 cells, under the control of transforming growth factor-b and IL-6. IL17 may mediate neutrophil recruitment and granulopoiesis through G-CSF-dependent and -independent mechanisms. Using a novel in vitro culture and differentiation system and an in vivo lung inflammation/injury model, we demonstrate that addition of G-CSF and IL17 promotes functional neutrophil generation. The numbers of neutrophils released upon IL-17 stimulation is comparable to or more than that of G-CSF stimulation. In confirmation of the result published by Kolls et al, IL17 induces modest G-CSF production. However, IL-17 also induces a dramatic increase in IL6 expression that is not observed in G-CSF-treated cultures. Furthermore, both G-CSF and IL17-induced granulopoiesis is attenuated in IL6-deficient mice. Our data also show an increased population of lin-/c-kit+/Sca1- cells in IL17 and IL6-treated cultures. The Stat3 and pAKT are activated upon IL17 stimulation and contribute to activation of IL6 transcription. Taken together, IL-17 produced by T cells stimulates granulopoiesis through the release IL-6 and G-CSF and is critical for the initiation of innate immunity.
Delineation of hypoxia versus inflammatory contributions to tissue injury by attenuation of G-alpha-i signaling (93.7)
Authors: Moratz Chantal  M , Egan R , Zacharia A , Simovic M , Gist I , Dalle Lucca J , Tsokos G C 
Published: January 2023

Abstract
The complement dependent pathology resulting from mesenteric ischemia/reperfusion insult instigates tissue injury in mesenteric and systemic organs. In this study the contribution of G-protein receptor signaling through G[alpha]i to stages of tissue injury is delineated. Global G[alpha]i inhibition, by PTX, attenuated mesenteric injury and production of lipid modulators such as leukotriene B4. Systemic inflammation was inhibited as assessed by lung tissue injury scores and monitoring cellular infiltrates. To delineate the role of G[alpha]i signaling in mesenteric versus systemic inflammatory responses, an inhibitor of pulmonary inflammatory chemotactic G[alpha]i signaling, anti-CXCR2 was used. As expected PTX and anti-CXCR2, abrogated pulmonary inflammation. In contrast, anti-CXCR2 was not effective in mitigating mesenteric injury. The results from global versus specific G[alpha]i inhibition in local and systemic injury indicate multiple mechanisms by which G[alpha]i signaling facilitate tissue injury. The G[alpha]i linked signaling effects of complement activation and chemokine induced migratory control are distinct and can be mitigated independently. Delineating which signals govern particular components of tissue injury will be useful in developing therapeutics which can control specific deleterious aspects of systemic inflammation after trauma while maintaining the protective aspects of complement activation.
A novel analysis of single particle trajectories quantifies the dynamic interactions between LFA-1 and the actin cytoskeleton (94.13)
Authors: Das Raibatak , Cairo Christopher W , Coombs Daniel 
Published: January 2023

Abstract
Dynamics of cell-surface proteins are strongly influenced by their localized interactions with cytoskeletal attachments or intracellular signaling molecules. We developed a technique to analyze single particle trajectories to quantify these interactions. We modeled trajectories of a cell membrane-associated protein interacting with a homogeneously distributed binding partner, and applied this model to analyze the motion of the integrin receptor LFA-1 on the surface of T cells. Our analysis reveals that the diffusion of LFA-1 is two-state, with distinct diffusion coefficients in its free and cytoskeleton-bound forms. We also quantify the rates of transition between the two states, and we show that cellular activation leads to large changes in the LFA-1-cytoskeleton interactions.
Macrophage migration inhibitory factor and the regulation of adhesion molecule expression on the blood brain barrier during experimental autoimmune encephalomyelitis (94.15)
Authors: Cox Gina Mavrikis , Kithcart Aaron P , Williams Jessica , Smith Kristen , Shawler Todd , Whitacre Caroline C 
Published: January 2023

Abstract
Multiple sclerosis (MS) is a chronic inflammatory and demyelinating disease of the central nervous system (CNS). Cerebrospinal fluid (CSF) isolated from patients during relapse was found to contain greater levels of macrophage migration inhibitory factor (MIF) relative to samples collected during remission, thus suggesting a role for MIF in the pathogenesis of MS. We have demonstrated that upon induction of experimental autoimmune encephalomyelitis (EAE), mice deficient in MIF exhibit reduced clinical signs as well as fewer inflammatory infiltrates in the brain and spinal cord. MIF deficient mice, however, do not differ from wild type controls in the function of autoreactive lymphocytes in the periphery. These findings suggest that MIF is necessary for progression of EAE in the CNS, particularly at the level of the blood brain barrier (BBB). Mice treated with anti-MIF antibody show impaired migration of lymphocytes into the CNS and down-regulation of adhesion molecules on the BBB. Thus, we hypothesize that MIF is acting at the level of the BBB, regulating adhesion molecule expression and therefore access to the CNS. We are analyzing the expression of adhesion molecules, such as VCAM-1 and ICAM-1 in the brain and spinal cord of MIF deficient and small molecule inhibitor-treated mice. (Supported by NIH grant AI 064320 and National MS Society grant RG3272)
Effect of the sphingolipid synthesis inhibitor, myriocin, in a murine pulmonary inflammation model (94.19)
Authors: Edling Andrea E , Mahan Amy , Leonard John , Williams John , Kaplan Johanne , Perricone Michael 
Published: January 2023

Abstract
Effect of the sphingolipid synthesis inhibitor, myriocin, in a murine pulmonary inflammation model Sphingolipids are signaling molecules involved in many aspects of cell function. Therapeutic modulation of the sphingolipid pathway is a potential approach to manage inflammation. Myriocin is a naturally occurring fungal metabolite that inhibits serine palmitoyltransferase (SPT), a vital enzyme in sphingolipid biosynthesis, and has been shown to have immunosuppressive properties. Using a murine acute pulmonary inflammation model as a surrogate inflammatory disease model, we evaluated the immune-modulatory effects of myriocin. Myriocin was administered at 0.3mg/kg, i.p., shortly after LPS challenge. Cellular infiltration into the lungs 24hrs and 48hrs after LPS challenge was significantly inhibited by myriocin. Macrophage, neutrophil, and lymphocyte numbers in the bronchoalveolar lavage fluid (BALf) were all significantly reduced by myriocin treatment as compared to the vehicle control group. Plasma ceramide levels were reduced 24hrs after myriocin treatment. There was no effect of myriocin on the pro-inflammatory cytokine TNFα, or chemokines KC and MIP-2 in the BALf. However, myriocin inhibited MCP-1 induced migration activity of BALf macrophages. These data provide support for the therapeutic modulation of the sphingolipid pathway as a strategy to suppress inflammation.
CD73-generated Adenosine Regulates Thrombomodulin Expression (94.29)
Authors: Qu Dongfeng , Esmon Naomi , Esmon Charles , Thompson Linda 
Published: January 2023

Abstract
Thrombomodulin (TM), as a thrombin modulator, plays a role in anti-coagulation. TM also plays an important role in regulating inflammation and vascular permeability. As TM expression levels are regulated by cAMP, and adenosine receptors (AR) modulate intracellular cAMP levels, we investigated whether TM expression can be regulated by AR signaling triggered by CD73-generated adenosine. Mouse endothelial cells were treated with either the general AR agonist NECA, or agonists specific for different AR subtypes, and TM expression levels were measured. Protein levels increased 2-fold in response to NECA; mRNA levels increased 4-fold. TM protein and mRNA levels also increased in response to A2AAR and A2BAR agonists, but not to A1AR or A3AR agonists. TM protein and mRNA levels also increased after AMP treatment and these increases were abrogated by a CD73 inhibitor. TM mRNA levels increased 4- to 6-fold in lung, heart, and liver of mice treated with NECA; protein levels increased 2-fold. Wild type and CD73-/- mice were treated with LPS and TM expression levels were measured in lung tissues. TM mRNA levels decreased further in lung tissue of CD73-/- mice than in wild-type mice after LPS treatment. These data suggest that AR signaling can modulate TM expression and that CD73-generated adenosine can be very important in regulating TM mRNA levels in vivo during an inflammatory response.
This work is funded by NIH grants AI18220 and P20RR016904.
Type-I interferon modulates monocyte recruitment and maturation in chronic inflammation (94.4)
Authors: Lee Pui Y , Li Yi , Kumagai Yutaro , Weinstein Jason , Nacionales Dina , Butfiloski Edward , Kellner Erinn , Akira Shizuo , Sobel Eric , Satoh Minoru , Reeves Westley 
Published: January 2023

Abstract
Chronic inflammation is characterized by continuous recruitment and activation of immune cells such as monocytes in response to a persistent stimulus. Production of pro-inflammatory mediators by monocytes leads to tissue damage and perpetuates the inflammatory response. However, the mechanism(s) responsible for the sustained influx of monocytes in chronic inflammation are not well defined. In chronic peritonitis induced by 2,6,10,14-tetramethylpentadecane (TMPD; pristane), the persistent recruitment of Ly6Chi inflammatory monocytes into the peritoneum was abolished in type-I interferon (IFN-I) receptor deficient mice but was unaffected by the absence of IFN-γ, TNF-α, IL-6 or IL-1. IFN-I signaling stimulated the production of chemokines (CCL2, CCL7, and CCL12) that recruited Ly6Chi monocytes via interactions with the chemokine receptor CCR2. Interestingly, after TMPD treatment, the rapid turnover of inflammatory monocytes in the inflamed peritoneum was associated with a lack of differentiation into Ly6Clo monocytes/macrophages, a more mature subset with enhanced phagocytic capacity. In contrast, Ly6Chi monocytes/macrophages differentiated normally into Ly6Clo cells in IFN-I receptor deficient mice. The effects of IFN-I were specific for monocytes as granulocyte migration was unaffected in the absence of IFN-I signaling. Taken together, our findings reveal a novel role of IFN-I in promoting the recruitment of inflammatory monocytes via the chemokine receptor CCR2. Continuous monocyte recruitment and the lack of terminal differentiation induced by IFN-I may help sustain the chronic peritoneal inflammatory response.
Differentiation of blood monocytes to dendritic cells following reverse transmigration across the blood-brain barrier (94.6)
Authors: Talreja Jaya , Prameya Rukmini , Dorovini-Zis Katerina 
Published: January 2023

Abstract
Dendritic cells (DC) are absent in the normal human central nervous system (CNS) but their presence has been documented in neuroinflammation. The origin of CNS DC is still controversial. The objective of this study was to determine whether monocytes that cross the blood-brain barrier (BBB) differentiate into DC. Primary human brain microvessel endothelial cells (HBMEC) were cultured on collagen gels. Purified human CD14+ monocytes were incubated with confluent HBMEC for 48h and the reverse transmigrated (RT) cells were characterized by FACS analysis. Approximately 50% of the monocytes reverse transmigrated across HBMEC monolayers and 70% of the RT cells expressed DC-SIGN. Monocytes incubated on gels in the absence of HBMEC retained their phenotype. The percentage of RT cells expressing CD83 increased from 25% to 70% when human CNS myelin was incorporated into the gels, and there was significant increase in CCR7 expression. Analysis of culture supernatants showed high levels of IL-6, TNF-α, PGE2 and IL-1β when HBMEC were cultured on myelin gels suggesting that these cytokines may contribute to the differentiation of monocytes to mature DC. There was an associated increase of MIP-3β, MCP-1, MIP-1α, and MIP-1β in the supernatants. Mixed lymphocyte reactions by co-culturing CFSE-labeled T cells and RT cells showed that RT cells were capable of stimulating allogeneic T cell proliferation. In conclusion, the results suggest that monocytes which reverse-transmigrate across HBMEC differentiate into mostly mature DC capable of inducing T cell proliferative responses. (Supported by MSS, Canada)