bluuebunny/crossref_metadata_2025_split huggingface dataset:
New Applications for Bispecific Antibodies in Wound Healing (94.16)
Authors: Cousens Leslie  P. , Terry Frances E , Falanga Vincent 
Published: January 2023

Abstract
Bispecific antibodies (BiAbs) have potential to confer injury-specific targeting to cells for enhancing wound repair. Studies are designed to test BiAbs for their ability to direct recruitment and trafficking of specific cell populations (i.e. stem cells or macrophages) to sites of injury. A new series of BiAbs for wound repair applications have been chemically heteroconjugated: monoclonal antibodies (mAbs) for specific cell populations (i.e. F4/80 on macrophages, or c-kit on stem cells) were linked to mAbs against injury-associated antigens (i.e. VCAM, fibrinopeptide A). Heteroconjugation efficiency was measured by SDS-PAGE/densitometry, demonstrating final product composition of 66.75%, 58.76%, and 24.80% active heteroconjugates of F4/80xFibA, F4/80xVCAM, and cKitxVCAM, respectively. Specific BiAb binding to macrophage or stem cell populations was observed by flow cytometry. Functional BiAb binding was quantified in a novel biological assay. With these three distinct BiAbs successfully heteroconjugated and validated in vitro, studies are underway to test trafficking of BiAb-armed cells to injured tissue in murine models of wound repair: full thickness skin wounds and external muscle crush injury.
CalDAG-GEF-I is required for E-selectin-dependent slow leukocyte rolling (94.2)
Authors: Zarbock Alexander , Mueller Helena , Bergmeier Wolfgang , Lowell Clifford A. , Van Aken Hugo , Wagner Denisa D. , Ley Klaus 
Published: January 2023

Abstract
During leukocyte recruitment, leukocytes are exposed to different adhesion molecules and chemokines. E-selectin binding to PSGL-1 and chemoattractant binding to Gαi-coupled receptors can modulate the rolling velocity by activating β2-integrins. The guanine nucleotide exchange factor Ca2+ and diacylglycerol-regulated guanine nucleotide exchange factor I (CalDAG-GEF-I) catalyzes GDP for GTP exchange in Rap1 and 2 and thereby controls the activation of β-integrins. CalDAG-GEF-I-/- neutrophils have an adhesion and migration defect in response to chemokine activation. To test the role of CalDAG-GEFI in E-selectin-mediated slow rolling, we investigated calDAG-GEF-I-/- mice by intravital microscopy of the cremaster muscle and in autoperfused flow chambers. In the flow chamber, rolling velocity of neutrophils from WT mice on E-selectin was 2.2 µm/s and adding ICAM-1 to E-selectin reduced rolling velocity to 1.0 µm/s. This velocity reduction was partially abrogated in calDAG-GEF-I-/- mice, suggesting that this molecule is involved in E-mediated slow rolling. Mixed chimeric mice confirmed that calDAG-GEF-I-/- leukocytes had an elevated rolling velocity in TNF-α-induced inflammation in vivo. The physiologic importance of our findings is shown by near complete inhibition of neutrophil recruitment into the inflamed peritoneal cavity of calDAG-GEF-1-/- leukocytes treated with pertussis toxin to block Gαi signaling. These data show that calDAG-GEF-1 partially mediates slow rolling following E-selectin engagement.
Molecular interaction between CD99 and CD99-related proteins (94.27)
Authors: Ma Min-Jung , Lee Hae Young , Choi Eun Young 
Published: January 2023

Abstract
Functions of CD99 have been well known to be related to apoptosis and homotypic aggregation of thymocytes, and transendothelial migration of lymphocytes and monotypes. Moreover, CD99- like 2 protein (CD99L2), a protein with homology to CD99, has also been found to be involved in cellular diapedesis. Despite these well known cellular functions, information on the biochemical and molecular characteristics of CD99 and the related proteins is relatively rare. The molecules have been suggested to interact in a hemophilic fashion during the cellular transendothelial migration. We investigated molecular interaction among the mouse CD99-related proteins using bimolecular fluorescence complementation analysis (BiFC). In BiFC analysis, the dimerization-induced fluorescence was detected when CD99 formed homodimers, at the perinuclear region, and the dimerization was formed via extracellular region of the molecules. Not only CD99 formed homodimer, but also did CD99 form dimmers with mouse CD99L2 or with an alternatively-spliced version of CD99L2. But the distribution pattern of the fluorescence emitted when CD99 formed dimers with CD99L2 or with an alteratively-spliced version of CD99L2 was different from the pattern observed when the CD99 formed homodimers, with more dispersed distribution inside of the cells, implying the change in molecular interaction according to the cellular location of CD99. This result suggests that molecular interactions among the CD99-related molecules may control the cellular localization of the CD99.
Endoglycan is a CD34-related marker of marginal zone B cells and is upregulated by TLR ligands (94.7)
Authors: Hirukawa Alison , Merkens Helen , Frohwerk Erin J , Lam Mindy , David Muriel , Furness Sebastian GB , Hughes Michael R , McNagny Kelly M 
Published: January 2023

Abstract
Endoglycan is a CD34-related sialomucin proposed to play a role in selectin-dependent adhesion of leukocytes to vasculature. Here we have investigated the distribution and function of endoglycan by generating a specific monoclonal antibody to an extracellular epitope of the protein. On non-hematopoietic cells, endoglycan is a relatively selective marker of smooth muscle cells of the gut, airway, and vasculature. Surprisingly, endoglycan does not appear to be a marker of vascular endothelia. Within the mature lineages of the hematopoietic system, endoglycan is restricted to a subset of CD4+CD8+ thymocytes and a subset of mature peripheral B cells. In the spleen of naïve mice, the highest levels of endoglycan surface expression is on marginal zone B (MZB) cells with much lower expression on follicular B cells. To further explore the regulation of endoglycan expression we treated splenocytes with a panel of toll-like receptor (TLR) ligands and cytokines. LPS, Pam3CSK4, CpG and IL-1 all enhanced endoglycan surface expression on splenic B cells approximately four-fold within 48 hrs. Interestingly, TLR ligand-dependent upregulation was blocked in the presence of IL-4, IL-5 or TGF-β, suggesting a selective requirement for endoglycan in T cell independent responses.
This work was funded by an operating grant from the Heart and Stroke Foundation of Canada. EJF is a recipient of an NSERC post-graduate scholarship.
Expression analyses of CHST4, a carbohydrate sulfotransferase required for L-selectin ligand biosynthesis, suggests novel role in epithelial cells (95.10)
Authors: Hopkins Deborah A , Allred Sean , Pretorius James , Harrison Janine , Hale Kari , Trueblood Esther , Ramachandran Chidambaram 
Published: January 2023

Abstract
Slowed lymphocyte rolling along high endothelial venules (HEV) is initiated by the L selectin: PNAd interaction, as an essential first step in a cascade of reactions leading to extravasation of lymphocytes across HEV and into the lymphoid tissues. CHST4, a golgi resident carbohydrate sulfotransferase, has been described as an HEV restricted sulfotransferase in part responsible for the sulfation step required for L selectin: PNAd interaction. The MECA79 antibody which recognizes sulfated PNAd provides a useful tool for monitoring sulfotransferase activity. The presence of both CHST4 and the MECA79 sulfated epitope have been described in HEV as well as lymphoid aggregates and tertiary lymphoid tissues at the sites of chronic inflammation. For the studies described, we investigated the expression of CHST4 with real time PCR, In Situ Hybridization (ISH), Immunohistochemistry (IHC) as well as MECA79 IHC. Expression patterns were examined in an extensive collection of normal human tissue panels as well as diseased tissue collected from several inflammatory conditions. Resulting data, in addition to confirming an HEV and lymphoid aggregate restricted expression profile also revealed expression of the gene in epithelial and ductal epithelial cells in a small set of diverse organ tissues. The functional consequences of such expression and the co-localized MECA79 epitope expression are currently unknown.
CD146+ T lymphocytes are increased in both the peripheral circulation and in the synovial fluids of patients with various musculoskeletal diseases and may facilitate T and B interactions (95.11)
Authors: McCoy, Jr J Philip , Tatlici Gulnaz , Samsel Leigh , Raghavachari Nalini , Liu Poching , Liu Delong , Munson Peter J , Gourley Mark 
Published: January 2023

Abstract
The endothelial junction adhesion molecule CD146 appears on activated lymphocytes in vitro and in very low percentages of circulating lymphocytes in healthy individuals. Preliminary evidence suggests that CD146 facilitates adherence of these cells to the endothelium, and perhaps transendothelial migration. 24 synovial fluids (SF) were obtained from 21 patients with various arthritic conditions. Peripheral blood (PB) was obtained from 6 of these patients and PB from 23 healthy individuals. Our data indicate CD4+CD146+ cells accumulate in SFs preferentially compared to PB and these cells display a unique genotypic profile from their CD146- counterparts. Using Affymetrix HG-U133+ PM arrays, comparative data analysis between CD146 positive and negative SF CD4+ cells revealed differential expression of 230 probesets. CD146+ CD4+ cells compared to CD146- CD4+ cells displayed increased expression of numerous genes including collagen type VI, alpha 3, CXCL13, and CD109 and decreased expression CCR7 and cytotoxicity-associated genes such as granzymes h and k, perforin, and CCL5. Both CD109 and CXCL13 have been shown to mediate T cell - B cell interactions and the upregulation of these genes on CD146+ T cells in SF may indicate these cells facilitate such interactions.
IQGAP1 mediates lymphocyte cytoskeletal polarization and peripheral effector functions (95.16)
Authors: Gorman Jacquelyn A , Gomez Timothy S , Billadeau Daniel D 
Published: January 2023

Abstract
IQGAP1 is a scaffolding protein that integrates the actin and microtubule cytoskeletons. Although IQGAP1 has been shown to participate in numerous cellular activities in fibroblasts and epithelial cells including E-cadherin-mediated cell-cell adhesion, cell polarity, motility, proliferation, and MAP kinase signaling, its function in thymocyte development or peripheral T cell effector functions has not yet been investigated. IQGAP1 localizes to the immune synapse in CD4+ and CD8+ T cells suggesting a possible role in T cell activation. Using wild type and IQGAP1-deficient mice, we demonstrate that neither thymocyte development nor peripheral T cells numbers are affected in the absence of IQGAP1. Surprisingly, IQGAP1-deficient CD4+ T cells make significantly more IL-2 compared to wild type cells. Despite this, IQGAP1-deficient T cells show diminished T cell proliferation and increased cell death following TCR and CD28 ligation, which is most likely due to increased activation-induced cell death. Interestingly, IQGAP1-deficient T cells spread abnormally in response to anti-CD3, fail to accumulate F-actin or polarize their microtubule-organizing center (MTOC) toward anti-CD3 coated beads. Consistent with the latter observation, CD8+ T cells do show defective cell-mediated killing. Furthermore, we find that IQGAP1-deficient T cells fail to polarize following integrin stimulation. Taken together, our data identify IQGAP1 as an important mediator of proper cytoskeletal polarization during T cell activation. This work was supported by NIAID grant AI065474 to D.D.B. and NIH -T32-AI07425 pre-doctoral Immunology Training Grant to J.A. Gorman.
Unexpected role of selectin ligands in T cell trafficking to the lung (95.6)
Authors: Onami Thandi M. , Harp John R. 
Published: January 2023

Abstract
Selectin mediated tethering represents one of the earliest steps in T cell extravasation into lymph nodes via HEVs. Fucosyltransferases (FucT) -IV and -VII are required for production of the sialyl Lewis X moiety, which is essential for selectin mediated T cell entry into LNs. We examined T cell trafficking to lymphoid and non-lymphoid compartments such as the lung using FucT -IV and -VII double knockout (FucT DKO) mice. Examination of uninfected FucT DKO mice revealed significantly increased numbers of T cells localized in lungs, with no evidence of induced bronchial associated lymphoid tissue (iBALT). Following LCMV viral infection, we noted similar numbers of antigen specific CD8 T cells, but reduced numbers of total activated CD8 T cells in the lungs compared to wildtype mice. Finally, we transferred bona fide naïve or memory CD8 T cells into recipient mice and show increased trafficking of T cells into lungs of uninfected FucT DKO mice. We conclude from these studies that while selectin mediated entry may play an important role in T cell trafficking to the inflamed lung, in the absence of selectin ligand expression, T cells preferentially migrate to and are retained in the lung under non-pathological conditions. In situations where normal trafficking of T cells to LNs are compromised, tissues such as the lung may become sites for T cell accumulation, and this may play a role in the initiation of T cell mediated lung diseases.
This work was supported by NIH grants AI057719, GM62116 (CFG), and University of Tennessee start up funds
Characterization of a new monoclonal antibody to human IL-17A (96.14)
Authors: Bonnevier Jody L , deMedeiros Ricardo , Cheng Qi , Coley Kristen , Galitz David , Munshi Cyrus , LaBresh Kurt , Herrera Julio , Ni Jessie H.T. 
Published: January 2023

Abstract
Although IL-17A is made by CD4+ TH17 cells and there is an ever-increasing body of literature on these cells, other cells make IL-17A, including CD8+ (Tc17) and gamma-delta T cells, NK cells, and neutrophils. There are many factors that contribute to published findings of the percentage of cells producing IL-17A, including the activation or differentiation conditions, time course of activation, and antibody used for detection. It has been published that high levels of IL-17A are secreted by human PBMCs after anti-CD3/CD28-treatment alone for only 24 hrs. We have developed a new monoclonal Ab (clone 41802) that is specific for human IL-17A. The specificity of this Ab has been rigorously tested. Clone 41802 detects an increase in IL-17A in PMA/ionomycin-treated human PBMCs over resting cells by intracellular flow cytometry. This finding correlated with real-time PCR data in activated vs resting CD4+ PBMCs. This clone also detects IL-17A in a population of activated CD3+ PBMCs that is also positive for IL-22 and IL-23R by flow cytometry. Importantly, clone 41802 detects IL-17A in transfectant cells overexpressing human IL-17A, but not in cells overexpressing human IL-17F. The specificity of clone 41802 was further confirmed by western blot. In conclusion, clone 41802 is specific for human IL-17A. Although a larger percentage of IL-17A+ cells are detected in activated PBMCs than with other commercially available clones, this indicates an interesting biological finding, not a lack of antibody specificity.
Pro-inflammatory adjuvants enhance the cognate helper activity of aged CD4 T cells (96.9)
Authors: Eaton Sheri , Maue Alexander C , Lanthier Paula A , Sweet Kathryn B , Blumerman Seth L 
Published: January 2023

Abstract
The humoral response in aged individuals is significantly reduced compared to young, thus contributing to the reduced efficacy of vaccines commonly seen in older populations. Using an adoptive transfer model, we have shown that age-related intrinsic declines in the function of naïve CD4 T cells from aged donors contribute significantly to reduced humoral responses. This decline leads to reduced B cell expansion and differentiation as well as reduced IgG production. In this study, we show that the helper function of aged CD4 T cells can be enhanced using a Toll-like receptor (TLR) binding adjuvant or an adjuvant containing proinflammatory (PI) cytokines. In addition, we show that the helper function of aged CD4 T cells can be enhanced by adding PI cytokines during in vitro effector generation. We observed enhanced expansion and differentiation of B cells and affinity maturation of IgG. Importantly, we found that PI cytokines also induced significant production of a variety of effector cytokines, including IL-4, IFNg, IL-17, and IL-21, by both young and aged CD4 T cells. The use of PI adjuvants can also improve humoral responses in intact aged animals. We propose that one of the mechanisms involved in the ability of adjuvants to enhance both young and aged T cell responses includes driving multifaceted T cell differentiation and production of multiple cytokines by responding T cells. This work funded by NIH grants AG21054 and AG02160 to LH.
A regulatory B cell subset with a unique CD1dhiCD5+ phenotype controls T cell-dependent inflammatory responses (97.1)
Authors: Yanaba Koichi , Bouaziz Jean-David , Sato Shinichi , Tedder Thomas F 
Published: January 2023

Abstract
B cells mediate multiple functions that influence immune and inflammatory responses. While B cells contribute to disease in most autoimmune mouse models, we have also found that B cells have regulatory functions and inhibit inflammation. Specifically, T cell-mediated inflammation in a mouse model of contact hypersensitivity was exaggerated in CD19-deficient (CD19-/-) mice and wild type mice depleted of CD20+ B cells, while inflammation was substantially reduced in mice with hyperactive B cells due to CD19 overexpression (hCD19Tg). These inflammatory responses were negatively regulated by a unique CD1dhiCD5+ B cell subset that was absent in CD19-/- mice, represented only 1-2% of spleen B220+ cells in wild type mice, but was expanded to ~10% of spleen B220+ cells in hCD19Tg mice. Adoptive transfer of these CD1dhiCD5+ B cells normalized inflammation in wild type mice depleted of CD20+ B cells and in CD19-/- mice. Remarkably, IL-10 production was restricted to this CD1dhiCD5+ B cell subset, with IL-10 production diminished in CD19-/- mice, yet increased in hCD19Tg mice. Thereby, CD1dhiCD5+ B cells represent a unique subset of potent regulatory B cells that we have labeled as "B10 cells" due to their unique production of IL-10 and the fact that other regulatory B cell subsets are likely to exist.
Interleukin-21 mediates unanticipated suppressive effects via its induction of IL-10 (97.5)
Authors: Spolski Rosanne , Leonard Warren J. 
Published: January 2023

Abstract
Interleukin-21 (IL-21) is a type I cytokine produced by CD4+ T cells and NK T cells that regulates the function of T, B, NK, and myeloid cells. IL-21 can augment both humoral and cell-mediated immune responses through its effects on lymphoid differentiation and function, and we and others have demonstrated a critical role for IL-21 in the autoimmune NOD mouse model of diabetes. Elevated levels of IL-21 were previously correlated with development of disease in the BXSB-Yaa mouse model of systemic lupus erythematosus, and interestingly, these mice also had increased serum IL 10. We now demonstrate that IL-21 potently induces IL-10 production by both T and B cells, with increased IL-10 mRNA and protein production in IL-21 transgenic mice but reduced production in IL-21R KO mice. TCR priming in the presence of IL-21 led to the accumulation of cells with IL 10-dependent immunosuppressive activity on both T cell and B cell function. Furthermore, we demonstrate that IL-21 is required for optimal induction of IL-10 by either IL-6 or IL-27. These results identify a novel mechanism for the induction and amplification of IL-10 production in both naive and activated T cells, and moreover identify unexpected immunosuppressive actions for IL-21.
Involvement of a receptor tyrosine kinase in the activation of cAMP and ERK1/2 by a retroviral-derived immunosuppressive peptide (98.13)
Authors: Haraguchi Soichi , Cianciolo George J , Day Noorbibi K 
Published: January 2023

Abstract
Several endogenous and exogenous retroviruses contain an evolutionarily conserved amino acid sequence (putative immunosuppressive domain, ISD) within the ectodomain of their transmembrane envelope proteins. We investigated the physiological and pathological role of ISD using a synthetic biopeptide composed of 17 amino acid CKS-17 homologous to the highly conserved region and showed that it not only exhibited important immune-regulatory functions, but also inhibited the production of IL-2, IL-12, IFN-γ, and TNF-α, while enhancing IL-10. Moreover, CKS-17 activated several intracellular signaling molecules including elevation of cAMP levels and phosphorylation of ERK1/2. Since the activation of the cAMP and ERK1/2 pathways has been shown to favor the development of Th2-type immune responses as well as influence viral replication and infectivity, these findings provided an important molecular mechanism(s) responsible for the observed cytokine patterns and virus-associated pathogenesis. In the present study, we show that AG879 (an inhibitor to TrkA, ErbB2, and FLK-1) completely blocks CKS-17-induced elevation of cAMP levels and phosphorylation of ERK1/2 using a monocytic cell line THP-1. By contrast, other receptor tyrosine kinase inhibitors (AG825, AG1296, AG1478) do not have this effect. We also show that while PI3K inhibitors (wortmannin, Ly294002) reduce the activation of ERK1/2 by CKS-17, these inhibitors have no effect on the elevation of cAMP levels by CKS-17. These results suggest that the action of the ISD may be mediated by an AG879-sensitive receptor tyrosine kinase that induces the phosphorylation of ERK1/2 through, at least, PI3K but elevates cAMP levels independently of PI3K. Supported in part by the Eleanor Naylor Dana Charitable Trust.
Monocytic THP-1 cells stimulated by normal human serum (NHS) release cytokine-bearing plasma membrane-derived vesicles (PMVs), and can be inhibited by methyl-beta-cyclodextrin, calpeptin and Rho-kinase inhibitor, Y-27632 (98.27)
Authors: Ansa-Addo Ephraim A , Cestari Igor dos Santos , Pathak Paras , Ramirez Marcel I , Inal Jameel M. 
Published: January 2023

Abstract
Stimulation of human acute monocytic cells with sublytic complement leads to an increase of intracellular calcium resulting in the release of two forms of microvesicles, one derived from the surface membrane (between 0.1-1 μm), which we term plasma membrane-derived vesicles (PMVs) and exosomes (50-100nm) released by exocytosis. Much work has been documented on the release of PMVs, using various stimuli, and the roles they play in disease, but comparatively little is available on inhibition of PMV release. Although upon stimulation cells release exosomes in addition to PMVs, little has been reported on ways of differentiating between these two microvesicle types. Our aim was therefore to identify inhibitors of PMV release and to effectively differentiate between PMVs and exosomes. Methyl-β -cyclodextrin, an inhibitor of lipid rafts, calpeptin, which inhibits calpain and Rho-kinase inhibitor, Y-27632 were found to inhibit effectively the release of PMVs. Exosomes separated from PMVs by sucrose gradient were confirmed by electron microscopy and compared to PMVs were found to express less phosphatidylserine, but more CD63. Of the various cytokines carried by PMVs and exosomes, macrophage migration inhibition factor was found at a four-fold higher level in PMVs.
Source of Research Support: Royal Society, London.