bluuebunny/crossref_metadata_2025_split huggingface dataset:
B Lymphocyte Selection and Age-Related Changes in VH Gene Usage in Mutant Alicia Rabbits
Authors: Zhu Xiaocui , Boonthum Anusorn , Zhai Shi-Kang , Knight Katherine L. 
Published: December 2022

AbstractYoung Alicia rabbits use VHa-negative genes, VHx and VHy, in most VDJ genes, and their serum Ig is VHa negative. However, as Alicia rabbits age, VHa2 allotype Ig is produced at high levels. We investigated which VH gene segments are used in the VDJ genes of a2 Ig-secreting hybridomas and of a2 Ig+ B cells from adult Alicia rabbits. We found that 21 of the 25 VDJ genes used the a2-encoding genes, VH4 or VH7; the other four VDJ genes used four unknown VH gene segments. Because VH4 and VH7 are rarely found in VDJ genes of normal or young Alicia rabbits, we investigated the timing of rearrangement of these genes in Alicia rabbits. During fetal development, VH4 was used in 60–80% of nonproductively rearranged VDJ genes, and VHx and VHy together were used in 10–26%. These data indicate that during B lymphopoiesis VH4 is preferentially rearranged. However, the percentage of productive VHx- and VHy-utilizing VDJ genes increased from 38% at day 21 of gestation to 89% at birth (gestation day 31), whereas the percentage of VH4-utilizing VDJ genes remained at 15%. These data suggest that during fetal development, either VH4-utilizing B-lineage cells are selectively eliminated, or B cells with VHx- and VHy-utilizing VDJ genes are selectively expanded, or both. The accumulation of peripheral VH4-utilizing a2 B cells with age indicates that these B cells might be selectively expanded in the periphery. We discuss the possible selection mechanisms that regulate VH gene segment usage in rabbit B cells during lymphopoiesis and in the periphery.
Expression of the Protein Kinase C Substrate Pleckstrin in Macrophages: Association with Phagosomal Membranes
Authors: Brumell John H. , Howard Jeffrey C. , Craig Karen , Grinstein Sergio , Schreiber Alan D. , Tyers Mike 
Published: December 2022

AbstractDespite evidence suggesting that protein kinase C (PKC) isoforms are important in phagocytosis by Fcγ receptors, the mechanisms by which the substrates of these kinases act are largely unknown. We have investigated the role of one PKC substrate, pleckstrin, in cells of the monocyte/macrophage lineage. Pleckstrin expression in mouse macrophages was induced severalfold in response to bacterial LPS and IFN-γ. In unstimulated cells, the protein was largely confined to the cytosol. Upon ingestion of IgG-opsonized zymosan particles (OPZ), however, pleckstrin accumulated on the phagosomal membrane. This association was transient, being maximal after 15 min and declining thereafter. Similar kinetics of association was also seen for both filamentous actin and the δ isoform of PKC. Ingestion of OPZ was found to induce phosphorylation of pleckstrin. To examine whether phosphorylation was required for phagosomal association, pleckstrin was expressed in CHO-IIA cells that stably express the FcγRIIA receptor and are competent for phagocytosis of OPZ. In these cells, both wild-type pleckstrin and mutants in which the phosphoacceptor sites had been mutated to either alanine (nonphosphorylatable) or glutamine (pseudophosphorylated) were found to accumulate on OPZ phagosomes. Thus, association of pleckstrin with phagosomes is independent of its phosphorylation. Our findings suggest that pleckstrin may serve as an intracellular adaptor/targeting protein in response to particulate stimuli. By targeting interacting ligands to the phagosomal compartment, pleckstrin may serve to regulate phagocytosis and/or early steps during maturation of the phagosome.
Role of 5-Lipoxygenase Products in the Local Accumulation of Neutrophils in Dermal Inflammation in the Rabbit
Authors: Marleau Sylvie , Fruteau de Laclos Bernard , Sanchez Ana B. , Poubelle Patrice E. , Borgeat Pierre 
Published: December 2022

AbstractStudies were undertaken to define the role of 5-lipoxygenase (5-LO) products and, in particular, of leukotriene (LT) B4 in the polymorphonuclear leukocyte (PMN) emigration process using a rabbit model of dermal inflammation. Our results show that i.v. administration to rabbits of MK-0591, a compound that inhibits LT biosynthesis in blood and tissues when administered in vivo, significantly reduced 51Cr-labeled PMN accumulation in response to intradermally injected chemotactic agonists, including IL-8, FMLP, C5a, and LTB4 itself. In addition, pretreatment of the labeled PMN with MK-0591 ex vivo before their injection in recipient animals was equally effective in reducing 51Cr-labeled PMN emigration to dermal inflammatory sites. These results support a role for de novo synthesis of 5-LO metabolites by PMN for their chemotactic response to inflammatory mediators. Other studies demonstrated that elevated intravascular concentration of LTB4 interferes with PMN extravasation inasmuch as a continuous i.v. infusion of LTB4, in the range of 5–300 ng/min/kg, dose-dependently inhibited extravascular PMN accumulation to acute inflammatory skin sites elicited by the chemoattractants LTB4, FMLP, C5a, and IL-8 and by TNF-α, IL-1β, and LPS; such phenomena may constitute a natural protective mechanism from massive tissue invasion by activated PMN in specific pathologic conditions such as ischemia (and reperfusion). These studies demonstrate additional functions of 5-LO products in the regulation of PMN trafficking, distinct from the well-characterized chemotactic activity of LTB4 present in the extravascular compartment.
A Deficiency of Placental IL-10 in Preeclampsia
Authors: Hennessy A. , Pilmore H. L. , Simmons L. A. , Painter D. M. 
Published: December 2022

Abstract
Accommodation of the fetoplacental unit in human pregnancy requires maternal immune tolerance to this “semiallograft”. Local antiplacental immunity is modified by synthesis of uncommon histocompatibility Ags (e.g., HLA-G), growth factors, and cytokines by the placenta. Placental interleukins have been identified in reproductive tissues, but their roles in adaptive maternal immunity and determining term pregnancy outcomes have not been fully clarified. This study examined the distribution of IL-10 and TNF-α staining in term placentas. Women with proteinuric hypertension (PE, n = 10) were compared with an age-matched group with normal pregnancy (NP, n = 14) and gestational hypertension (GH, n = 6). Using immunohistochemistry of parrafin-fixed tissues, trophoblast cells were identified by cytokeratin 7 and cytokeratin 18 staining. The cytokine binding of villous trophoblast cells was scored depending on the extent of circumferential cytoplasm staining (&lt;25%; intermediate or &gt;75%). The cytokine positive decidual cells were scored as a percentage of total extravillous trophoblast cells. There was a reduction in villous IL-10 immunostaining compared with normal term placenta (PE, 10.2 ± 1.1, mean ± SEM; NP, 14.07 ± 1.16 Mann-Whitney U test; p = 0.02). In these patients, there was an increase in TNF-α immunostaining. Sparse endovascular extravillous trophoblast cells demonstrated nuclear IL-10 staining in 30% of patients with preeclampsia. Serum IL-10 was diminished in women with preeclampsia compared with normal pregnancy. In conclusion, villous trophoblast demonstrated diminished immunostaining of IL-10 in preeclampsia. This abnormality may be associated with heightened maternal antifetal immunity and therefore inadequate placental development in preeclampsia.
Cloning the Antibody Response in Humans with Inflammatory Central Nervous System Disease: Analysis of the Expressed IgG Repertoire in Subacute Sclerosing Panencephalitis Brain Reveals Disease-Relevant Antibodies That Recognize Specific Measles Virus Antigens
Authors: Burgoon Mark P. , Owens Gregory P. , Smith-Jensen Tracey , Walker David , Gilden Donald H. 
Published: December 2022

Abstract
The presence of increased IgG in the brains of humans with infectious and inflammatory CNS diseases of unknown etiology such as multiple sclerosis may be a clue to the cause of disease. For example, the intrathecally synthesized oligoclonal bands (OGBs) in diseases such as subacute sclerosing panencephalitis (SSPE) or cryptococcal meningitis have been shown to represent Ab directed against the causative agents, measles virus (MV) or Cryptococcus neoformans, respectively. Using SSPE as a model system, we have developed a PCR-based strategy to analyze the repertoire of IgG V region sequences expressed in SSPE brain. We observed abnormal expression of germline V segments, overrepresentation of particular sequences that correspond to the oligoclonal bands, and substantial somatic mutation of most clones from the germline, which, taken together, constitute features of Ag-driven selection in the IgG response. Using the most abundant or most highly mutated γ H chain and κ or lambda L chain sequences in various combinations, we constructed functional Abs in IgG mammalian expression vectors. Three Abs specifically stained MV-infected cells. One Ab also stained cells transfected with the MV nucleoprotein, and a second Ab stained cells transfected with the MV-fusion protein. This technique demonstrates that functional Abs produced from putative disease-relevant IgG sequences can be used to recognize their corresponding Ags.
Expression, Regulation, and Function of B Cell-Expressed CD154 in Germinal Centers
Authors: Grammer Amrie C. , McFarland Richard D. , Heaney Jonathan , Darnell Bonnie F. , Lipsky Peter E. 
Published: December 2022

AbstractActivated B cells and T cells express CD154/CD40 ligand in vitro. The in vivo expression and function of B cell CD154 remain unclear and therefore were examined. Tonsillar B and T cells expressed CD154 at a similar density both in situ and immediately ex vivo, whereas a significantly higher percentage of the former expressed CD154. CD154-expressing B cells were most frequent in the CD38positiveIgD+ pre-germinal center (GC)/GC founder, CD38positive GC and CD38−IgD− memory populations, and were also found in the CD38−IgD+ naive and CD38brightIgD+ plasmablast subsets, but not in the CD38brightIgD− plasma cell subset. B cell expression of CD154 was induced by engaging surface Ig or CD40 by signals that predominantly involved activation of AP-1/NF-AT and NF-κB, respectively. The functional importance of CD154-mediated homotypic B cell interactions in vivo was indicated by the finding that mAb to CD154 inhibited differentiation of CD38positiveIgD− GC B cells to CD38−IgD− memory cells. In addition, mAb to CD154 inhibited proliferation induced by engaging sIg or CD40, indicating the role of up-regulation of this molecule in facilitating B cell responsiveness. Of note, CD154 itself not only functioned as a ligand but also as a direct signaling molecule as anti-CD154-conjugated Sepharose beads costimulated B cell responses induced by engaging surface Ig. These results indicate that CD154 is expressed by human B cells in vivo and plays an important role in mediating B cell responses.
Characterization of RNA Binding Proteins Associated with CD40 Ligand (CD154) mRNA Turnover in Human T Lymphocytes
Authors: Rigby W. F. C. , Waugh M. G. , Hamilton B. J. 
Published: December 2022

AbstractCD154 (CD40 ligand (CD40L)) has been demonstrated to play an essential role in the development of humoral and cellular immunity through its interaction with CD40. While earlier studies have examined the regulation of CD154 expression by transcriptional and posttranslational pathways, scant data exist on its regulation at a posttranscriptional level. In this report we demonstrate that CD154 mRNA is rapidly turned over in primary culture of activated human T lymphocytes. Moreover, we demonstrate that CD154 mRNA is unstable, but can be stabilized by treatment with either phorbol esters or calcium ionophores. To address this lability of CD154 mRNA, we examined the ability of cytoplasmic proteins to bind to its 3′ untranslated region (3′UTR). Two major proteins (p25 and p50) capable of binding the 3′UTR of CD154 were identified. The p25 binding activity was associated with polysomes and appeared to correlate with CD154 mRNA instability. Intriguingly, these proteins did not appear to bind to the AU-rich elements present in the 3′UTR of CD154. Rather, their binding was localized to unique sites between nt 471–811 of the 3′UTR, which lack any classical AU-rich elements. These data suggest that these proteins interact with distinct cis-acting elements that are important in the posttranscriptional regulation of CD154 expression. As such, identifying these proteins will help us understand the signals that are necessary for CD154 expression by activated T cells.
Failure of Rearranged TCR Transgenes to Prevent Age-Associated Thymic Involution
Authors: Lacorazza H. Daniel , Guevara Patiño Jose A. , Weksler Marc E. , Radu Dorel , Nikolić-Z̆ugić Janko 
Published: December 2022

AbstractAfter puberty, the thymus undergoes a dramatic loss in volume, in weight and in the number of thymocytes, a phenomenon termed age-associated thymic involution. Recently, it was reported that age-associated thymic involution did not occur in mice expressing a rearranged transgenic (Tg) TCRαβ receptor. This finding implied that an age-associated defect in TCR rearrangement was the major, if not the only, cause for thymic involution. Here, we examined thymic involution in three other widely used MHC class I-restricted TCRαβ Tg mouse strains and compared it with that in non-Tg mice. In all three TCRαβ Tg strains, as in control mice, thymocyte numbers were reduced by ∼90% between 2 and 24 mo of age. The presence or absence of the selecting MHC molecules did not alter this age-associated cell loss. Our results indicate that the expression of a rearranged TCR alone cannot, by itself, prevent thymic involution. Consequently, other presently unknown factors must also contribute to this phenomenon.
Up-Regulation of NK Cytotoxic Activity Via IL-15 Induction by Different Viruses: A Comparative Study
Authors: Fawaz Lama M. , Sharif-Askari Ehsan , Menezes José 
Published: December 2022

AbstractIL-15 is a recently identified cytokine that belongs to the four α-helix bundle cytokine family and possesses biological activities similar to those of IL-2. Its ability to induce effectors of NK activity suggests its involvement in innate immunity. In this study, we analyzed the effect of different viruses (HSV, EBV, respiratory syncitial virus, vesicular stomatitis virus, influenza virus, reovirus, and Sendai virus) on the up-regulation of NK activity in vitro. Exposure of human PBMC to the these viruses resulted in an immediate up-regulation of NK activity of PBMC via IL-15 induction; this effect was abrogated in the presence of mAbs to IL-15. Results of experiments conducted in parallel using mAbs to IL-15, as well as to other cytokines (IL-2, IL-12, IFN-γ, and TNF-α), clearly indicated that IL-15 was specifically responsible for the observed effect. Furthermore, supernatants of virus-infected PBMC cultures significantly enhanced NK activity of uninfected PBMC in vitro. An increase of IL-15 protein levels 20 h postinfection was also confirmed in a bioassay using the IL-2-dependent cell line CTLL. Kinetic analysis of IL-15 mRNA expression using a semiquantitative RT-PCR revealed that the level of IL-15 messages peaked at different time points (up to 12 h) postinfection, depending on the nature of the virus. Taken together, these results suggest that the IL-15 response of the host to viral infection and the subsequent NK cell activation represent an important effector mechanism of the innate immune surveillance of the host against viral infections.
Ectosomes Released by Human Neutrophils Are Specialized Functional Units
Authors: Hess Christoph , Sadallah Salima , Hefti Andreas , Landmann Regine , Schifferli Jürg-Alfred 
Published: December 2022

AbstractHere we show that human polymorphonuclear leukocytes (PMN) release ectosomes independently of complement attack during their activation both in vitro and at the site of inflammation in vivo. Patterns of biotinylated proteins on the surface of PMN and on PMN-derived ectosomes indicated a specific sorting of cell surface proteins into and out of ectosomes. Ectosomes expressed clusters of complement receptor 1 (CR1), which allowed them to bind efficiently to opsonized bacteria. Myeloperoxidase and human leukocyte elastase, both stored within the azurophilic granules of PMN, were found to colocalize on ectosomes with CR1. Furthermore, myeloperoxidase colocalized with human leukocyte elastase. In contrast, not present on CR1-expressing ectosomes were CD63, a selective marker for the azurophilic granules, and CD14, which is located within the same granules and the secretory vesicles as CR1. Of the other complement regulatory proteins expressed by PMN, only CD59 colocalized with CR1, while CD55 and CD46 were almost absent. Ectosomes released by activated PMN at the site of inflammation may function as a well organized element (ecto-organelle), designed to focus antimicrobial activity onto opsonized surfaces.
Normal Induction of Oral Tolerance in the Absence of a Functional IL-12-Dependent IFN-γ Signaling Pathway
Authors: Mowat Allan McI. , Steel Margaret , Leishman Andrew J. , Garside Paul 
Published: December 2022

AbstractThere is considerable evidence that regulatory cytokines play an important role in mediating the systemic tolerance found after oral administration of protein Ags. Although most existing work has focused on cytokines such as IL-4, IL-10, and TGF-β, recent evidence from TCR transgenic systems suggests that the induction of oral tolerance is accompanied by priming of Ag-specific IFN-γ production. IFN-γ has also been implicated as a mediator of T cell tolerance in other models in vivo and in vitro, including that induced by aerosol administration of protein. We show here that feeding tolerogenic doses of OVA primes for IFN-γ production in the spleen of mice with a normal T cell repertoire. However, depleting IFN-γ at the time of feeding OVA had no effect on the induction of tolerance. In addition, tolerance was induced normally in both IFN-γ receptor knockout (IFN-γR−/−) and IL-12 p40 knockout (IL-12−/−) mice. This was the case for all components of the systemic immune response and also with a variety of feeding protocols, including those believed to induce distinct regulatory mechanisms. We conclude that IL-12-dependent IFN-γ-mediated regulation does not play an essential role in oral tolerance.
Defective CD8+ T Cell Activation and Cytolytic Function in the Absence of LFA-1 Cannot Be Restored by Increased TCR Signaling
Authors: Shier Peter , Ngo Karen , Fung-Leung Wai-Ping 
Published: December 2022

AbstractSignaling through the TCR as well as engagement of costimulatory molecules are required for efficient T cell activation and progression into differentiated effector cells. The β2 integrin LFA-1 (CD11a/CD18) has been implicated in TCR costimulation as well as in cell-cell adhesion function, but its exact role is still ambiguous. The present study focuses on the requirement for LFA-1 in CD8+ T cell activation and effector function using LFA-1-deficient cells expressing the 2C transgenic TCR as a model system. The lack of LFA-1 expression in 2C T cells resulted in severely diminished proliferative response toward allogeneic BALB/c splenocytes. Increase in TCR signaling alone by pulsing stimulators with high affinity peptides, p2Ca or QL9, had minimal effects in restoring proliferation. Addition of exogenous IL-2, however, enhanced the effect of peptide pulsing on proliferation of LFA-1-deficient 2C T cells. LFA-1-deficient 2C CTLs generated from alloantigen stimulation exhibited a defective cytotoxic activity when tested on a variety of target cells. Cytolysis could be improved, but not fully rectified by peptide pulsing of target cells. Thus, in the 2C TCR model, LFA-1 has a requisite role for optimal CD8+ T cell activation and effector function, which cannot be overcome by increasing peptide/MHC density on either the APCs or target cells, respectively.
Influence of Lymphocytes on the Presence and Organization of Dendritic Cell Subsets in the Spleen
Authors: Crowley Mary T. , Reilly Christina R. , Lo David 
Published: December 2022

AbstractStudies were undertaken to clarify the roles of individual leukocyte populations in maintaining the presence and organization of splenic dendritic cells (DCs). Using Abs specific for DC subsets, we found that the distinct types of DC maintained appropriate compartmentalization within the white pulp of lymphocyte-deficient mice despite an unusual overall distribution of DCs. Even in mice lacking both B and T lymphocytes, the central arteriole remained the structure around which T area DCs were organized. Marginal zone area DCs remained in a peripheral sheath excluded from the T area DCs. Additionally, we revealed an important role for splenic B cells in the presence and organization of marginal zone cells. B-deficient or B- and T-deficient mice lacked sialoadhesin+ marginal zone macrophages and lacked MAdCAM-1 expression in marginal zone reticular endothelial cells. Adoptive transfer of B lymphocytes induced MAdCAM-1 expression but failed to recruit marginal zone macrophages. Taken together, our results demonstrate that the arrival, localization, and persistence of DCs in spleen are events not solely dependent upon signals from the mature B and T cells or marginal zone macrophages. We suggest that specific stromal elements in the vicinity of the central arteriole are primarily responsible for providing directional cues to the DC.
A Novel Role for H-Ras in the Regulation of Very Late Antigen-4 Integrin and VCAM-1 Via c-Myc-Dependent and -Independent Mechanisms
Authors: Liu Zhao-Jun , Tanaka Yoshiya , Fujimoto Hiroko , Mine Shinichiro , Morinobu Akio , Yagita Hideo , Okumura Ko , Oishi Isao , Udagawa Jun , Yamamura Hirohei , Minami Yasuhiro 
Published: December 2022

AbstractDespite extensive studies on the crucial functions of Ras and c-Myc in cellular proliferation and transformation, their roles in regulating cell adhesion are not yet fully understood. Involvement of Ras in modulating integrin activity by inside-out signaling has been recently reported. However, in contrast to R-Ras, H-Ras was found to exhibit a suppressive effect. Here we show that ectopic expression of a constitutively active H-Rasv12, but not c-Myc alone, in a hemopoietic cell line induces activation of very late Ag-4 (VLA-4, α4β1) integrin without changing its surface expression. Intriguingly, coexpression of H-Rasv12 and c-Myc in these cells results in not only the activation of VLA-4, but also the induction of expression of VCAM-1, the counterreceptor for VLA-4, thereby mediating a marked homotypic cell aggregation. In addition, H-Rasv12-induced VLA-4 activation appears to be partly down-regulated by coexpression with c-Myc. Our results represent an unprecedented example demonstrating a novel role for H-Rasv12 in the regulation of cell adhesion via c-Myc-independent and -dependent mechanisms.