bluuebunny/crossref_metadata_2025_split huggingface dataset:
An activation-responsive element in the murine IL-4 gene is the site of an inducible DNA-protein interaction.
Authors: Tara D , Weiss D L , Brown M A 
Published: December 2022

Abstract
IL-4 is a pleiotropic cytokine whose expression is limited to a subset of activated T cells and cells of the basophil/mast cell lineage. It plays a key role in regulating many immune responses; however, little is known about the intracellular signaling events that lead to the selective and transient IL-4 expression in either of these cell types. In this study, the molecular basis of stimulation-dependent transcription in T cells was explored. To identify cis elements that regulate IL-4 gene transcription, various amounts of the 5' flanking region of the murine IL-4 gene were linked to a chloramphenicol acetyl transferase (CAT) reporter gene and tested for the ability to modulate CAT gene transcription in PMA-stimulated EL-4 T cells. These experiments indicate that multiple positive and negative-acting elements contribute to the overall level of IL-4 transcription. These elements are located both proximal and distal to the transcription initiation site (TIS). An activation responsive element is located within 87 bp of the IL-4 gene TIS. This sequence is sufficient to confer responsiveness to PMA-mediated signals and results in a 10- to 20-fold induction of CAT reporter gene activity compared to activity detected in unstimulated cells. Proteins that specifically bind sequences within this region (-88 to -60) are detected in both unstimulated and stimulated EL-4 T cell nuclear extracts. An additional DNA-protein interaction is detected only when extracts from stimulated cells are analyzed. Base substitutions within the -88 to -60 sequence affect both transactivation function and protein/DNA interactions and demonstrate that sequences between -78 and -69 bp are critical. Together, these data support a model in which T cell activation signals stimulate binding of a nuclear protein(s) to a preexisting IL-4 DNA-protein complex. Proteins detected in these promoter proximal DNA-protein complexes are likely to be key elements in facilitating stimulation-dependent IL-4 transcription.
Pulmonary T helper lymphocytes are CD44hi, CD45RB- effector/memory cells in mice vaccinated with attenuated cercariae of Schistosoma mansoni.
Authors: Coulson P S , Wilson R A 
Published: December 2022

Abstract
Percutaneous exposure of C57Bl/6 strain mice to radiation-attenuated schistosome cercariae stimulates proliferation of Ag-specific T lymphocytes in the draining lymph nodes (LN), followed by an intense leukocytic infiltration into the pulmonary parenchyma and airways. The airway T cells persist at elevated levels at least up to 10 wk and will secrete IFN-gamma and IL-3 upon antigenic stimulation in vitro. We report here that more CD4+ T cells from the airways responded rapidly to mitogen by up-regulating the p55 subunit of the IL-2R than did splenocytes from the same animal, suggesting that the bulk of the pulmonary Th infiltrate comprised previously activated cells. A four-fold higher response to Ag indicated the greater abundance of schistosome-specific cells in the lungs than the spleen. Virtually all of the pulmonary CD4+ T cells expressed high levels of the memory marker CD44 (Pgp-1), in contrast with the situation in the draining LN and circulation where only a minority were in that category. Very few Th cells from the inflamed lung bound antibody to an epitope coded for by the B exon of CD45, unlike those from the LN and circulation; the antibody normally binds strongly to naive and weakly to memory CD4+ T cells. Overall, the airway CD4+ T lymphocytes in the schistosome-vaccinated mouse display the functional and phenotypic characteristics of short-term effector/memory cells. We believe that their presence endows the lung with the ability to respond rapidly to recall Ag in the form of challenge parasites.
Acute cerebral toxoplasmosis is induced by in vivo neutralization of TNF-alpha and correlates with the down-regulated expression of inducible nitric oxide synthase and other markers of macrophage activation.
Authors: Gazzinelli R T , Eltoum I , Wynn T A , Sher A 
Published: December 2022

Abstract
C57BL/6 mice infected with the ME-49 strain of Toxoplasma gondii develop a progressive encephalitis culminating in 100% mortality between 12 and 15 wk after intraperitoneal inoculation of the parasite. Moreover, when injected at 4 wk after infection with anti-IFN-gamma mAb, progression of toxoplasmic encephalitis is markedly accelerated, resulting in death of the animals by 9 to 12 days posttreatment. In this study, we investigated the expression of mRNAs encoding cytokines as well as lymphocyte and macrophage markers during the development of toxoplasmic encephalitis. High levels of lymphocyte CD4 and CD8 surface Ag transcript were detected in the brains of mice throughout the infection. In addition from 2 to 4 wk we found elevations of Th1 (IFN-gamma and IL-2) but not of Th2 (IL-4 and IL-5) cytokine mRNAs. The elevation in Th1 cytokines was accompanied by increases in the expression of monokine (IL-1, IL-6, IL-10, granulocyte macrophage-colony stimulating factor [GM-CSF], and TNF-alpha) mRNAs, as well as markers expressed by activated macrophages (major histocompatibility class II [Ia], inducible nitric oxide synthase [iNOS] and macrophage activation gene 1 [Mag-1]). Interestingly, after 8 wk of infection with T. gondii we observed a dramatic decrease of Th1 cytokine and most monokine (IL-1, IL-6, GM-CSF, and TNF-alpha) as well as Mag-1 and iNOS mRNA levels. This down-regulation was associated with enhanced necrosis and neutrophilic infiltrates in the brain accompanied by increased expression of genes expressed specifically by the tachyzoite stage of T. gondii (T. gondii surface antigen 1 [SAG-1] and T. gondii surface antigen 2 [SAG-2]). Similarly, in mice chronically infected with T. gondii and treated with anti-IFN-gamma mAb the resulting pathology was associated with decreased expression of TNF-alpha and iNOS and increased expression of SAG-1 and SAG-2. Moreover, treatment with anti-TNF-alpha mAb also resulted in enhanced pathology, which correlated with low levels of iNOS mRNA and high levels of tachyzoite-specific mRNAs. Together these results suggest that reactivation of T. gondii results from a down-regulation of IFN-gamma and TNF-alpha expression leading to decreased macrophage or microglial cell activation, release of parasite growth, and subsequent tissue damage.
IL-6 down-modulates the cytokine-enhanced antileishmanial activity in human macrophages.
Authors: Hatzigeorgiou D E , He S , Sobel J , Grabstein K H , Hafner A , Ho J L 
Published: December 2022

Abstract
IL-6 is a cytokine synthesized by T cells and macrophages (M phi). It has pleiotropic effects on diverse cell types and is recognized for its "pro-inflammatory" properties. In mice, IL-4, IL-5, IL-6, and IL-10 are produced by Th-2 cells. Because IL-10 suppresses Th-1 clones, and IL-4 broadly deactivates M phi, experiments were carried out to investigate the in vitro effects of recombinant human IL-6 on cytokine activation of human M phi. Pretreatment with IL-6 induced a dose- and time-dependent suppression of IFN-gamma (1000 U/mL) and TNF-alpha (25 ng/mL) activation of M phi for the killing of L. amazonensis. At doses greater than 0.1 to 100 ng/mL, IL-6 inhibited IFN-gamma and TNF-alpha activation by 21 to 93% and 36 to 82%, respectively. IL-6 alone had no effect on M phi viability and intracellular L. amazonensis growth. Blockade of M phi activation was greatest when IL-6 was added 24 or 48 h before infection and treatment with IFN-gamma or TNF-alpha. Furthermore, mAb against IL-6 abrogated the inhibitory activity of IL-6. Similarly IL-6 pretreatment suppressed M phi activation for antileishmanial capacity by IL-3, granulocyte-monocyte-CSF (GM-CSF) and IL-1 beta. Because cytokine induction of antileishmanial activity is associated with enhancement of oxidative capacity, the effect of IL-6 on this mechanism was evaluated. Pretreatment with IL-6 down-modulated TNF-alpha (25 ng/mL) enhancement of M phi oxidative capacity in a dose- and time-dependent manner. A similar depression of oxidative capacity was observed for GM-CSF and IL-3 but not for IFN-gamma. Furthermore, NG-monomethyl-L-arginine (a nitric oxide synthase inhibitor) had no effect on IFN-gamma and TNF-alpha activation of antileishmanial activity and nitrites/nitrates were not reliably assayed from M phi culture supernatants. These findings suggest that IL-6 down-modulates cytokine activation of M phi antileishmanial capacity by inhibiting oxygen-dependent and undefined oxygen-independent mechanisms.
IL-4- and IL-5-secreting lymphocyte populations are preferentially stimulated by parasite-derived antigens in human tissue invasive nematode infections.
Authors: Mahanty S , King C L , Kumaraswami V , Regunathan J , Maya A , Jayaraman K , Abrams J S , Ottesen E A , Nutman T B 
Published: December 2022

Abstract
Helminth infections in humans and animals are associated with strong T helper 2 (Th2) responses. To determine whether parasite-derived Ag preferentially expand a Th2-like cell population, a filter immunoplaque assay was used to enumerate the frequencies (F0) of PBMC and CD4(+)-enriched PBMC from individuals with helminth infections secreting selected cytokines in response to parasite-derived (PAg) and nonparasite antigens (NPAg). In 20 individuals with lymphatic filariasis, frequency analysis of PBMC secreting IL-4 and IFN-gamma indicated that the F0 of PAg-specific IL-4-secreting cells (geometric mean F0 (GM): 1/12,100) was 57-fold higher than the corresponding F0 of NPAg-reactive cells (GM: 1/692,000; p &lt; 0.02). In marked contrast, the F0 of IFN-gamma-secreting cells responding to PAg (GM: 1/2,700) did not differ from those of cells specific for NAPg (GM: 1/3,400; p = 0.83). In another group of helminth-infected individuals, the F0 of highly enriched CD4+ cells secreting IL-4 and IL-5 in response to PAg (GMs: 1/2,600 and 1/5,600 CD4+ cells, respectively) were also found to be significantly higher than those specific for NPAg (GMs: 1/291,000 and 1/303,000 CD4+; p &lt; 0.05 and p &lt; 0.01, respectively), whereas the corresponding F0 of IFN-gamma- and granulocyte-macrophage-CSF-secreting cells were equivalent for PAg and NPag. Furthermore, the proportion of PAg-specific IL-4- and IL-5-secreting CD4+ cells relative to all cells secreting the given cytokine were approximately 29-fold higher than the proportion of NPAg-specific cells secreting these cytokines. Again, the corresponding proportions of Ag-specific IFN-gamma-and GM-CSF-secreting CD4+ cells were equivalent for PAg and NPAg. Thus, in this ex vivo system, a circulating population of IL-4- and IL-5-secreting (Th2-like) cells has been shown to exist in humans; PAg appears to expand these cells preferentially.
Regulation of human lung mast cell function by phosphatase inhibitors.
Authors: Peachell P T , Munday M R 
Published: December 2022

Abstract
Okadaic acid, a cell-permeant inhibitor of serine/threonine phosphatases (PP) was found to attenuate the IgE-mediated release of both performed (histamine) and newly formed (leukotriene C4) mediators from human lung mast cells (HLMC). Optimal inhibition (82 +/- 6%) of the IgE-triggered release of histamine was observed after a 2-h incubation of HLMC with okadaic acid (1 microM). Shorter (&lt; 2 h) incubation times and lower (&lt; 1 microM) concentrations of okadaic acid were less effective at inhibiting histamine release from HLMC. The extent to which okadaic acid prevented histamine release was highly dependent upon the strength of the stimulus with lower levels of stimulation more readily modulated. The efficacy of okadaic acid as an inhibitor was not dependent upon the nature of the stimulus because matched levels of histamine release (approximately 30%) induced by either IgE-mediated processes or the calcium ionophore A23187 were attenuated to comparable extents. A series of analogues of okadaic acid and a structurally-distinct PP inhibitor, calyculin, were also found to inhibit the IgE-induced release of mediators from HLMC with the following rank order of potency; calyculin (approximate IC50; 0.015 microM) &gt; okadaic acid (0.2 microM) &gt; okadaol (3.3 microM) &gt; nor-okadaone (&gt; 10 microM). These same PP inhibitors displayed a similar rank order of activity for the inhibition of mediator release induced by the ionophore A23187. Extracts of purified HLMC were found to liberate 32P from radiolabeled glycogen phosphorylase and this PP activity was inhibited by both low (2 nM) and high (5 microM) concentrations of okadaic acid suggesting that HLMC contain both PP type 2A (PP2A) and PP type 1 (PP1). The okadaic acid, analogues of okadaic acid, and calyculin attenuated mediator release at concentrations and with a rank order of activity that closely parallels their activities as inhibitors of PP suggests that PP are important in the regulation of HLMC function. This contention is further supported by the finding that PP activities are constitutively associated with HLMC and that these activities could be inhibited by okadaic acid.
Mapping studies reveal unique epitopes on IgG recognized by rheumatoid arthritis-derived monoclonal rheumatoid factors.
Authors: Bonagura V R , Artandi S E , Davidson A , Randen I , Agostino N , Thompson K , Natvig J B , Morrison S L 
Published: December 2022

Abstract
We have used chimeric IgG antibodies and their genetically engineered variants prepared by a combination of site-directed mutagenesis and exon exchange to define the structure(s) on IgG recognized by monoclonal rheumatoid factor (RF) autoantibodies from rheumatoid arthritis (RA) patients. Nineteen RF produced by EBV-transformed cell lines from the synovium or blood of RA patients were analyzed. Their binding patterns differ significantly from those seen with RF obtained from patients with Waldenstrom's macroglobulinemia (WMac). Half of the RA-derived RF bound IgG1, 2, and 4, but not 3 (Ga specificity), the common pattern in WMac. However, heterogeneity in fine specificity within the Ga reactivity pattern was observed. Moreover, seven others bound all four IgG subclasses, a pattern observed for only one WMac-derived RF from a patient who also had RA. Three RF had subclass specificities unlike any observed with WMac-derived RF. Most RA-derived RF bound IgG at a discontinuous epitope comprised of residues from both the CH2 and CH3 H chain constant regions. However, unlike any WMac-derived RF, one RA-derived RF bound IgG in CH2, another in CH3, and a third at an undetermined site outside of the CH2-CH3 interface. Some RA-derived RF bound aglycosylated IgG4 less well than glycosylated IgG4, suggesting that the carbohydrate moiety was important in establishing their binding epitope in CH2. These studies demonstrate that the repertoire of RF expressed by RA patients contains some unique binding specificities for IgG epitopes not found among our panel of WMac-derived RF. Our results therefore call into question whether WMac-derived RF with their limited diversity are appropriate models for disease-related RF. In addition, RF with their multiple specificities can serve as probes of antibody structure.
Alkali hydrolysis of recombinant proteins allows for the rapid identification of class I MHC-restricted CTL epitopes.
Authors: Gavin M A , Gilbert M J , Riddell S R , Greenberg P D , Bevan M J 
Published: December 2022

Abstract
The characterization of the epitopes recognized by CTL provides insights into the nature of protective immune responses and facilitates the development of methods to enhance immunity to human pathogens. However, no easily applicable approach for CTL epitope identification has been developed. We present a rapid and efficient method for locating CTL epitopes within a protein. The gene encoding the protein of interest is inserted into an inducible prokaryotic expression vector. Random peptides are then generated by alkali digestion of intact or lysed Escherichia coli expressing the protein and assayed for the presence of the epitope by coating target cells for a standard CTL targeting assay. A large panel of clones containing serial 3'-deletions of the gene is then generated by exonuclease III digestion, and the expressed truncated proteins are similarly analyzed for the presence of the antigenic peptide. The epitope is located by determining the deletion points of clones expressing sequential truncations and differing in Ag expression. This technique was used to identify the H-2Ld-restricted nonamer in E. coli beta-galactosidase, with residues 876-884 representing the naturally processed epitope. To test the applicability of this method to other proteins, two genes from human CMV, an often fatal pathogen in immunocompromised individuals, were screened for HLA class I-restricted epitopes. An HLA-B18-restricted epitope from the CMV major immediate-early protein was found to lie between residues 378 and 389, and an HLA-B35-restricted epitope from the CMV pp65 matrix protein was characterized as residues 123 to 131. The results demonstrate that this technique can be used to rapidly identify CTL epitopes within a chosen protein and should be useful for assaying viral isolates or neoplasms for loss of epitopes after mutation and selection by host immune responses.
Heterogeneous populations of class II MHC+ cells in human dermal cell suspensions. Identification of a small subset responsible for potent dermal antigen-presenting cell activity with features analogous to Langerhans cells.
Authors: Meunier L , Gonzalez-Ramos A , Cooper K D 
Published: December 2022

Abstract
Little is known regarding the identification, classification, and function of class II MHC+ dendritic cells in the perivasculature of human connective tissues, such as the dermis. We developed a method for preparing papillary dermal cell suspensions from human keratome strips. Among the class II MHC+ populations of the dermis identified using triple color flow cytometry, cells of monocyte/macrophage lineage (CD45+ CD1- CD11b+ CD11clo-mid CD32+ CD36+ or - CD11a-) and mesenchymal cells of non-bone marrow origin (CD45-) were identified and characterized. Another distinct class II MHC+ subset was identified, which expressed a number of features analogous to epidermal Langerhans cells (LC) and other dendritic APC. These were a numerically minor population comprising only 2.7% +/- 1% (n = 7) of dermal cells. Like LC, they express HLA-DR, CD45, CD1a (albeit at a lower level of expression), CD1c, and CD32 and lack constitutive CD11a or ICAM-1. In contrast to LC, this dermal CD1a+CD1c+ subset expresses CD1b, CD11b, a higher level of CD11c, and intracytoplasmic factor XIIIa. Alloantigen presentation by unfractionated dermal cells was reduced by prior removal of this CD1b+ subset to the same degree achieved by removal of the entire DR+ population (20% of dermal cells), indicating that this was the critical DR+ subset. Cocultures of CD4+ T lymphocytes with cells sorted by flow cytometry into CD1c+DR+, CD1c-DR+ and DR- dermal cell subsets positively identified the CD1c+DR+ population as the most potent of potential APC subsets in human dermis. Thus, in distinction to other dermal macrophage and mesenchymal subsets with elongate morphology, the CD1aloCD1b,c+CD11c(hi)CD11b+CD32+DR+ population in human dermis is highly analogous to cells of LC/dendritic APC lineage in its phenotype and in its exclusive ability to potently present Ag to T lymphocytes. These studies identify and characterize the APC subset most potent in inducing activation of T cells initially entering the perivasculature of human dermis to be of LC/dendritic APC, and not tissue macrophage, lineage.
Analysis of Ig H chain gene segment utilization in human fetal liver. Revisiting the "proximal utilization hypothesis".
Authors: Pascual V , Verkruyse L , Casey M L , Capra J D 
Published: December 2022

Abstract
The utilization of Ig H chain gene segments during ontogeny is postulated to result from an immature recombination machinery that preferentially rearranges genes according to chromosomal position. We have tested the "proximal utilization hypothesis" using the two functional members of the VH5 family as a model. Nucleotide sequence analyses of transcripts involving these VH gene segments in 11- to 12-wk fetal liver samples revealed that they were utilized frequently, even though they are 500 kb apart. Although 50% of the H chain rearrangements in our study use the DQ52 gene segment, the most 3' end proximal human D segment, the preference for rearranging D gene segments based on 3' end proximity does not apply to other D segments because different members of the DLR family that are interspersed throughout the D locus are equally rearranged in our sample. The recurrent utilization of the DQ52 and JH4 gene segments and a propensity to preserve the two nucleotides flanking the 3' end of the VH gene segment to generate the amino acid residue at the V-D junction appear to be the major biases in the generation of an otherwise diverse fetal repertoire.
TNF-alpha, IL-1 beta, and hepatocyte growth factor cooperate in stimulating specific acute phase plasma protein genes in rat hepatoma cells.
Authors: Baumann H , Morella K K , Wong G H 
Published: December 2022

Abstract
In rat hepatoma H-35 cells, TNF-alpha, like IL-1 beta, stimulates the synthesis and secretion of type 1 acute phase plasma proteins, including alpha 1-acid glycoprotein (AGP), complement component 3 (C3), hemopexin, and haptoglobin. TNF and IL-1 in combination act additively to synergistically on the expression of AGP and C3 but not on hemopexin and haptoglobin. The cytokine stimulation of AGP and C3 genes is further enhanced by hepatocyte growth factor. The effect of TNF is mediated by the type I TNF receptor as judged from the TNF-like effect elicited by the agonist antibodies to type I but not type II TNF receptor. The data suggest that, although TNF and IL-1 share considerable overlap in their signal transduction mechanism, cytokine-specific signal pathways exist that affect a subset of acute phase plasma protein genes. A potential regulatory role is attributed to the transcription factors C/EBP beta and NF-kB based on studies on transiently transfected H-35 cells.
C1Q, a subunit of the first component of complement, enhances the binding of aggregated IgG to rat renal mesangial cells.
Authors: van den Dobbelsteen M E , van der Woude F J , Schroeijers W E , Klar-Mohamad N , van Es L A , Daha M R 
Published: December 2022

Abstract
Previous reports have shown the presence of C1Q-R on monocytes, macrophages, polymorphonuclear cells, fibroblasts, platelets, lymphocytes, and endothelial cells. The present study demonstrates a functional C1Q-R on rat renal mesangial cells (MC). Incubation of MC with increasing concentrations of [125I]C1Q resulted in a dose-dependent binding of [125I]C1Q to MC; the binding of [125I]C1Q was inhibitable by excess unlabeled C1Q or C1Q stalks whereas BSA and C1Q globular heads had no effect. Scatchard analysis of the data revealed the presence of 6.2 x 10(7) binding sites/cell with an affinity of 4.9 x 10(6) M-1 for C1Q. Immunoprecipitation of 125I-labeled MC membrane proteins with C1Q or mAb directed against human C1Q-R revealed a single 66- to 68-kDa band under reducing conditions. We have shown previously that soluble stable aggregates of IgG bind to rat MC in a dose-dependent fashion. In addition the presence of a receptor for IgG has been described on rat MC. In order to find out whether there is a cooperative effect between C1Q and AlgG in binding of [125I]AlgG to MC, we incubated [125I]AlgG in the presence of increasing concentrations of C1Q, and showed a 5- to 15-fold enhancement of binding of [125I]AlgG to MC. Neither heat-inactivated C1Q nor C1Q stalks were able to enhance the binding of [125I]AlgG to MC. Enhanced binding by C1Q was only observed when aggregated IgG was used; the binding of monomeric IgG to MC was not affected by C1Q. These studies indicate that there is a cooperative effect between Fc gamma R and C1Q-R on MC in the recognition of immune complexes.