bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation of human basophils through the IL-8 receptor.
Authors: Krieger M , Brunner T , Bischoff S C , von Tscharner V , Walz A , Moser B , Baggiolini M , Dahinden C A
Published: December 2022

IL-8 and its structural analogs derived from blood platelets have been proposed as stimuli of IgE-independent basophil activation. In order to clarify the mechanism of action of these peptides, we examined the effects of pure IL-8, connective tissue-activating peptide III (CTAP-III), neutrophil-activating peptide 2 (NAP-2), and platelet factor 4 (PF-4) on blood basophils with and without pretreatment by IL-3, which modulates mediator release. After pretreatment with IL-3, significant histamine release was observed with 10(-8) M and 10(-7) M IL-8 and 10(-7) M NAP-2, but not with the other peptides. At higher concentrations (10(-6) M), however, all IL-8 analogs, as well as the unrelated cationic peptides poly-D-lysine, histone VS, and lysozyme, induced histamine release to variable degrees. Binding and competition studies with [125I]IL-8 revealed specific IL-8R on basophils from a patient with chronic myelogenous leukemia and normal individuals. From 3500 to 9600 receptors with a mean Kd value of 0.15 nM were found on average per chronic myelogenous leukemia and normal basophil, respectively. NAP-2 weakly competed for IL-8 binding. IL-8 and, to a lesser extent, NAP-2 led to a transient rise of cytosolic free calcium concentration ([Ca2+]i), which was independent of a preexposure to IL-3. IL-8 prevented the [Ca2+]i rise induced by NAP-2, but did not influence [Ca2+]i responses to other agonists, e.g. C5a, C3a, or platelet-activating factor. IL-8 induced [Ca2+]i changes and histamine release in IL-3-primed basophils were pertussis toxin sensitive. CTAP-III or PF-4 did not compete for IL-8 binding, did not induce [Ca2+]i changes, and did not influence the [Ca2+]i response to IL-8 and NAP-2. This study shows that IL-8 and NAP-2 activate human basophils by a receptor-mediated mechanism similar to that operating in neutrophils. At high concentrations histamine release can also be induced by cationic peptides by a mechanism that does not involve the IL-8R, and probably depends on cationic interactions.
Localization of IFN-gamma receptor in first trimester placenta to trophoblasts but lack of stimulation of HLA-DRA, -DRB, or invariant chain mRNA expression by IFN-gamma.
Authors: Peyman J A , Hammond G L
Published: December 2022

Trophoblasts do not express MHC class II Ag on their cell surface, but it is not known at which level the regulation of expression of these IFN-gamma-responsive genes occurs. We localized the IFN-gamma R Ag to the trophoblasts of normal human first trimester placenta. As previously shown, treatment with IFN-gamma of short term cultures of placental explants did not induce expression of HLA-DR Ag in trophoblasts. We demonstrated that in situ hybridization with probes for DRA, DRB, and MHC class II-associated invariant chain genes gave no detectable signals for the presence of the corresponding mRNA in cytotrophoblasts or syncytiotrophoblasts of first trimester placenta with or without IFN-gamma. Maternal leukocytes in these placental preparations as well as IFN-gamma-treated HeLa cell control cultures expressed these mRNA. In addition, Northern analysis of RNA isolated from the trophoblast cell line Jar after treatment with IFN-gamma showed that induction of transcription did not occur from the HLA-DRA, HLA-DRB, or invariant chain genes. Jar cells expressed the IFN-gamma R Ag. These data suggest that down-regulation of MHC class II Ag on trophoblasts in the first trimester placenta and in a cell line include a lack of productive intracellular signaling by the IFN-gamma R and maintenance, in a coordinate manner, of low steady state levels of the mRNA encoded by the DRA, DRB, and invariant chain genes.
Role of cathepsin D in antigen presentation of ovalbumin.
Authors: Rodriguez G M , Diment S
Published: December 2022

Modification of protein Ag by proteolysis is one of the principal steps in the presentation of Ag to Th cells. However, little is known about the enzymes participating in these events, their specificity or the characteristics of the natural fragments that they produce. Cathepsin D (CD) is an aspartyl protease identified in endosomes of APC. In this report, the role of CD in the processing of OVA has been investigated. OVA digested in vitro with purified CD was able to stimulate IL-2 secretion by three different OVA-specific I-Ad restricted Th cell hybridomas when it was presented by fixed APC. The digest of OVA was recognized in the context of I-Ad, but not by I-Ak-restricted OVA-specific Th cells. No difference was observed in the ability of OVA digested with CD to stimulate Th cells in the absence of FCS or in the presence of protease inhibitors indicating that extracellular proteases were not likely to contribute to processing of OVA. Taken together, these results suggest that CD is necessary and sufficient for the generation of an antigenic epitope from OVA. A fragment containing the epitope was isolated from the OVA digest by reverse phase HPLC. This fragment, which migrates in SDS-PAGE as a 10-kDa polypeptide, is a potent epitope. Its capacity to activate Th cells is compared to that of the tryptic peptide OVA323-339.
Regulation of transcription of the germ-line Ig alpha constant region gene by an ATF element and by novel transforming growth factor-beta 1-responsive elements.
Authors: Lin Y C , Stavnezer J
Published: December 2022

Inasmuch as transcription of unrearranged, or germ-line, Ig CH genes appears to direct switch recombination, understanding the regulation of this transcription is essential for understanding the regulation of class switching. Transforming growth factor-beta 1 (TGF-beta 1) induces germ-line alpha transcripts and increases class switching to IgA in the I.29 mu B lymphoma and in Peyer's patch and splenic B cells. It has been previously demonstrated that induction of germ-line alpha transcripts by TGF-beta occurs at the transcriptional level in I.29 mu cells. We now demonstrate that the DNA segment located 5' to the initiation sites of germ-line alpha RNA drives expression of a luciferase reporter gene construct in transient transfection experiments. Full constitutive expression requires no more than 106 bp of the 5' flanking segment. By creating a series of deletion and substitution mutations, we have demonstrated that an ATF/CRE site residing within this region is very important for constitutive expression of the germ-line alpha promoter, but mutation of this motif does not diminish TGF-beta induction. Inducibility by TGF-beta requires additional sequences residing between -128 to -106 relative to the first RNA initiation site. Two copies of a tandemly repeated sequence 5' CA-CAG(G)CCAGAC 3' (termed Ig alpha TGF-beta-RE) are located in the region from -127 to -105. An oligonucleotide containing multimers of these repeats confers TGF-beta inducibility to a heterologous promoter. An additional copy of the TGF-beta-RE was identified at -41/-30 and its deletion reduces the TGF-beta response. Thus, we conclude that tandem repeats of a novel TGF-beta-RE are the positive regulatory element for the TGF-beta response. Our study provides further evidence that TGF-beta directs class switching to IgA through induction of transcription of the germ-line C alpha gene and demonstrates that TGF-beta can activate the promoter for the germ-line alpha gene.
Independent and synergistic effects of disulfide bond formation, beta 2-microglobulin, and peptides on class I MHC folding and assembly in an in vitro translation system.
Authors: Ribaudo R K , Margulies D H
Published: December 2022

We have examined the post-translational processing, intrachain disulfide bond formation, folding, and assembly of MHC class I H chains with beta 2-microglobulin after coupled in vitro translation of homogeneous mRNA and transport of nascent chains into canine microsomal vesicles. The formation of native alpha 3 domain conformation was dependent on conditions that optimized intrachain disulfide bond formation, and efficient folding of the alpha 1 alpha 2 domain required exposure to antigenic peptide. beta 2-microglobulin and peptide acted synergistically in forming native alpha 1 alpha 2 domain structure, and a small proportion of molecules with native alpha 1 alpha 2, but non-native alpha 3 structure were detected, indicating that alpha 3 domain folding is not an absolute prerequisite for the formation of native alpha 1 alpha 2 domain structure.
Inhibition of macrophage protein kinase C-mediated protein phosphorylation by Leishmania donovani lipophosphoglycan.
Authors: Descoteaux A , Matlashewski G , Turco S J
Published: December 2022

The cell surface lipophosphoglycan (LPG) from Leishmania donovani promastigotes is a potent inhibitor of purified protein kinase C (PKC) activity in vitro. In this study, we have investigated the effect of LPG on the activation process of PKC in murine bone marrow-derived macrophages. The extent and kinetics of calcium ionophore A23187-induced [3H] phorbol dibutyrate binding to macrophages were not affected by LPG pretreatment or infection with either wild-type or LPG-deficient promastigotes, indicating no effect on the association of calcium-dependent PKC with the plasma membrane. In contrast, LPG inhibited the phosphorylation of both the PKC-specific VRKRTRLLR substrate peptide and MARCKS, and endogenous PKC substrate, in 1-oleoyl-2-acetyl-glycerol-stimulated macrophages. These observations provide direct evidence that LPG effectively inhibits PKC activity in intact macrophages. Finally, depletion of PKC rendered macrophages more permissive for the proliferation of L. donovani, suggesting that inhibition of PKC-dependent events contributes to the survival of this parasite within its host cell.
Studies in Specific Hypersensitiveness
Authors: Grove Ella F.
Published: December 2022

The assumption that hay-fever could be explained on the basis of anaphylactic hypersensitiveness (1) and the suggestion (2) that the dyspnea of the asthmatic patient resembles the bronchospasm of the guinea pig in severe anaphylactic shock, were the starting points of a number of investigations into the mechanism of human hypersensitiveness.

Many workers claim to have demonstrated anaphylactic antibodies in the serum of human individuals afflicted with the diverse forms of hypersensitiveness (3, 4, 5, 6, 7), but the results are in general unconvincing (8, 9).

In 1921 Prausnitz and Küstner (9) discovered an important feature of atopic hypersensitiveness in a case of an individual sensitive to fish. They were able to demonstrate sensitizing substances in the blood of this individual by local passive transfer not to the skin of the guinea pig but to the skin of another human individual not sensitive to fish.

A most interesting and suggestive case was reported by Ramirez (12).
The Effect of Formaldehyde on Specific Complement Fixation Systems
Authors: Reyner C. E.
Published: December 2022

In work already reported (1) (2) it has been shown that formaldehyde added in the proper concentration to diluted Wassermann antigen has a uniformly constant effect in enhancing the capacity of the system to fix complement. Under such conditions not only is the total amount of complement to be fixed very definitely increased, but the velocity at which the fixation takes place is markedly accelerated. These effects can not be demonstrated through any action of the formaldehyde upon the antigen, the reagin, or the complement alone, but are only manifest when the formaldehyde operates upon the entire antigen-reagin-complement complex and even here a period of incubation is essential. Wassermann systems so enhanced in activity lose none of their apparent specificity and if quantitative relationships are properly adjusted there is no evidence that false reactions take place.

With the antigen-reagin complex as used in the Wassermann system it seemed to be impossible to demonstrate the mechanism through which the formaldehyde acted.
Anaphylaxis
Authors: Downs Cornelia M.
Published: December 2022

Turtles may be actively sensitized to mammalian serum. The heart in situ responds in a specific characteristic way to injection of the antigen used for sensitization. Desensitization can be demonstrated. Precipitins are present in the serum of a certain number of the actively sensitized animals and this serum seems to confer passive anaphylaxis. The specific response resembles vagus stimulation of the turtle heart.
Experimental Inoculation of Chickens with Hodgkin's Nodes
Authors: L'Esperance Elise S.
Published: December 2022

Five healthy chickens were inoculated intravenously with emulsions of lymph nodes from 2 cases of clinically and histologically characteristic Hodgkin's disease.

All of these chickens developed either a typical or an atypical tuberculosis and in the tissue smears, stained by Ziehl-Neilsen method, acid-fast granules and rods, extra- and intracellular, were demonstrated in two of them, and non-acid-fast granules in one of them.

Reinoculation of material from the lesions of one of these chickens into another chicken produced an identical though more extensive manifestation of the disease.

An atypical tuberculosis developed in a guinea pig by inoculation of the tissue from the fourth chicken. A pure growth of bacteria with the staining and cultural characteristics of the avian tubercle bacillus was produced on egg media from material from this guinea pig.

Cultures from these birds on egg media were contaminated in all instances. In one there was a symbiotic growth of a blastomyces-like organism with an acid-fast bacillus.
Studies on the “Toxins” of Hemolytic Streptococci
Authors: Ando K. , Kurauchi K. , Ozaki K.
Published: December 2022

Summary and Conclusions

Hemolytic streptococci isolated from pathologic foci, scarlatinal and non-scarlatinal, produce a toxic principle capable of evoking a cutaneous reaction in human subjects. Production of toxin varies considerably according to strains. Generally speaking, strains of hemolytic streptococci isolated from scarlet fever and erysipelas cases have been found to produce a toxic principle capable of evoking a marked cutaneous reaction in a larger proportion of human subjects tested than in the case of the toxins produced by suppurative streptococci. The toxins produced by various strains isolated from the same pathological conditions, e.g., scarlet fever, erysipelas or other suppurative lesions, may differ qualitatively when tested simultaneously on the same subject. If, therefore, only a small number of persons were tested simultaneously with two monovalent toxins of streptococcus hemolyticus from the same pathological conditions, the reactions to both toxins might be entirely similar or entirely dissimilar to each other. This dissimilarity may become more marked, if scarlatinal, erysipelas and suppurative toxins were compared in their monovalent form. But, in comparing the scarlatinal, erysipelas and suppurative toxins by skin reaction, we employed polyvalent toxin and demonstrated that the reactions to them were similar to the majority of cases tested. This similarity was marked in the greatest degree in the case of scarlatinal and erysipelas toxin, and marked in a lesser degree in the case of erysipelas and suppurative toxin, while it was not marked in the case of scarlatinal and suppurative toxin. The effectiveness of the toxin for both Dick test and preventive immunization depends largely on the selection of suitable strains. It seems to be reasonable to use polyvalent toxin by mixing toxins of several strains which are most potent in strength and inclusiveness.
Studies on the “Toxins” of Hemolytic Streptococci
Authors: Ando K. , Kurauchi K.
Published: December 2022

We have prepared three sorts of polyvalent toxins, scarlatinal, erysipelas and suppurative, by mixing several toxic preparations selected through skin reactions on human subjects and three sorts of antitoxin by immunization with these toxins. From the neutralization tests of polyvalent and monovalent toxins by these antitoxins on white pigs the following conclusions may be drawn: Scarlatinal, erysipelas and suppurative antitoxic sera equally neutralize scarlatinal, erysipelas and suppurative toxins.The neutralizing action of these sera is most marked in the case of homologous toxin.Scarlatinal antitoxic serum prepared by us is more comprehensive in its neutralizing action than the erysipelas serum.The results of the neutralization tests with antiscarlatinal serum of monovalent toxins of various strains have shown that the majority of toxic preparations (32 out of 34 strains), regardless of their sources, were neutralized with the exception of that produced by one erysipelas and one scarlatinal strain.

A part of the expenses of this investigation was defrayed by a grant from the Antiscarlatinal Committee of the Kwantung Government.
Studies on the Antigenic Properties of the Ultraviruses
Authors: Schultz E. W. , Bullock L. T. , Brewer H. V.
Published: December 2022

No evidence for the presence of specific complement fixing antibodies against the virus of rabies was found in the serum of rabbits which had been immunized in various ways with fixed virus brains. These immune sera also failed to present any evidence of specific precipitins. Rabicidal antibodies were demonstrated in all of the immune sera tested for this property.
Blood Grouping of Baffin Island Eskimos
Authors: Heinbecker Peter , Pauli Ruth H.
Published: December 2022

The blood groupings of 166 Eskimos of Baffin Island are recorded.

Further evidence is presented to support the view that the pure blooded Eskimo falls into blood group I.
The Primary Toxicity of Goat-Serum
Authors: Aronson Joseph D.
Published: December 2022

That the serum of certain normal animals may prove toxic when injected into normal animals of another species has long been known and a number of explanations have been offered of the mechanism involved.

Landois (1) believed that the toxicity of the serums of sheep, man, dog and cat is due to their ability to produce intravascular hemagglutination and intravascular coagulation. Leob, Strickler and Tuttle (2) concluded that the toxic action of beef-serum for rabbits is due to its hemagglutinating action, but that the toxic action of dog-serum for rabbits is due to its hemolytic action. Coca (3) believed that hemagglutination is responsible for the shock that follows the injection of washed heterologous cells.

That hemagglutination cannot be the cause of the toxic action of heterologous serum has been shown by Zinsser (4). This author found that while the hemolytic and toxic property of goat-serum for rabbits was lost when the serum was heated at 56°C. for twenty minutes, the hemagglutinative property was not affected.
General and Local Immunity to Cl. Botulinum Toxin
Authors: Nevin Mary , Hazen Elizabeth Lee
Published: December 2022

In spite of the fact that much work has been done on the subject of absorption of Cl. botulinum toxin from the gastro-intestinal tract and the relative susceptibility of various experimental animals to this toxin when administered per os, up to the present time very little work has been reported on the immunization of animals with Cl. botulinum toxin utilizing the gastro-intestinal tract as the route of administration. This is probably due to the very large amounts of the toxin necessary to produce any symptoms by this route together with the fact that although no symptoms of botulism may be evidenced a large percentage of the animals become extremely emaciated and subsequently die with cachetic symptoms.

Van Ermengem (1) made the first observation upon the differences in susceptibility of laboratory animals to obtulinum toxin and the corresponding differences when the toxin was administered by different routes.
A Study of the Reactions between Toxins and Antisera Produced with Hemolytic Streptococci
Authors: Kirkbride Mary B. , Wheeler Mary W. , Hendry Jessie L.
Published: December 2022

In previous studies, already reported (1, 2), in which the toxins of over two hundred strains of hemolytic streptococci from various infections including, among others, scarlet fever, erysipelas, tonsillitis, septic sore throat, and otitis media, were tested on goats, it was found that neutralization tests with a monovalent scarlet-fever antistreptococcus serum produced with the Dochez N. Y. No. 5 strain of hemolytic streptococcus failed to demonstrate any fundamental qualitative or quantitative differences between the toxins of the different strains. These results in general have been confirmed by other workers.

Recently in studying the reactions induced in goats with mixtures of a standard concentrated scarlet-fever goat antitoxin and toxins from different hemolytic streptococci marked variations were noted in the proportion of antitoxin required for neutralization of certain heterologous toxins which apparently induced reactions no stronger or in fact less strong than that induced by one skin-test dose of the standard scarlet-fever toxin.
Cis-urocanic acid stimulates human peripheral blood monocyte prostaglandin E2 production and suppresses indirectly tumor necrosis factor-alpha levels.
Authors: Hart P H , Jones C A , Jones K L , Watson C J , Santucci I , Spencer L K , Finlay-Jones J J
Published: December 2022

Photoisomerization of trans-urocanic acid (UCA) in the stratum corneum has been implicated in the immunosuppression detected after irradiation with UVB (UV wavelength of 280-320 nm). In this study, cis-urocanic acid suppressed human monocyte production of TNF-alpha by a PGE2-dependent mechanism. This contrasted with the mechanism involving histamine type 2 receptors by which the UCA structural analogue, histamine, suppressed monocyte TNF-alpha production. Histamine type 1 receptor antagonists were without effect on both the cis-UCA- and histamine-induced suppression of monocyte TNF-alpha levels. As indomethacin can reverse UVB-immunosuppression in murine models, we may have identified one of the cellular mechanisms responsible for reduced delayed-type hypersensitivity responses. Decreased TNF-alpha levels, by restricting further cytokine recruitment, may also limit the development of the inflammatory components of hypersensitivity responses.
Induction of experimental autoimmune encephalomyelitis in severe combined immunodeficient mice reconstituted with allogeneic or xenogeneic hematopoietic cells.
Authors: Jones R E , Bourdette D N , Whitham R H , Offner H , Vandenbark A A
Published: December 2022

Severe combined immunodeficient (SCID) C.B-17-scid/scid (H-2d) strain mice are deficient for T and B lymphocytes and lack all of the immune functions associated with these cell types. Experimental autoimmune encephalomyelitis (EAE) was induced in chimeric SCID mice that had been previously reconstituted with allogeneic mouse or xenogeneic rat hematopoietic stem cells from EAE-susceptible donor strains. Encephalitogenic, myelin Ag-specific, T lymphocytes selected from SJL mice, Lewis rats, or Buffalo rats transferred passive EAE into chimeric SCID mice reconstituted with SJL mouse, Lewis rat, or Buffalo rat hematopoietic cells, respectively. SCID mice reconstituted with Lewis rat hematopoietic tissue and thymus were also susceptible to EAE induced by active immunization with the myelin proteolipid protein synthetic peptide PLP S139-151. T lymphocytes recovered from the spleens of SCID mouse-rat chimeras with EAE proliferated upon in vitro stimulation with myelin Ag presented by APC syngeneic to the transplant donor, and rat T lymphocytes selected in vitro from SCID mouse-rat chimeras with EAE transferred EAE back into naive recipient rats. Thus, the immunodeficiency present in SCID mice can be overcome at least partially by hematopoietic tissue transplantation from allogeneic or xenogeneic donors. Furthermore, allogeneic SJL mouse and xenogeneic Lewis or Buffalo rat myelin Ag-specific T cells can transfer EAE between strains and species, respectively, into recipient SCID mouse chimeras.
Dipeptidyl peptidase I is enriched in granules of in vitro- and in vivo-activated cytotoxic T lymphocytes.
Authors: Brown G R , McGuire M J , Thiele D L
Published: December 2022

Recent studies have suggested that dipeptidyl peptidase I (DPPI) is the major post-translational processing enzyme responsible for generating activated myeloid and lymphoid granule serine proteases. The current studies assessed the relative levels of DPPI and granzyme A (BLT esterase) in B6 anti-H-2d-specific CTL generated in mixed lymphocyte cultures (in vitro-activated CTL), by infusion of B6 spleen cells into irradiated H-2d mice (graft-vs-host, GVH CTL) or by 1 degree and 2 degrees peritoneal immunization of B6 mice with P815 (H-2d) cells (PE CTL). In contrast to low levels of DPPI activity in unstimulated CD4+ spleen T cells, both unstimulated CD8+ spleen T cells and in vitro-activated CTL populations were several-fold enriched in DPPI activity, while PE CTL and GVH CTL expressed even higher levels of DPPI. Depletion of DPPI-enriched cells by treatment with Leu-Leu-OMe resulted in loss of cytolytic effector function from each CTL population. However, PE CTL and GVH CTL were more sensitive to the toxicity of Leu-Leu-OMe than were in vitro-activated CTL. While standard BLT esterase assays detected much higher levels of this serine protease activity in GVH CTL or in vitro-activated CTL than in PE CTL, levels of BLT esterase activity significantly above the basal levels present in unstimulated CD8+ or CD4+ T lymphocytes were found in association with immunoreactive granzyme A in lysates of PE CTL. In both PE CTL and in vitro-activated CTL, DPPI and BLT esterase activity co-localized in the granule fraction of cell lysates, and similar percentages of total cellular BLT esterase and DPPI were exocytosed upon cross-linking of surface CD3. Thus, both in vivo- and in vitro-activated CTL were found to possess functional granules containing readily detectable albeit somewhat different levels of DPPI and granzyme A activity.
Induction of early response genes by cross-linking membrane Ig on B lymphocytes.
Authors: Mittelstadt P R , DeFranco A L
Published: December 2022

Cross-linking of membrane Ig (mIg) on B lymphocytes induces protein tyrosine phosphorylation and phosphoinositide hydrolysis, events that are thought to mediate the diverse biologic responses of B cells to Ag binding. mIg stimulation also induces the expression of the putative transcriptional regulators c-myc, c-fos, egr-1, and jun-B. In this report, normal murine B cells and two murine B lymphoma cell lines were examined for the induction of mRNA expression of seven early response genes first identified in fibroblasts. Expression of four of the seven genes (nur77, nup475, pip92, and 3CH134), encoding two putative transcriptional regulators, a protein of unknown function, and a putative protein phosphatase, was induced after cross-linking of mIg in resting B cells isolated from mouse spleen. In the 2PK-3 and WEHI-231 B lymphoma cell lines three and two, respectively, of these four genes were induced. Expression of these genes could be induced in 2PK-3 cells by activating the phosphoinositide-signaling pathway independently of the tyrosine phosphorylation pathway by signaling through an M1 muscarinic acetylcholine receptor introduced by transfection. Additionally, in all but one case, these early response genes could be induced by directly activating protein kinase C with phorbol esters. In the cell line 2PK-3, the gene 3CH134 was not induced by phorbol ester treatment, but was induced by elevation of intracellular calcium. Thus, a subset of the early response genes identified in serum-stimulated fibroblasts is also induced by Ag-receptor stimulation in B lymphocytes, and this induction appears to be mediated by the phosphoinositide signaling pathway and, for the most part, protein kinase C.
Response of human NK cells to IL-6 alterations of the cell surface phenotype, adhesion to fibronectin and laminin, and tumor necrosis factor-alpha/beta secretion.
Authors: Rabinowich H , Sedlmayr P , Herberman R B , Whiteside T L
Published: December 2022

In vitro effects of human recombinant IL-6 (1-1000 U/ml) on highly enriched human NK CD3-CD56+ cells (94% +/- 2; mean +/- SEM; n = 8), obtained from PBL were studied. IL-6 induced low levels of NK cell proliferation (7- to 30-fold during 6-day incubation), which was IL-2-independent, because IL-6 did not induce detectable IL-2 production by NK cells. Two-color flow cytometry analysis demonstrated that incubation of NK cells with IL-6 at the optimal concentration of 250 U/ml for 6 days significantly increased the proportion of NK cells expressing the following activation Ag: CD25 (26% +/- 17, mean +/- SEM vs 4% +/- 1 in control, n = 5), CD54 (44% +/- 17 vs 9% +/- 3), HLA-DR (29% +/- 13 vs 12% +/- 4), CD69 (45% +/- 7 vs 12% +/- 3), and CD71 (34% +/- 17 vs 6% +/- 2). The mean fluorescence intensity of these activation Ag was increased as well. IL-6 induced expression of CD49b (alpha-chain of VLA-2, 20% +/- 11 vs 2% +/- 1) and CD49c (alpha-chain of VLA-3, 43% +/- 17 vs 5% +/- 3), which are not expressed on resting NK cells. IL-6 also enhanced the fluorescence intensity of beta 1 integrins, CD49d, CD49e, and CD49f, expressed on NK cells. IL-6-stimulated NK cells showed significantly increased integrin-mediated adhesion to fibronectin- or laminin-coated plates (26 +/- 3 mean % cells adhering +/- SEM vs 15 +/- 4 in control for FN and 19 +/- 1 vs 11 +/- 1 for LM, p < 0.05 for both) as determined in a 3 h binding assay. As assessed by inhibition of adhesion using mAb to the VLA-2, -3, -4, -5, and -6, NK cell adhesion to fibronectin was mediated by VLA-4 and 5, and their adhesion to laminin by VLA-3 and -6. NK cells incubated in the presence of IL-6 were found to produce a factor cytostatic to WEHI-164 clone 13 target cells. This effect was partly, although significantly, blocked by neutralizing antibodies to TNF-alpha or TNF-beta. Our data demonstrate that IL-6 can directly activate human NK cells, but is a less potent NK cell activator, for all activation and functional parameters studied, than IL-2.
Human CD4+ cytolytic T cells kill antigen-pulsed target T cells by induction of apoptosis.
Authors: Rahelu M , Williams G T , Kumararatne D S , Eaton G C , Gaston J S
Published: December 2022

Stimulation of human CD4+ T cell clones with appropriate specific peptides or with lectins in the absence of APC induces a substantial degree of cell death. We have investigated the mechanisms of induction of this cell death and show that it occurs by apoptosis, identified by morphology and the characteristic pattern of DNA degradation. We also investigated whether this T cell death was a result of a suicide process activated in the T cell after "inappropriate" recognition of Ag on the surface of another T cell clone (and in the absence of other accessory signals), or was due to a conventional lethal hit delivered by one cytolytic T cell to another "target" T cell. Our results strongly suggest that peptide-induced cell death of human CD4+ CTL is due to active killing of Ag-presenting target T cells by effector T cells of the same clone. The cell death that is induced in the target T cell occurs via apoptosis that requires de novo RNA transcription and translation in the effector T cell.
Functional thrombin receptors on human T lymphoblastoid cells.
Authors: Tordai A , Fenton J W , Andersen T , Gelfand E W
Published: December 2022

Human alpha-thrombin stimulated five different T lymphoblastoid cell lines to increase intracellular free Ca2+ concentrations, whereas five B cell lines did not similarly respond. The T cell intracellular free Ca2+ increased rapidly, plateaued within 10 to 20 s, and then declined without any measurable sustained intracellular free Ca2+ elevation. Liberation of inositol trisphosphate peaked within 60 s, and a rapid and sustained activation of protein kinase C was induced. The thrombin-specific inhibitor, hirudin, completely blocked the response to alpha-thrombin. Catalytically inactivated forms of alpha-thrombin failed to stimulate the T cells, whereas trypsin, gamma-thrombin (which lacks fibrinogen clotting activity), or the synthetic 14-amino acid thrombin receptor-activation peptide stimulated T cells, but required approximately 10-, approximately 15-, or approximately 100-fold higher concentrations, respectively, when compared to alpha-thrombin. Stimulation by thrombin receptor-activation peptide was desensitized by alpha-thrombin, and vice versa, but alpha-thrombin did not desensitize stimulation by mAb to the TCR/CD3 Ag. The presence of the receptor on T but not on B cells was confirmed by flow-cytometric analysis using a mAb against the human thrombin receptor. These findings demonstrate functional thrombin receptors on T lymphoblastoid cells that may be capable of activating the cells, independently of an ongoing immune response.
Elevated cyclic AMP inhibits endothelial cell synthesis and expression of TNF-induced endothelial leukocyte adhesion molecule-1, and vascular cell adhesion molecule-1, but not intercellular adhesion molecule-1.
Authors: Pober J S , Slowik M R , De Luca L G , Ritchie A J
Published: December 2022

We have investigated the role of cAMP as a signal transducer for TNF-induction of leukocyte adhesion molecule expression in cultured human umbilical vein endothelial cells (EC). Forskolin, a stimulator of adenylate cyclase, either alone or in combination with isobutyl methylxanthine (IBMX), an inhibitor of phosphodiesterase, fails to induce expression of endothelial leukocyte adhesion molecule 1 (ELAM-1 or E-selectin), of vascular cell adhesion molecule 1 (VCAM-1) or of intercellular adhesion molecule 1 (ICAM-1 or CD54). Unexpectedly, this combination of cAMP-elevating drugs inhibits TNF induction of ELAM-1 and VCAM-1 but not ICAM-1 expression. Similar results were observed with the membrane-permeant cAMP mimetics 8 bromoadenosine 3':5' cyclic monophosphate (8Br-cAMP) and N(6)2'-O-dibutyryladenosine 3':5'-cyclic monophosphate. Inhibition was greater at lower TNF concentrations (< 10 U/ml), at higher 8 Br-cAMP concentrations (> 100 microM), and at early times (2 h). Forskolin plus IBMX selectively inhibits TNF-induced increases in ELAM-1 and VCAM-1 mRNA, indicating that the action of cAMP is to block synthesis of these molecules. TNF, through stimulation of prostaglandin synthesis, produces slight elevations in the levels of endothelial cAMP. However, these increases in cAMP appear too small compared to those induced by forskolin plus IBMX to inhibit adhesion molecule expression. Indeed, complete inhibition of the TNF-mediated rise in cAMP, achieved by blocking cyclooxygenase with indomethacin, does not alter ELAM-1 expression. We conclude that cAMP is neither an intracellular mediator nor a physiological regulator of TNF-induced adhesion molecule expression in EC. However, our findings suggest that pharmacological elevations of cAMP in EC, by inhibiting TNF-induced synthesis of ELAM-1 and VCAM-1, could serve to limit inflammation.
Expression of the Fc receptor for IgG (Fc gamma RII/CDw32) by human circulating T and B lymphocytes.
Authors: Mantzioris B X , Berger M F , Sewell W , Zola H
Published: December 2022

Tissue-specific isoforms of the human FcR for IgG Fc gamma RII (CDw32) have previously been described by using mAb. These mAb were shown to exhibit different patterns of reactivity with lymphocytes. Among human PBL, Fc gamma RII has been detected on B cells but not T cells when assessed by flow cytometry and microscopy with the use of mAb KB61 and 41H16. Although KB61 and 41H16 were found to react with B cells, the mAb IV.3, CIKM5, and 2E1 did not react with any PBL subset. In this study, we show that KB61 and 41H16 react strongly with the majority (93-96%) of B cells (CD20+), and weakly with a proportion (18-42%) of T cells (CD3+), including 10 to 14% of CD4+ and 27 to 69% of CD8+ cells. In addition, mRNA for Fc gamma RII was detected in purified CD3+CD8high+ lymphocytes by polymerase chain reaction. KB61 and 41H16 also reacted with a majority of CD3-CD16/CD56+ cells, and CD3-CD20- cells. These findings indicate that a subset of T cells and non-T/non-B cells express Fc gamma RII, and are of interest in the light of previous studies which postulate that human Fc gamma R+ cells and Fc gamma RII+ murine T cells suppress the B cell immune response.
Identification of functional domains in murine granulocyte-macrophage colony-stimulating factor using monoclonal antibodies to synthetic peptides.
Authors: Greenfield R S , Braslawsky G R , Kadow K F , Spitalny G L , Chace D , Bull C O , Bursuker I
Published: December 2022

Granulocyte-macrophage (GM)-CSF is an important hematopoietic cytokine that regulates proliferation and differentiation of macrophages, neutrophils, and eosinophils. In this study, we generated mAb to five synthetic peptides that correspond to regions along the murine GM-CSF molecule. The ability of anti-peptide mAb to bind to and inhibit biologic activity of murine (m) GM-CSF was determined. mAb with the highest neutralization titers were derived from mice immunized with peptide II, which correspond to amino acids 27 to 38 of mGM-CSF. Immunochemical studies showed that peptide II specifically blocked binding of anti-peptide II mAb to GM-CSF. mAb to two other peptides in the N-terminal half corresponding to residues 7 to 17 and 47 to 58, respectively, of mGM-CSF also inhibited GM-CSF-dependent proliferation and differentiation of murine bone marrow precursors for macrophages and granulocytes. Anti-peptide mAb also inhibited growth of a murine hematopoietic cell line FDCP1 and a murine T cell line HT-2, which was shown to be dependent on GM-CSF for growth in vitro. Biologic activity of both natural and recombinant mGM-CSF was neutralized by anti-peptide mAb. These findings indicate that epitopes in the N-terminal region of mGM-CSF are important for biologic activity, and the epitope defined by peptide II (residues 27 to 38) lies within a particularly important functional domain of the mGM-CSF molecule.
Characterization of the EBV/C3d receptor on the human Jurkat T cell line: evidence for a novel transcript.
Authors: Sinha S K , Todd S C , Hedrick J A , Speiser C L , Lambris J D , Tsoukas C D
Published: December 2022

EBV is a human herpes virus that causes mononucleosis and is associated with various tumors. EBV infects cells via the CR2 that was previously thought to be expressed only on the surface of B cells and certain epithelial cells. Recent findings in our laboratory and those of others, however, have shown that the EBV receptor is also present on T cells. Our study shows that Jurkat human T cells have a molecule that reacts with both anti-CR2 antibodies and the third component of complement, C3. Furthermore, the data indicate that this molecule binds EBV detected by incubation with biotin-conjugated virus and streptavidin phycoerythrin. Viral binding is specific, as it is inhibited by nonconjugated virus, with anti-CR2 antibodies, and with an antibody reactive with the glycoprotein (gp350) that EBV uses to bind CR2. In addition, EBV variably infects Jurkat cells as demonstrated by the presence of transcripts of Epstein Barr nuclear Ag (EBNA-1) using the polymerase chain reaction. Immunoprecipitation experiments with anti-CR2 antibodies and SDS-PAGE analysis reveal a protein with an apparent molecular mass of 155 kDa which is higher than the one seen in B cells. The size of this molecule is reduced to 119 kDa upon endoglycosidase F treatment. Northern blot analysis of Jurkat poly(A)+ RNA shows a transcript of 4.7 kb upon probing with the B cell CR2 cDNA. This size is consistent with that of B cell CR2 mRNA. Two cDNA clones were identified upon screening of a Jurkat cell cDNA library with the B cell CR2 cDNA. One of the clones possesses an identical nucleotide sequence to the one corresponding to B cell CR2, whereas the other represents a differentially spliced transcript which lacks the exon 8b of B cell CR2. Analysis of Jurkat and Raji mRNA by PCR demonstrated the presence of this novel splice variant in both cell lines.
Beta 2-microglobulin-deficient mice can be protected against influenza A infection by vaccination with vaccinia-influenza recombinants expressing hemagglutinin and neuraminidase.
Authors: Epstein S L , Misplon J A , Lawson C M , Subbarao E K , Connors M , Murphy B R
Published: December 2022

Immunity to viral infections includes both antibody and T cell components. The contributions of humoral and cell-mediated immune responses vary depending on virus and host factors. We have used an in vivo challenge system to examine protective immunity to influenza A(H1N1) virus infection in immunocompetent B6 (H-2b) mice, and in H-2b mice homozygous for disruption of the gene for beta 2-microglobulin, termed beta 2 mu(-/-) mice. The latter mice do not express conventional MHC class I complexes on cell surfaces and lack CD8+ class I-restricted T cells. Ten vaccinia virus recombinants, each expressing 1 of the 10 proteins of influenza virus, were used to immunize the mice. Normal mice were protected against challenge with influenza virus by vaccination with HA-VAC and NA-VAC, but not by any of the vaccinia vectors expressing one of the eight other influenza virus proteins nor by a mixture of all eight of the latter vectors. Similar results were observed in mice of H-2d or H-2k MHC haplotypes. The beta 2 mu(-/-) mice were also protected by immunization with HA-VAC and NA-VAC, demonstrating that classical CD8+ CTL responses were not required for protection. Depletion of CD4+ T cells in either normal or beta 2 mu(-/-) mice at the time of challenge had little or no effect on protection induced by HA-VAC or NA-VAC, suggesting that preformed antibody is the dominant mediator of protective immunity induced by these recombinants. Antibody responses to vaccinia virus Ag and expressed influenza virus Ag were lower in titer in beta 2 mu(-/-) mice than in normal B6 mice, suggesting an influence of MHC class I complexes, CD8+ T cells, or their products on antibody production.
Neutrophil CD14: biochemical properties and role in the secretion of tumor necrosis factor-alpha in response to lipopolysaccharide.
Authors: Haziot A , Tsuberi B Z , Goyert S M
Published: December 2022

CD14 is a myeloid cell differentiation Ag expressed primarily by monocytes and macrophages. CD14 has recently been shown to function as a receptor for a complex of LPS and LPS binding protein (LBP), an acute phase serum protein also present in normal serum in trace amounts. In the presence of LBP, LPS strongly activates monocytes via CD14 as measured by TNF secretion. This pathway of monocyte activation is thought to be a major contributor to the symptoms of endotoxin shock. Another major cell type involved in the response to Gram-negative infection is the neutrophil. Recent studies have shown that neutrophils also express CD14 and suggest that they can respond to LPS through a similar pathway. However, the biochemical nature of neutrophil CD14 has not previously been described. In this report, we have analyzed several biochemical characteristics of neutrophil CD14. We show that CD14 is actively synthesized by neutrophils as a glycosylphosphatidyl-inositol-anchored protein, indistinguishable in size from monocyte CD14. Furthermore, neutrophils, like monocytes, shed a smaller soluble form of CD14 into culture supernatants. In addition, like monocytes, neutrophils respond to LPS/LBP complexes via CD14 by releasing TNF-alpha. The described properties and function of neutrophil CD14 suggest that it may directly participate in the acute inflammatory response and in endotoxin shock.
IL-1 and transforming growth factor-beta inhibit platelet-derived growth factor-AA binding to osteoblastic cells by reducing platelet-derived growth factor-alpha receptor expression.
Authors: Yeh Y L , Kang Y M , Chaibi M S , Xie J F , Graves D T
Published: December 2022

Platelet-derived growth factor (PDGF) is thought to play a significant role in bone repair and regeneration. We previously demonstrated that PDGF-AA-induced chemotaxis and proliferation can be modulated by IL-1. We now report that IL-1 and transforming growth factor-beta (TGF-beta) significantly decrease the number of PDGF-AA binding sites in both normal and tumor-derived human osteoblastic cells, whereas PDGF-BB binding is minimally affected. The affinity of PDGF-AA binding remains unchanged in the presence of IL-1, but is slightly reduced by TGF-beta as demonstrated by Scatchard analysis. We also showed that tyrosyl kinase phosphorylation after PDGF-AA binding is decreased in the presence of both IL-1 and TGF-beta. Northern blot analysis indicates that both IL-1 and TGF-beta decrease the expression of PDGF-alpha receptor mRNA. These results suggest that IL-1 and TGF-beta have the potential to regulate PDGF-AA-induced biologic activity in normal human osteoblastic cells and in human osteoblastic sarcoma cells by decreasing the levels of the PDGF-alpha receptor.
A peptide variant of an arthritis-related T cell epitope induces T cells that recognize this epitope as a synthetic peptide but not in its naturally processed form.
Authors: Wauben M H , van der Zee R , Joosten I , Boog C J , van Dijk A M , Holewijn M C , Meloen R H , van Eden W
Published: December 2022

The immune system has the potential to utilize a diverse T cell repertoire for the recognition of Ag in the context of MHC molecules. Here we describe the analysis of two rat T cell clones, both of which recognize a synthetic peptide comprised of the arthritis-associated 180-188 amino acid sequence of the mycobacterial 65-kDa heatshock protein (hsp65 180-188), but which differ in the recognition of the naturally processed hsp65. The arthritogenic T cell clone A2b, generated by immunization with whole Mycobacterium tuberculosis, recognized the hsp65 180-188 synthetic peptide as well as the processed hsp65, whereas T cell clone ATL11, generated after immunization with a single amino acid substituted peptide analog of hsp65 180-188, recognized peptide hsp65 180-188 but not the processed hsp65. For both T cell clones the minimal stimulatory sequence was hsp65 180-186. However, within this minimal stimulatory sequence marked differences between the clones were found with regard to peptide residues interacting with the TCR. Furthermore, addition of extra residues at the N terminus of the hsp65 180-186 sequence abrogated the recognition by clone ATL11, but not by A2b. These findings demonstrate that, upon in vivo immunization with a synthetic peptide comprised of a single amino acid variant of a T cell epitope, T cells can be triggered that recognize a peptide comprised of the original epitope sequence, but that do not recognize this epitope in its naturally processed protein fragment. The possibility of triggering such T cells by immunization with synthetic peptides, may well have consequences for the design of peptide vaccines or peptide immunomodulatory agents.