bluuebunny/crossref_metadata_2025 huggingface dataset:
Cloning, sequence, and expression of bovine interferon-gamma.
Authors: Cerretti D P , McKereghan K , Larsen A , Cosman D , Gillis S , Baker P E
Published: December 2022

Bovine interferon-gamma (IFN-gamma) sequences have been isolated by screening a cDNA library with a human IFN-gamma cDNA probe. The cDNA library was constructed from RNA isolated from concanavalin A-stimulated bovine lymph node cells. The open reading frame predicts that the bovine IFN-gamma precursor is composed of 166 amino acids with a predicted m.w. of 19,393. Alignment of the amino acid sequence with human IFN-gamma indicates that mature bovine IFN-gamma is composed of 143 amino acids with a predicted m.w. of 16,858. It has an amino acid homology of 63% with human IFN-gamma, and 47% with murine IFN-gamma. Biologically active bovine IFN-gamma was synthesized in an Escherichia coli expression system.
Murine B cell hybridomas bearing ligand-inducible Fc receptors for IgE.
Authors: Lee W T , Conrad D H
Published: December 2022

Interest in the regulation of IgE synthesis has generated investigation of low-affinity Fc receptors for IgE (Fc epsilon R) and the related immunoregulatory IgE-binding factors. In an effort to facilitate biochemical analysis of the B lymphocyte Fc epsilon R, hybridoma technology has been used to create stable cell lines that maintain Fc epsilon R in high numbers. Fusion of the HAT-sensitive B lymphoma, M12.4.5, with murine B cells from Nippostrongylus brasiliensis infected BALB/c mice led to the formation of hybrid cells of B cell phenotype, all of which were Fc epsilon R+, including several that had greater than 50,000 Fc epsilon R/cell. The Fc epsilon R on these cells were biochemically identical to the Fc epsilon R on normal B cells with respect to binding affinity (approximately equal to 10(8) M-1), m.w. (49,000), and tryptic peptides. Each hybridoma cell line specifically increased its Fc epsilon R level between twofold and fourfold when cultured with rat or mouse IgE. Additional studies demonstrated that the increased IgE binding ability was due to an increase in receptor number rather than an affinity change, and the Fc epsilon R increase was seen on the entire cell population. Dose studies indicated that oligomeric IgE was 10-fold more effective than monomeric IgE in causing upregulation, and the effective concentrations required indicated that induction occurred only if IgE was present in saturating concentrations. Upon addition of IgE, peak Fc epsilon R levels were reached after 15 to 20 hr of culture; blocking protein synthesis with cycloheximide largely blocked the increase in Fc epsilon R levels. Additionally, the inductive signal IgE must constantly be present to maintain upregulated Fc epsilon R levels in that its removal from the culture resulted in a rapid decline of Fc epsilon R from induced to normal levels. Because Fc receptor upregulation is important to several systems describing Ig isotype-specific regulation, the ability to examine such receptor upregulation at a clonal level should aid in discerning the role of the Fc epsilon R in the regulation of IgE antibody synthesis.
Structural analysis of the oligosaccharides of DR1 and DQw1 molecules.
Authors: Iturbe S , Narasimhan S , Letarte M
Published: December 2022

The major glycopeptide fractions of the alpha- and beta-chains of HLA-DR1 and DQw1 molecules were isolated on columns of immobilized concanavalin A (Con A), Lens culinaris (Lens), Ricinus communis agglutinin Type I (RCA), and leuko-phytohemagglutinin. Oligosaccharides were prepared from these fractions by enzymatic digestion with Endoglycosidases H or F and were analyzed on Bio-Gel P-6. The glycopeptides tightly bound to Con A (ConA III) were mostly associated with alpha-chains and were resolved as a single oligosaccharide peak (Kd = 0.72) on Bio-Gel P-6 after Endo H digestion. Man-5 is the minimal polymannosyl structure which can be deduced for the ConA III fractions of either DQw1 or DR1 oligosaccharides. The major component of the glycopeptides of the alpha-chains of either DR1 or DQw1 molecules which were weakly bound to Con A (ConA II fraction) did not interact with RCA before or after mild acid hydrolysis or neuraminidase treatment. This component represents a biantennary complex with neither terminal galactose nor sialic acid residues with a minimal structure terminating in N-acetyl glucosamine on the Mannose alpha 1----6 arm, referred to as GnM. The ConA II fractions, which constitute 10% of the total glycopeptides of beta-chains, are associated primarily with fucosylated, sialylated biantennary oligosaccharides not seen on the alpha-chains. The ConA I unbound fractions of either alpha- or beta-chains were mostly bound to RCA after mild acid hydrolysis, suggesting that the minimal structure was a sialylated triantennary structure. The major component associated with the beta-chains was bound to Lens such that a more definite structural assignment can be made, i.e., a triantennary structure with the Mannose on the alpha 1----6 arm substituted at C-2 and C-6. The oligosaccharides of alpha- and beta-chains were resolved as broad peaks on Bio-Gel P-6, suggesting that a mixture of tri- and tetraantennary structures with variable degrees of sialylation and galactosylation were present. The structural differences reported here between oligosaccharides of alpha- and beta-chains of DQw1 and of the two subsets of DR1 molecules could be responsible in part for the differential recognition properties expected of human class II molecules encoded by distinct loci.
Differential stimulation of the respiratory burst and lysosomal enzyme secretion in human polymorphonuclear leukocytes by synthetic diacylglycerols.
Authors: Cox C C , Dougherty R W , Ganong B R , Bell R M , Niedel J E , Snyderman R
Published: December 2022

Binding of chemoattractants to receptors on human polymorphonuclear leukocytes (PMN) stimulates the phosphodiesteric cleavage of phosphatidylinositol 4,5-bisphosphate to produce inositol 1,4,5-trisphosphate and 1,2-diacylglycerols. To investigate the possible second messenger function of diacylglycerols in PMN activation, we tested the ability of a series of synthetic sn 1,2-diacylglycerols, known to stimulate protein kinase C in other systems, to promote superoxide anion release, oxygen consumption, lysosomal enzyme secretion, and chemotaxis. None of the diacylglycerols initiated the chemotactic migration of PMN. Several of the diacylglycerols however, were, active in stimulating superoxide anion release and lysozyme secretion, with dioctanoylglycerol (diC8) being the most potent. Unexpectedly, didecanoylglycerol (diC10) induced lysosomal enzyme secretion, but failed to stimulate superoxide production or oxygen consumption. All other biologically active diacylglycerols tested displayed similar EC50 for stimulating lysozyme secretion and superoxide production. The ability of the diacylglycerols to compete for phorbol dibutyrate (PDBu) binding in intact PMN suggested a mechanism for the divergent biological activity of diC10. Although the compounds that stimulated both superoxide production and lysosomal enzyme secretion competed for essentially all [3H]PDBu binding from its receptor, diC10, which only stimulated secretion, competed for 45% of the bound [3H]PDBu. Thus diacylglycerols can selectively activate certain functions of leukocyte chemoattractant receptor. The data suggest that a discrete pool of protein kinase C may mediate activation of the respiratory burst in PMN.
Development of receptors for leukotriene B4 on HL-60 cells induced to differentiate by 1 alpha,25-dihydroxyvitamin D3.
Authors: Goldman D W , Olson D M , Payan D G , Gifford L A , Goetzl E J
Published: December 2022

The incubation of HL-60 human promyelocytic leukemia cells for 7 days with 100 nM 1 alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3] induced differentiation into monocyte-like cells, as assessed by morphologic and biochemical characteristics. Stereospecific receptors for leukotriene B4 (LTB4) developed on the surface of the HL-60 cell-derived monocytes that had the capacity to transduce LTB4 stimulation of a transient increase in the cytosolic concentration of calcium ([Ca+2]in). HL-60 cell-derived monocytes, but not undifferentiated HL-60 cells, expressed a high affinity subset of 6400 +/- 3700 receptors per cell with a dissociation constant (Kd) of 2.3 +/- 1 nM (mean +/- SD, n = 3) and a low affinity subset of approximately 2.2 X 10(6) receptors per cell with an apparent Kd of 680 +/- 410 nM. Derivatives of LTB4 inhibited the binding of [3H]LTB4 to HL-60 cell-derived monocytes with a rank order of potency of LTB4 greater than 20-OH-LTB4 greater than 3-aminopropyl amide-LTB4, which is similar to the order for LTB4 receptors of human blood PMNL. In contrast, leukotrienes C4 and D4 and formyl-methionyl chemotactic peptides did not inhibit the binding of [3H] LTB4, which demonstrates the specificity of these receptors for isomers of 5,12-dihydroxy-eicosatetraenoic acid. LTB4 stimulated an increase in [Ca+2]in in HL-60 cell-derived monocytes which reached 50% of the maximal level at an LTB4 concentration of 0.5 nM (EC50). Preincubation of HL-60 cell-derived monocytes with 10 nM LTB4 resulted in a selective loss of high affinity receptors, as assessed by binding of [3H]LTB4, and a 200-fold increase in the EC50 for stimulation by LTB4 of increases in [Ca+2]in, without alterations in either the low affinity receptors for LTB4 or the responsiveness of [Ca+2]in to formyl-methionyl chemotactic peptides. HL-60 cells that are induced to differentiate into monocytes thus develop stereospecific receptors for LTB4 with binding and transductional characteristics similar to those of human blood PMNL.
Platelet-activating factor (PAF) mediation of rat anaphylactic responses to soluble immune complexes. Studies with PAF receptor antagonist L-652,731.
Authors: Doebber T W , Wu M S , Biftu T
Published: December 2022

A new synthetic compound, L-652,731 (trans-2,5-(3,4,5-trimethoxyphenyl) tetrahydrofuran), which has been demonstrated by Hwang et al. to be a potent and specific platelet-activating factor (PAF) receptor antagonist causes 100% inhibition of 1 microM PAF-induced neutrophil degranulation at 50 microM, but has no effect on neutrophil degranulation induced by precipitating immune complexes (323 micrograms/ml), fMet-Leu-Phe (10(-7) M), or the calcium ionophore A23187 (10(-5) M). Intravenous infusion of 1 mumol L-652,731 results in almost 100% inhibition of hypotension induced by PAF but not that induced by isoproterenol, histamine, bradykinin, or acetylcholine. With the use of this novel PAF receptor antagonist, the in vivo mediator role of PAF in the soluble immune complex-induced hypotension, extravasation, vascular lysosomal hydrolase secretion, and neutropenia in rats was determined. The hypotension, extravasation, and lysosomal hydrolase release induced by immune complex infusion take 2 to 10 min longer to occur than the same responses elicited by PAF infusion. The neutropenia response is immediate with both stimuli. L-652,731 when orally administered to rats (20 mg/kg, 1.5 hr before PAF infusion) inhibited PAF-induced hypotension (69%), extravasation (76%), vascular lysosomal hydrolase release (79%), and neutropenia (73%). The same L-652,731-dosing regimen inhibited immune complex-stimulated hypotension (87%), extravasation (77%), and vascular lysosomal hydrolase release (31%). The initial and complete neutropenia induced by immune complex infusion was not inhibited in L-652,731-pretreated rats, but the rate of return of neutrophils to the blood was faster in the latter rats. Rats with blocked circulation to the liver still exhibited extensive extravasation and vascular lysosomal hydrolase release in response to PAF, but there was no extravasation and greatly reduced hydrolase release in response to immune complexes. Thus PAF is indicated to be a major mediator of soluble immune complex-induced hypotension and vascular permeability and a minor mediator of immune complex-induced lysosomal hydrolase release in rats. PAF probably does not mediate the initial and complete neutropenia stimulated by immune complexes. The liver is probably the major site for PAF production in response to circulating immune complexes.
Clonal relationship of the lymphoblastic cell line P388 to the macrophage cell line P388D1 as evidenced by immunoglobulin gene rearrangements and expression of cell surface antigens.
Authors: Bauer S R , Holmes K L , Morse H C , Potter M
Published: December 2022

A lymphocytic tumor of early B cell lineage, P388, and a mature macrophage tumor, P388D1, appear to have been derived from a common precursor based on identical immunoglobulin gene rearrangements and shared cell surface antigens. These cell lines may have resulted from differential maturation of a transformed cell that is an immediate cellular precursor to both the B cell and myeloid lineages. This supports earlier findings that suggest a close relationship between early B cell and myeloid differentiation pathways.
Recycling of receptor-bound IgE by rat basophilic leukemia cells.
Authors: Furuichi K , Rivera J , Buonocore L M , Isersky C
Published: December 2022

Rat basophilic leukemia (RBL) cells have receptors for immunoglobulin E (IgE). We previously showed that unlike some other ligands, the binding of monomeric rat or mouse IgE to RBL cells does not induce endocytosis. However, aggregation of the cell-bound, monomeric mouse IgE anti-dinitrophenyl (DNP) with DNP-protein conjugates leads to endocytosis of the aggregated mouse IgE and to the co-endocytosis of some unaggregated monomeric rat IgE. In this study we analyzed and compared the fate of co-endocytosed and endocytosed IgE. We found that co-endocytosed rat IgE recycled back to the cell surface within 3 to 4 hr. In contrast, endocytosed, immunochemically cross-linked, receptor-bound mouse IgE anti-DNP was partially degraded and was released into the medium, with no observable recycling of receptors, by 3 hr. However, addition of the hapten, DNP-lysine, resulted in rapid recycling (t1/2 10 min) of the endocytosed receptor-bound IgE to the plasma membrane and blocked additional endocytosis. Recycling of the endocytosed mouse IgE was more pronounced when the hapten was added to cells within 20 min of the initiation of endocytosis. When the hapten was added to the cells at later times (60 to 180 min), progressively less IgE recycled to the surface. This may reflect shuttling of the internalized IgE from a 'prelysosomal' to a 'lysosomal' compartment. Thus we provide evidence for recycling of monomeric IgE receptor complexes, sorting between cross-linked and non-cross-linked IgE receptor complexes, the freeing of receptor-bound monomeric IgE from the endocytosed immune-complexed IgE, and the apparent dependence of the recycling efficiency upon intracellular localization.
1,25-Dihydroxyvitamin D3 activates secretion of hydrogen peroxide by human monocytes.
Authors: Cohen M S , Mesler D E , Snipes R G , Gray T K
Published: December 2022

Human blood monocytes cultured in the presence of 1,25(OH)2D3 developed enhanced competence for secretion of H2O2 relative to cells suspended in media. This effect was maximal at a concentration of 10(-8) M 1,25(OH)2D3. After 3 days of incubation, monocyte-derived macrophages (MDM) exposed to 1,25(OH)2D3 demonstrated competence for secretion of H2O2 equivalent to cells exposed to recombinant IFN-gamma. Both IFN-gamma and 1,25(OH)2D3 offset decay of this function among cells in culture after 7 days. Simultaneous exposure of cells to 1,25(OH)2D3 and IFN-gamma did not activate competence for H2O2 secretion more than either agent alone. 24,25(OH)2D3 and 25(OH)2D3 activated MDM but at higher concentration than required for 1,25(OH)2D3. Progesterone did not affect H2O2 production. Incubation of MDM with a monoclonal antibody directed against IFN-gamma inhibited activation induced by lymphokine, and to a lesser extent by cells activated with IFN-gamma; this antibody had an insignificant effect on cells treated with 1,25(OH)2D3. These results suggest that 1,25(OH)2D3 exerts a receptor-mediated effect on monocyte function that results in cellular activation as manifested by enhanced competence for secretion of H2O2. It is possible that smaller concentrations of 1,25(OH)2D3 present in serum are permissive for macrophage activation, or that monocytic phagocytes are exposed to high concentrations of vitamin D metabolites under some clinical circumstances.
A chymotrypsin-type serine protease in rat basophilic leukemia cells: evidence for its immunologic identity with atypical mast cell protease.
Authors: Kido H , Izumi K , Otsuka H , Fukusen N , Kato Y , Katunuma N
Published: December 2022

Activity of a chymotrypsin-type serine protease was found in a subline of rat basophilic leukemia (RBL-2H3) cells. The protease was immunologically cross-reactive with anti-atypical mast cell protease immunoglobulin (Ig) G, and its activity was inhibited in a dose-dependent manner by the antibody. The apparent m.w. of the protease that reacted with the antibody was 25,000, which was identical with that of atypical mast cell protease in rat mucosal mast cells. These results show that the chymotrypsin type serine protease in RBL-2H3 cells is immunologically identical with atypical mast cell protease, which was first purified from rat small intestine. Immunohistochemical studies showed that the protease was located not only in intracytoplasmic granules but also in organelles synthesizing protein, such as cisternae of the rough endoplasmic reticulum, perinuclear spaces, and the Golgi apparatus. However, no immunoreactivity was demonstrated in rat basophils. The activity of the protease increased in the exponential phase of growth of RBL-2H3 cells in which some activity was also detected in the medium, and it decreased in the late stationary phase.
In vivo treatment with anti-I-A antibodies: differential effects on Ia antigens and antigen-presenting cell function of spleen cells and epidermal Langerhans cells.
Authors: Aberer W , Kruisbeek A M , Shimada S , Katz S I
Published: December 2022

The in vivo activation of T cells by a variety of antigens can be inhibited by the administration of anti-I-A antibodies (Ab) at the time of antigen priming. This inhibition can partially be explained by the temporary loss of Ia molecules from Ia-bearing antigen-presenting cells (APC) in the spleen. In this study, the effects of i.p. injected monoclonal Ab specific for I-A glycoproteins of different H-2 haplotypes on Ia antigen expression and APC function of spleen cells and epidermal Langerhans cells were compared. It was found that anti-I-A Ab quickly bound to both spleen cell and Langerhans cell Ia antigens. Although spleen cell Ia antigens were modulated and thus temporarily disappeared, Ia antigen expression by epidermal Langerhans cells was not modulated. In functional studies, the capacity of spleen cells and epidermal cells from anti-I-A Ab treated vs control animals to function as APC for antigen-specific, I-A- or I-E-restricted T cell clones was tested. A single injection of anti-I-A Ab completely abolished the APC function of spleen cells as shown in several inbred mouse strains, F1 animals, and with the use of several different Ab and T cell clones. In contrast, Langerhans cell-dependent APC function of epidermal cells remained completely unaltered. Even multiple injections of high doses of Ab never caused any inhibition of Langerhans cell function. Experiments with anti-I-Ak or anti-I-Ad Ab in an (H-2k X H-2d)F1 animal showed abrogation of APC function of spleen cells, but again not of Langerhans cells. Thus in vivo anti-I-A Ab administration appears to differentially affect Ia antigen expression and APC function from spleen and epidermis: Ia antigens are modulated from spleen cells but not from epidermis, and APC function disappears in the spleen but not in the epidermis. The abrogation of splenic but not of Langerhans cell APC function with anti-I-A Ab will facilitate the dissection of the relative contributions of Langerhans cells as compared with other APC in the generation of cutaneous immune responses.
In situ and in vitro characterization of the cellular immune response in erythema nodosum leprosum.
Authors: Modlin R L , Mehra V , Jordan R , Bloom B R , Rea T H
Published: December 2022

We sought to evaluate cell-mediated immune responses in erythema nodosum leprosum (ENL), a reactional state occurring in lepromatous leprosy. Skin biopsies from patients with leprosy were studied with monoclonal antibodies against T lymphocyte antigenic determinants, interleukin 2 (IL 2), and IL 2 receptors (Tac) by using immunoperoxidase staining of frozen sections. Peripheral blood lymphocytes from 18 ENL patients were tested in vitro for lepromin-induced suppression of Con A stimulation. Serial studies of seven lepromatous patients who developed ENL during the course of the study showed increases in both the Leu-3a:Leu-2a ratio and the number of IL 2-positive cells. IL 2-positive cells comprised 0.3% of the cells in all of the ENL lesions studied as compared with the 0.03% found in nonreactional lepromatous lesions (P less than 0.001). Lepromin-induced suppression of the Con A response, present in nonreactional lepromatous patients, significantly decreased in patients developing the ENL reaction, but returned after recovery from ENL. These changes in tissues and peripheral blood suggest that the pathogenesis of ENL is related to cell-mediated immune processes. Despite these immunologic changes, however, ENL patients do not recover antigen-specific skin tests or eliminate Mycobacterium leprae.
Prolongation of murine skin allograft survival by immunologic manipulation with anti-interleukin 2 receptor antibody.
Authors: Granstein R D , Goulston C , Gaulton G N
Published: December 2022

Administration of a rat monoclonal antibody (M7/20) directed against the murine interleukin 2 (IL 2) receptor in combination with sublethal x-irradiation of the recipient significantly enhanced the survival of skin allografts, both when the grafts were MHC disparate from the hosts and when only minor histocompatibility differences were present compared with untreated controls. No prolongation in graft survival was seen with either treatment alone at the dose employed. M7/20 and x-ray-treated allograft recipients also displayed significantly decreased alloantigen-specific reactivity against donor-strain spleen cells in both delayed-type hypersensitivity and cytotoxicity assays. Thus, such combination treatment reduces expression of host immune reactivity against graft determinants by several criteria. This work provides additional evidence that monoclonal antibodies directed against the IL 2 receptor may be useful in clinical transplantation.
Characterization of Theiler's murine encephalomyelitis virus (TMEV)-specific delayed-type hypersensitivity responses in TMEV-induced demyelinating disease: correlation with clinical signs.
Authors: Clatch R J , Lipton H L , Miller S D
Published: December 2022

After intracerebral inoculation of Theiler's murine encephalomyelitis virus (TMEV), certain mouse strains develop a persistent central nervous system (CNS) infection and inflammatory demyelinating lesions containing infiltrates of mononuclear cells and macrophages. Previous findings demonstrating a strong correlation between disease incidence, the presence of particular H-2 region genotypes, and development of high levels of TMEV-specific delayed-type hypersensitivity (DTH) supported an immune-mediated basis for myelin breakdown. These findings led us to examine whether a possible causal relationship would be supported by a temporal analysis comparing the onset of clinical disease and the development of TMEV-specific cellular or humoral immune responses in susceptible and resistant strains. In susceptible SJL/J mice, TMEV-specific DTH and T cell proliferative (Tprlf) responses developed within 10 to 14 days postinfection, preceded the onset of clinical signs, and remained elevated for 6 mo. In contrast, resistant BALB/c mice developed low levels of TMEV-specific Tprlf and no measurable DTH. However, both strains attained comparable levels of TMEV-specific serum antibody responses with parallel kinetics. Both DTH and Tprlf responses in susceptible SJL/J mice were shown to be specific for TMEV and mediated by L3T4+, Lyt-1+2-, class II-restricted T cells. A model is proposed implicating an effector role for TMEV-specific DTH, wherein lymphokine release by virus-specific DTH T cells leads to the recruitment, accumulation, and activation of macrophages in CNS tissue, which cause bystander myelin destruction and provide a permissive population of host cells for TMEV persistence.
The suppressive effect of gangliosides upon IL 2-dependent proliferation as a function of inhibition of IL 2-receptor association.
Authors: Robb R J
Published: December 2022

Gangliosides inhibited the proliferation of mitogen-activated human peripheral blood lymphocytes and the IL 2-dependent growth of murine T cell lines and 5-day-old human PHA lymphoblasts. In the case of the murine cell lines and PHA lymphoblasts, most of the effect of gangliosides could be reversed by the addition of high levels of IL 2. In the case of freshly-stimulated mitogen blasts, however, the ganglioside-induced inhibition could not be reversed by increasing exogenous IL 2 levels. These results indicate that inhibition of proliferation by gangliosides can be divided into IL 2-reversible and IL 2-irreversible mechanisms, the latter of which were predominant during the initial stage of cellular activation. Inclusion of gangliosides in receptor binding assays for radiolabeled IL 2 indicated that the IL 2-reversible mechanism likely involved competition between gangliosides and the cellular receptor for the binding of IL 2. Gangliosides blocked binding of radiolabeled IL 2 to both the high and low affinity forms of the IL 2 receptor, and this effect was most noticeable when the gangliosides and IL 2 were preincubated before addition of the target cells. In contrast, treatment of cells with gangliosides had no effect on the affinity of the cellular IL 2 receptor if the free gangliosides were removed immediately before the binding assay. Gangliosides also blocked the binding of radiolabeled IL 2 to anti-IL 2 antibodies, supporting the notion that their inhibitory effect is mediated via a direct interaction with IL 2. Thus, one major mechanism by which gangliosides block the IL 2-dependent proliferation of activated cells is by the sequestering or inactivation of the IL 2 molecule. This effect is reversible with the addition of excess IL 2, which distinguishes it from other mechanisms of ganglioside-dependent inhibition operating during the cellular activation process.
Immunochemical analysis of the Ia polymorphisms among the family of DR7-associated HLA-D specificities.
Authors: Karr R W
Published: December 2022

Among cells that bear the serologically defined Ia alloantigen DR7, four T cell-defined HLA-D specificities have been described: Dw7, Dw17, Dw11, and Dw7L. Ia molecules expressed by cells homozygous for these specificities have been compared by using immunofluorescence and two-dimensional gel electrophoresis in order to identify the DR and DQ polymorphisms among the family of DR7-associated HLA-D specificities. Cells homozygous for each of the four HLA-D specificities have in common one DR molecule that is indistinguishable by these methods. Two DR-specific monoclonal antibodies, IIIE3 and 109d6, detect a second distinct DR molecule on Dw7, Dw17, and Dw7L cells. This second DR molecule is also very similar from cells of the three specificities. In contrast, a second DR molecule was not detected on four Dw11 homozygous cells. Therefore, these data raise the possibility that all DR homozygous cells do not express the same number of DR molecules. The DQ molecules expressed by DQw2-positive Dw7, Dw17, and Dw7L cells are also very similar, whereas DQw3-positive Dw11 DQ molecules are structurally different. Therefore, no DR or DQ structural polymorphisms were detected to correlate with the Dw7, Dw17, and Dw7L T cell-defined Ia polymorphisms.
Generation of cytotoxic T lymphocytes from thymocyte precursors to trinitrophenyl-modified self antigens. I. Requirement of both killer-helper factor(s) and interleukin 2 for CTL generation from a subpopulation of thymocytes.
Authors: Takatsu K , Kikuchi Y , Kanatani T , Okuno K , Hamaoka T , Tominaga A , Sano Y
Published: December 2022

The requirement for signals in the induction of cytotoxic T lymphocytes (CTL) from thymocyte precursors has been investigated. Either unfractionated or peanut agglutinin-binding (PNA+) C3H/He thymocytes were stimulated with mitomycin C(MMC)-treated, 2,4,6-trinitrophenyl(TNP)-modified syngeneic spleen cells in the presence of a variety of lymphokine preparations. Cellfree supernatant (CFS) from purified protein derivatives(PPD-CFS) stimulated Mycobacterium tuberculosis (Tbc)-primed cells, or partially purified interleukin 2 (IL 2) mediated strong cytotoxic responses in unfractionated thymocytes, whereas only PPD-CFS at final concentrations beyond 30% was active for CTL generation in PNA+ thymocytes. Neither IL 2 at concentrations of below 60 U/ml nor a low concentration of PPD-CFS (at final below 10%) had such a capacity. The addition of monoclonal anti-IL 2 receptor antibody completely blocked CTL generation induced by PPD-CFS in PNA+ thymocytes. In contrast, anti-immune interferon (IFN-gamma) antibody showed a marginal effect. PPD-CFS (10%) and IL 2 (10 U/ml) could synergistically trigger PNA+ thymocytes to induce CTL generation. These results suggested that both IL 2 and "helper" factors other than IL 2 are required for CTL generation from PNA+ thymocytes. We refer to these kinds of helper factors as killer helper factors (KHF). Partially purified IL 2-free KHF show two peaks of activities at apparent m.w. 14,000 to 34,000 and 44,000 to 90,000, and are heterogeneous with respect to isoelectric point, which is between 4.5 and 5.1. Cultures that received TNP-modified syngeneic cells and KHF on day 0 and IL 2 on day 2 generated potent CTL responses, whereas the addition of IL 2 on day 0 followed by the addition of KHF on day 2 to the culture was ineffective, suggesting that KHF is required in the early phase of the culture to achieve optimal CTL responses.
Regulation of immune responses by T cell subsets. Role of helper and suppressor T cells in the development and expression of MHC-restricted antibody responses to L-glutamic acid60-L-alanine30-L-tyrosine10 (GAT) by (responder X responder)F1 spleen cells.
Authors: Lake J P , Kapp J A , Pierce C W
Published: December 2022

The roles of helper and suppressor T cells in the development and expression of antibody responses to GAT were studied in (responder X responder)F1 mice immunized with parental GAT-M phi. Spleen cells from (B10 X B10.D2)F1 mice primed in vivo with B10 or B10.D2 GAT-M phi developed secondary in vitro plaque-forming cell (PFC) responses only when stimulated by GAT-M phi syngeneic with the GAT-M phi used for in vivo priming. By contrast, virgin F1 spleen cells developed comparable primary PFC responses to both parental GAT-M phi Co-culture of T cells from (B10 X B10.D2)F1 mice primed in vivo by B10 GAT-M phi with virgin (B10 X B10.D2)F1 spleen cells demonstrated the presence of suppressor cells that inhibited the primary response of virgin spleen cells stimulated by B10.D2 GAT-M phi. Spleen cells from (B10 X B10.D2)F1 mice primed in vivo with B10.D2 GAT-M phi had suppressor T cells that suppressed primary responses stimulated by B10 GAT-M phi. The suppressor T cell mechanism was composed of at least two regulatory T cell subsets. Suppressor-inducer T cells were Lyt-2-, I-J+ and must be derived from immune spleen cells. Suppressor-effector T cells can be derived from virgin or immune spleens and were Lyt-2+ cells. When the suppressor mechanism was disabled by treatment with 1000 rad gamma irradiation or removal of Lyt-2+ cells, Lyt-2-helper T cells from (B10 X B10.D2)F1 mice primed with B10 GAT-M phi provided radioresistant help to virgin F1 B cells stimulated by B10 but not B10.D2 GAT-M phi. Suppressor inducer Lyt-2-,I-J+ cells from B10 GAT-M phi-primed (B10 X B10.D2)F1 mice were separated from the primed Lyt-2-,I-J-helper T cells. In the presence of Lyt-2+ suppressor effector cells, the Lyt-2-,I-J+ suppressor-inducer suppressed the primary response of virgin spleen or virgin T plus B cells stimulated by both B10 and B10.D2 GAT-M phi. Therefore, suppressor T cells were able to suppress primary but not secondary GAT-specific PFC responses stimulated by either parental GAT-M phi. These results showed that immunization of (responder X responder)F1 mice with parental GAT-M phi results in the development of antigen-specific helper and suppressor T cells. The primed helper T cells were radioresistant and were genetically restricted to interact with GAT in association with the major histocompatibility complex antigens of the M phi used for in vivo priming.(ABSTRACT TRUNCATED AT 400 WORDS)
Expression of idiotype on the surface of human B cells producing anti-DNA antibody.
Authors: Tamate E , Sasaki T , Muryoi T , Takai O , Otani K , Tada K , Yoshinaga K
Published: December 2022

We analyzed the idiotype (Id) expression on the surface of human anti-DNA antibody-producing cells. Murine monoclonal anti-Id antibodies with a specificity for determinants associated with the antigen-binding sites of human monoclonal anti-DNA autoantibodies were prepared. One anti-Id antibody reacted only with surface Id on anti-ssDNA-producing cells, but not with those on anti-dsDNA-producing B cell clones. Another anti-Id antibody did bind the surface Id on anti-dsDNA clones, but not those on anti-ssDNA clones. The interaction between anti-Id and surface Id was inhibited by pretreatment of the clones with DNA or appropriate polynucleotide antigens, or by preabsorption of anti-Id antibodies with free anti-DNA antibodies. Surface IgM and IgD expressed the same Id as the antibody secreted from the clones. The treatment of Id-positive clones by anti-Id antibody induced the redistribution of surface Id on the cells, indicating that these cells serve as targets for the regulatory action of anti-Id antibody.
Studies on human blood lymphocytes with iC3b (type 3) complement receptors. I. Granular, Fc-IgG receptor positive and negative subsets in healthy subjects and patients with systemic lupus erythematosus.
Authors: Bakke A C , Gray J D , Abo W , Quismorio F P , Lash A , Cooper S M , Horwitz D A
Published: December 2022

By using the OKM1 monoclonal antibody and the fluorescence-activated cell sorter to identify lymphocytes bearing iC3b (type 3) complement receptors, two principal populations of OKM1+ lymphocytes have been identified in human peripheral blood. One subset exhibited azurophilic granules and Fc receptors for IgG stained by Leu-11. The other population did not display FcR, but was enriched in cells reacting with OKT3 and OKT8 (low intensity). In healthy subjects, approximately 60% of CR3+ lymphocytes were granular FcR-bearing cells and only 18% co-expressed OKT3 determinants. In patients with systemic lupus erythematosus (SLE), CR3+ lymphocytes were predominantly FcR negative cells and 71% lacked granules. Only 33% reacted with Leu-11, but 50% co-expressed OKT3, 44% reacted with OKT8+, and 15% were OKT4+. We tested the hypothesis that agranular OKT3+ Leu-11- lymphocytes, such as those found in SLE patients, contained the precursors of natural killer (NK) cells. Leu-11+ cells were removed from normal lymphocytes by complement lysis, and the remaining cells were treated with recombinant IFN-alpha, IFN-gamma, or IL 2. These procedures were ineffective in generating typical NK effector cells. Our studies do not support the hypothesis that CR3+ Leu-11- lymphocytes are the precursors of granular Leu-11+ NK cells.
Modulation of in vitro myelopoiesis by LGL: different effects on early and late progenitor cells.
Authors: Matera L , Santoli D , Garbarino G , Pegoraro L , Bellone G , Pagliardi G
Published: December 2022

We describe here the modulatory activity of human peripheral blood natural killer (NK) cells on the growth and differentiation of myeloid progenitor cells at different stages of maturation. NK-enriched cell fractions containing 54 to 75% large granular lymphocytes (LGL) and displaying high levels of NK activity significantly inhibited the growth of late (7 day) granulocyte-macrophage colony-forming cells (CFU-GM) from about 50% of normal human bone marrow samples. However, the same fractions strongly enhanced the growth of early (14 day) stem cells from peripheral blood. Enhancing activity on early CFU-GM from blood was greater in highly purified NK cell preparations containing 96% LGL than in NK-depleted T cell preparations from the same donors. Analogous to the results when using the NK-enriched fractions, the NK-purified preparations inhibited late CFU-GM and stimulated the early ones. We conclude from these observations that human LGL have a modulatory effect on myelopoiesis depending on the maturation stage of the progenitor cell.
Induction of macrophage growth by lipids.
Authors: Yui S , Yamazaki M
Published: December 2022

Lipoproteins from ascitic tumors in mice and lipids extracted from these lipoproteins induced growth of murine peritoneal macrophages in vitro. The lipid components with activity were examined by use of lipid vesicles or liposomes. Liposomes prepared from egg-yolk PC alone did not induce macrophage growth, but those prepared from mixtures of egg-yolk PC and cholesterol or cholesteryl esters other than cholesteryl oleate, or triglycerides other than triolein, enhanced 3H-TdR incorporation into macrophages. The free fatty acids examined had no effect on 3H-TdR incorporation. These results suggest that growth of macrophages is induced by ordinary lipids present in lipoproteins or cell membranes that the macrophages scavenge in the body.
Expression of the transferrin receptor gene during the process of mononuclear phagocyte maturation.
Authors: Hirata T , Bitterman P B , Mornex J F , Crystal R G
Published: December 2022

The expression of transferrin receptors by blood monocytes, human alveolar macrophages, and in vitro matured macrophages was evaluated by immunofluorescence, radioligand binding, and Northern analysis, using the monoclonal anti-human transferrin receptor antibody OKT9, [125I]-labeled human transferrin and a [32P]-labeled human transferrin receptor cDNA probe, respectively. By immunofluorescence, the majority of alveolar macrophages expressed transferrin receptors (86 +/- 3%). The radioligand binding assay demonstrated the affinity constant (Ka) of the alveolar macrophage transferrin receptor was 4.4 +/- 0.7 X 10(8) M-1, and the number of receptors per cell was 4.4 +/- 1.2 X 10(4). In marked contrast, transferrin receptors were not present on the surface or in the cytoplasm of blood monocytes, the precursors of the alveolar macrophages. However, when monocytes were cultured in vitro and allowed to mature, greater than 80% expressed transferrin receptors by day 6, and the receptors could be detected by day 3. Consistent with these observations, a transferrin receptor mRNA with a molecular size of 4.9 kb was demonstrated in alveolar macrophages and in vitro matured macrophages but not in blood monocytes. Thus, although blood monocytes do not express the transferrin receptor gene, it is expressed by mature macrophages, an event that probably occurs relatively early in the process of monocyte differentiation to macrophages.
Decrease in IgE Fc receptor expression on mouse bone marrow-derived mast cells and inhibition of paf-acether formation and of beta-hexosaminidase release by dexamethasone.
Authors: Benhamou M , Ninio E , Salem P , Hieblot C , Bessou G , Pitton C , Liu F T , Mencia-Huerta J M
Published: December 2022

The effect of dexamethasone (DM) on the immunologic and nonimmunologic release of paf-acether and of the granule marker beta-hexosaminidase (BHEX) from mouse bone marrow-derived mast cells (BMMC) was studied. BMMC (1 X 10(6] in a modified Tyrode's solution containing 0.25% bovine serum albumin (BSA) were sensitized with an optimal dose of dinitrophenyl (DNP)-specific monoclonal IgE, and were washed before challenge with 40 ng/ml of DNP coupled to BSA. Preincubation of BMMC for 24 hr with 1 nM to 1 microM DM inhibited in a dose-dependent fashion the immunologic release of paf-acether and of BHEX as compared with control cells, with a half-maximal effect at 20 nM and 4 nM respectively. By contrast, the ionophore A23187 (1 microM)-induced release of paf-acether and of BHEX was unaffected by DM pretreatment. Finally, the antigen-induced increase in acetyltransferase activity, used as an index of cellular activation, was inhibited by 37 +/- 16% in 1 microM DM-treated BMMC as compared with untreated cells. Preincubation of BMMC with DM for 24 hr caused a dose-dependent inhibition of 125I-IgE binding to the cells, with a half-maximal effect at 14 nM. As determined by Scatchard analysis, the number of IgE Fc receptors was decreased by 55% in 1 microM DM-treated BMMC as compared with untreated cells, although the dissociation constants were comparable (control: 12.6 +/- 4.1 nM; DM-treated cells: 14.1 +/- 6.7 nM; mean +/- 1 SD; n = 3). Cytofluorometer analysis of BMMC sensitized with a saturating amount of purified monoclonal IgE, followed by addition of a fluoresceinated anti-mouse IgG (heavy and light chains), revealed a single cellular population for both DM-treated and untreated BMMC. This demonstrates that the DM-induced decrease in IgE Fc receptor expression was exhibited by every BMMC. The possible link between the decreased sensitization of the cells consequent to the reduction in IgE Fc receptor expression and the alteration of the secretory response and acetyltransferase activity was investigated. BMMC were incubated with IgE under experimental conditions giving half-sensitization of the cells. Upon antigen challenge, a 10.5 +/- 3.7% decrease in acetyltransferase activity and a 29.2 +/- 3.5% decrease in paf-acether release were observed with half-sensitized cells as compared with cells sensitized with a saturating amount of IgE.(ABSTRACT TRUNCATED AT 400 WORDS)
Characterization of the formyl peptide chemotactic receptor appearing at the phagocytic cell surface after exposure to phorbol myristate acetate.
Authors: Gardner J P , Melnick D A , Malech H L
Published: December 2022

We examined the biochemistry and subcellular source of new formyl peptide chemotactic receptor appearing at the human neutrophil and differentiated HL-60 (d-HL-60) cell surface after stimulation with phorbol myristate acetate (PMA). Formyl peptide receptor was analyzed by affinity labeling with formyl-norleu-leu-phe-norleu-[125I]iodotyr-lys and ethylene glycol bis(succinimidyl succinate) followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and densitometric analysis of autoradiographs. PMA, a specific granule secretagogue, increases affinity labeling of formyl peptide receptors on the neutrophil surface by 100%, and on d-HL-60, which lack specific granule markers, by 20%. Papain treatment markedly reduces surface labeling of formyl peptide receptor in both neutrophils and d-HL-60, and results in the appearance of a lower m.w. membrane-bound receptor fragment. PMA stimulation of papain-treated cells increases uncleaved surface receptor on neutrophils by 400%, and on d-HL-60 by only 45%. This newly appearing receptor is the same apparent m.w. (55,000 to 75,000 for neutrophils; 62,000 to 80,000 for d-HL-60) and yields the same papain cleavage product (Mr, 31,000 for neutrophils; Mr, 29,000 for d-HL-60) as receptor on the surface of unstimulated cells. Formyl peptide receptor detected by affinity labeling in neutrophil specific granule-enriched subcellular fractions is identical to receptor found on the surface of unstimulated cells appearing as equal amounts of two isoelectric forms (isoelectric points, 5.8 and 6.2) at Mr 55,000 to 70,000. There is twice as much receptor present in the specific granule-enriched fraction per cell equivalent compared with plasma membrane. Azurophil granules contain trace amounts of receptor. Similar analysis of neutrophils treated with papain before subcellular fractionation shows that papain cleaved receptor fragment is detectable almost exclusively in the plasma membrane-enriched fraction. Most of the affinity-labeled formyl peptide receptor present in specific granule enriched fraction is present in membranes other than plasma membrane or Golgi membrane, because specific granule-enriched fraction contains only a small amount of plasma membrane marker and an amount of Golgi membrane marker equal to that found in plasma membrane-enriched fraction.(ABSTRACT TRUNCATED AT 400 WORDS)
Natural cytotoxicity against mouse hepatitis virus-infected target cells. I. Correlation of cytotoxicity with virus binding to leukocytes.
Authors: Holmes K V , Welsh R M , Haspel M V
Published: December 2022

Spleen cells from uninfected control mice selectively lysed BALB/c 3T3 fibroblasts infected with mouse hepatitis virus (MHV), a murine coronavirus. Lysis of infected cells occurred within 3 hr, and histocompatibility between effector and target cells was not required. This natural, cell-mediated, virus-associated cytotoxicity differed from NK cell- and T cell-mediated lysis. Spleen cells from animals infected with MHV were enriched in NK activity and were more cytotoxic to YAC-1 target cells, but did not show enhanced cytotoxicity for MHV-infected target cells. Spleen cells from beige mice, which are deficient in NK cell activity, were able to lyse MHV-infected target cells, as were spleen cells from nude mice, which are deficient in T cell activity. Lysis of MHV-infected target cells could be mediated by cells from the spleen and, to a lesser extent, by cells from the bone marrow, but not by resident peritoneal cells or thymocytes. We suggest the term "virus killer (VK) activity" for this phenomenon. VK activity of splenocytes from different mouse strains correlated with the ability of the splenocytes to bind purified radiolabeled MHV virions. MHV virions caused agglutination of spleen leukocytes from susceptible mouse strains, indicating that leukocyte agglutination or adsorption may provide a useful assay for coronaviruses such as MHV which lack hemagglutinating activity. SJL mouse splenocytes did not bind MHV and did not lyse infected targets. MHV bound relatively well to splenocytes of other mouse strains, but poorly to thymocytes and erythrocytes. Binding of MHV to leukocytes was not influenced by 6 mM EDTA or EGTA, indicating a lack of requirement for Mg++ or Ca++. VK activity was also resistant to EDTA and EGTA, in contrast to NK activity, which was sensitive to those chelating agents. VK activity was also unaffected by actinomycin D, cycloheximide, or puromycin, indicating that new protein synthesis was not required for lysis. Antibody to interferon-alpha/beta did not block lysis, nor was there substantially enhanced lysis mediated by leukocytes from mice infected with virus and thus exposed to high levels of interferon. VK activity was blocked by antibody directed against the peplomeric glycoprotein E2 of MHV. VK activity required infected target cells, because cells with adsorbed MHV virions were not lysed by splenocytes.(ABSTRACT TRUNCATED AT 400 WORDS)