A monoclonal antibody (mAb) was isolated that blocked the binding and bioactivity of both human and murine interleukin 1 beta (IL-1 beta) on murine IL-1 receptor-bearing cells. This mAb recognized a protein that was distinct from the Type I and Type II IL-1 receptors, suggesting that an additional protein exists that is involved in IL-1 biological responses. By expression cloning in COS-7 cells, we have isolated a cDNA from mouse 3T3-LI cells encoding this putative auxiliary molecule, which we term the IL-1 receptor accessory protein (IL-1R AcP). Sequence analysis of the cDNA predicts an open reading frame that encodes a 570-amino acid protein with a molecular mass of approximately 66 kDa. The IL-1R AcP is a member of the Ig superfamily by analysis of its putative extracellular domain and also bears limited homology throughout the protein to both Type I and Type II IL-1 receptors. Northern analysis reveals that murine IL-1R AcP mRNA is expressed in many tissues and appears to be regulated by IL-1. In mammalian cells expressing natural or recombinant Type I IL-1R and IL-1R AcP, the accessory protein forms a complex with the Type I IL-1R and either IL-1 alpha or IL-1 beta but not IL-1ra. The recombinant accessory protein also increases the binding affinity of the recombinant Type I IL-1R for IL-1 beta when the two receptor proteins are coexpressed. Therefore, the functional IL-1 receptor appears to be a complex composed of at least two subunits.
A cytokine that can synergize with interleukin 2 to activate cytotoxic lymphocytes was purified to homogeneity. The protein, provisionally called cytotoxic lymphocyte maturation factor (CLMF), was isolated from a human Blymphoblastoid cell line that was induced to secrete lymphokines by culture with phorbol ester and calcium ionophore. The purification method, utilizing classical and high-performance liquid chromatographic techniques, yielded protein with a specific activity of 8.5 x 107 units/mg in a T-cell growth factor assay. Analysis of the purified protein by sodium dodecyl sulfate/polyacrylamide gel electrophoresis demonstrated that CLMF is a 75-kDa heterodimer composed of disulfide-bonded 40-kDa and 35-kDa subunits. Determination of the N-terminal amino acid sequences of the two subunits revealed that both subunits are not related to any previously identified cytokine. Purified CLMF stimulated the proliferation of human phytohemagglutinin-activated lymphoblasts by itself and exerted additive effects when used in combination with suboptimal amounts of interleukin 2. Furthermore, the purified protein was shown to synergize with low concentrations of interleukin 2 in causing the induction of lymphokine-activated killer cells.The potential utility of cytokines in the treatment of neoplasia and as immunoenhancing agents has recently been demonstrated in studies using human recombinant interleukin 2 (rIL-2) (1-6). However, the clinical use of rIL-2 has been complicated by the serious side effects that it may cause (2, 3). One approach to improving the efficacy of cytokine therapy while reducing toxicity is to use two or more cytokines in combination. For example, synergistic antitumor activity has been shown to result when rIL-2 is administered to tumor-bearing mice together with recombinant interferon a (rIFN-a) (7,8) or with recombinant tumor necrosis factor a (rTNF-a) (9). The antitumor effects of rIL-2 are thought to be mediated by host cytotoxic effector lymphocytes, which are activated by rIL-2 in vivo (10). rIFN-a (11) and rTNF-a (12, 13) have been shown to synergize with rIL-2 in activating cytotoxic effector cells in vitro as well as to exert synergistic antitumor effects when given in combination with rIL-2 in vivo (7-9). Hence, a cytokine was sought that could synergize with rIL-2 to activate cytotoxic lymphocytes in vitro and thus might also have utility as an antitumor agent when administered in combination with rIL-2 in vivo.Previously we demonstrated that IL-2-depleted lymphokine-containing cell supernatant solutions from cultures of human peripheral blood lymphocytes activated with phytohemagglutinin (PHA) or in mixed lymphocyte cultures contained such a factor, provisionally called cytotoxic lymphocyte maturation factor (CLMF) (14, 15). However, the quantities of human CLMF produced by peripheral blood lymphocytes were too low to permit its purification to homogeneity. Therefore, human lymphoid cell lines were screened for the production of cytokines that could synergize with rIL-2 to a...
SummaryIt has been suggested that epidermal Langerhans cells (LC) bearing immunoglobulin E (IgE) may be involved in the genesis of atopic disease. The identity of the IgE receptor(s) on LC remained unclear, although it represents a crucial point in understanding cellular events linked to the binding of allergens to LC via IgE. In this report, we demonstrate that epidermal LC express the high affinity receptor for the Fc fragment of IgE (FceRI) which has, so far, only been described on mast cells and basophils. Epidermal LC react with antibodies specific for the u subunit of the tetrameric (oL,~,23') FceRI. Specific transcripts for Fc~RIo~ and FceRI3~ were detected in LC and correspond to those of human basophils and of the human basophil cell line KU812. Furthermore, human basophils, KU812 cells, and LC express the putative B subunit. Thus human LC express the complete structure of FceRI. This finding opens new perspectives in the putative functional role of this structure on antigen-presenting cells.T he demonstration of IgE molecules on epidermal Langerhans cells (LC) 1 in patients with atopic dermatitis has implied that these cells should perform a major function in the pathophysiology of atopic disease (1, 2). Although initially, only receptors for the Fc fragment of IgG were identified on epidermal LC (3, 4), the low affinity receptor for IgE FceRII/CD23 (5) and the human IgE binding protein (eBP) (6) have now also been found on these cells in lesional, as well as in normal skin. However, attempts to completely block IgE binding on LC by a variety of anti-FceRII/CD23, antieBP, and/or anti-Fc3,R reagents remained unsuccessful, suggesting the presence of a third IgE-binding structure actually responsible for a part of the IgE-binding capacity of LC. We report here, that normal human LC also express the high a~nity receptor for IgE, Fcelkl, demonstrating that the presence of this structure is not restricted to mast cells and basophils. Our results also document the presence of the putative/~ chain on both human basophils and LC.
Interleukin-2 (IL-2) binds to both high- and low-affinity classes of IL-2 receptors on activated T lymphocytes. Only the high-affinity receptors are involved in receptor-mediated endocytosis and normally transduce the mitogenic signals of IL-2; however, the structural features distinguishing the high- and low-affinity receptors are unknown. When 125I-labeled IL-2 was chemically cross-linked to activated human T lymphocytes, two major bands were identified. First, as predicted, a 68- to 72-kilodalton band, consisting of IL-2 (15.5 kilodaltons) cross-linked to the IL-2 receptor (55 kilodaltons), was observed. Second, an unpredicted 85- to 92-kilodalton moiety was detected. This band was not present when IL-2 was cross-linked to transfected C127 cells, which exclusively express low-affinity receptors. The data presented are most consistent with the existence of a 70- to 77-kilodalton glycoprotein subunit (p70) which, upon associating with the 55-kilodalton low-affinity receptor (p55), transforms it into a high-affinity site. It is proposed that p55 and p70 be referred to as the alpha and beta subunits, respectively, of the high-affinity IL-2 receptor.
SummaryThe Shwartzman reaction is elicited by two injections of lipopolysaccharide (LPS) in mice. The priming LPS injection is given in the footpad, whereas the lethal LPS challenge is given intravenously 24 h later. The injection of interferon 3, (IFN-'y) or interleukin 12 (IL-12) instead of the LPS priming injection induced the lethal reaction in mice further challenged with LPS. Antibodies against IFN-q/when given together with the priming agent, prevented the lethal reaction in mice primed with either LPS, IL-12, or IFN-3'. Antibodies against IL-12, when given together with the priming agent, prevented the lethal reaction in mice primed with either LPS or IL-12 but not with IFN-% These results strongly suggest that LPS induces the release of IL-12, that IL-12 induces the production of IFN-% and that IFN-7 is the cytokine that primes macrophages and other cell types. Upon LPS challenge, the lethal Shwartzman reaction is induced by a massive production of inflammatory cytokines that act on the target sites already sensitized by IFN-7. If mixtures of TNF and IL-1 or mixtures of TNF and IFN-3' are used to challenge mice previously primed with IFN-7 or ILo12, mortality is induced. In the same conditions, the individual cytokines or a mixture of IL-1 and IFN-3' do not replace the LPS challenge. When the mice are primed with LPS, the combination of TNF, IL-1, and IFN-7 induced only a partial mortality incidence suggesting that the involvement of other LPS-induced factors. Bacterial LPS is a cell wall component of Gram-negative bacteria that is responsible for most of the toxic manifestations associated with bacterial infections. In mice, a lethal shock syndrome, known as the generalized Shwartzman reaction, can be elicited by two consecutive injections of LPS (for review see 1). A priming dose of LPS, injected intradermall), in the footpad (f.p.)l, is followed after 24 h by an intravenous challenge injection of LPS. After this challenge injection, which is not lethal per se, the mice die within the following 48 h from disseminated intravascular coagulation, vascular occlusion, hemorrhage, perivascular accumulation of leukocytes, and necrosis (2). This hypersensitivity reaction occurs only if the time interval between these injections is crucially fixed at between 18 and 24 h. There is no response when the intravenous challenge of LPS is given either earlier or later, or when the order of priming and challenge 1 Abbreviations used in thispaper: f.p., footpad; h, human; m, mouse; MIF, macrophage inhibiting factor; R, receptor. injections is reversed. FinaUy, the reaction does not occur if both the LPS injections are intradermal or if the priming dose of LPS exceeds an optimum. The careful dosage and timing of the LPS injections and the need of specific routes of administration indicate that the Shwartzman reaction is elicited by induced endogenous factors acting in a precise time sequence. IFN-'y, TNF, and IL-1 are known to be involved in the pathogenesis of the generalized Shwartzman reaction. IFN-7 seems important ...
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