Showing papers on "Receptor expression published in 1986"

Recombinant human granulocyte-macrophage colony-stimulating factor stimulates in vitro mature human neutrophil and eosinophil function, surface receptor expression, and survival.

Abstract: A purified recombinant human granulocyte-macrophage colony stimulating factor (rH GM-CSF) was a powerful stimulator of mature human eosinophils and neutrophils. The purified rH GM-CSF enhanced the cytotoxic activity of neutrophils and eosinophils against antibody-coated targets, stimulated phagocytosis of serum-opsonized yeast by both cell types in a dose-dependent manner, and stimulated neutrophil-mediated iodination in the presence of zymosan. In addition, rH GM-CSF enhanced N-formylmethionylleucylphenylalanine(FMLP)-stimulated degranulation of Cytochalasin B pretreated neutrophils and FMLP-stimulated superoxide production. In contrast, rH GM-CSF did not promote adherence of granulocytes to endothelial cells or plastic surfaces. rH GM-CSF selectively enhanced the surface expression of granulocyte functional antigens 1 and 2, and the Mo1 antigen. rH GM-CSF induced morphological changes and enhanced the survival of both neutrophils and eosinophils by 6 and 9 h, respectively. These experiments show that granulocyte-macrophage colony stimulating factor can selectively stimulate mature granulocyte function.

Decreased production of interferon-gamma by human neonatal cells. Intrinsic and regulatory deficiencies.

Abstract: Human neonatal lymphocytes produced little macrophage activation factor in response to mitogens. This correlated with decreased production of interferon-gamma (IFN gamma): adult lymphokines contained 894.2 +/- 177.1 U/ml, whereas neonatal cord and peripheral lymphokines contained 66.9 +/- 17.0 and 116.7 +/- 29.6 U/ml by bioassay. Results by radioimmunoassay (RIA) for IFN gamma were similar. In contrast, the interleukin 2 content of cord lymphokines was greater (P less than 0.01) and that of neonatal peripheral blood lymphokines similar to that of adults. Interleukin 1 production and interleukin 2 receptor expression and affinity were similar for adult and neonatal cells. Interleukins 1 and 2 in amounts comparable to those in adult lymphokines did not increase production of macrophage activation factor or IFN gamma by neonatal cells. Neonatal cells did not contain intracellular IFN or degrade exogenous IFN. Excess suppressor activity was not found in neonatal cultures. Addition of IFN alpha, 10,000-50,000 U/ml of interleukin 2 or phorbol myristate acetate (PMA) to cord mononuclear cells or of adult monocytes or PMA to cord T cells increased IFN gamma production compared to cells stimulated with concanavalin A (ConA) alone. Nevertheless, under optimal conditions (T cells + PMA + Con A), adult cells produced much more IFN gamma (1,360 +/- 261 U/ml by RIA) than cord cells (122 +/- 37 U/ml). Staphylococcal enterotoxin A (SEA) stimulated cord cell IFN gamma production at low cell densities; nevertheless, adult cells produced more IFN in response to SEA 1,341 +/- 350 U/ml) than cord cells (350 +/- 33 U/ml). Decreased production of IFN gamma by neonatal cells appears to be due both to differences in their intrinsic capacity to produce IFN gamma and to differences in regulatory mechanisms.

The human interleukin-2 receptor: normal and abnormal expression in T cells and in leukemias induced by the human T-lymphotropic retroviruses.

Abstract: The human receptor for interleukin-2 (T-cell growth factor) plays a critical role in the growth of T cells and is required for full expression of the normal immune response. Through hybridoma and recombinant DNA techniques, the interleukin-2 receptor protein has been biochemically characterized and purified; full-length copies of its complementary DNA have been molecularly cloned, sequenced, and expressed in eukaryotic cells; and the receptor gene has been characterized. Transient expression of the interleukin-2 receptor gene occurs during normal T-cell activation, and high- and low-affinity forms of the membrane receptor exist. A naturally occurring, soluble receptor has also been isolated, and its levels in serum correlate with the activity of various diseases. Deregulation of interleukin-2 receptor expression occurs in T-cell leukemias produced by the human T-lymphotropic retroviruses types I and II (HTLV-I and -II) and has been causally linked to the action of the trans-activator (taf) gene of these viruses. Monoclonal antibodies specific for the interleukin-2 receptor are being evaluated in the treatment of HTLV-I-induced leukemias and other conditions involving the inappropriate function of activated T cells.

Decreased expression of the C3b/C4b receptor (CR1) and the C3d receptor (CR2) on B lymphocytes and of CR1 on neutrophils of patients with systemic lupus erythematosus

Abstract: Decreased numbers of complement receptor type 1 (CR1) have been observed on erythrocytes of patients with systemic lupus erythematosus (SLE) and on glomerular podocytes of patients having proliferative nephritis of SLE. In the present study, the analysis of the cellular expression of CR1 has been extended to include leukocytes. In addition, expression by B lymphocytes of the C3d receptor (CR2), which also serves as the receptor for the Epstein-Barr virus, was assessed. Receptor expression was measured by 2-color fluorescent flow cytometry of peripheral blood B cells, identified by the presence of the B1 antigen, that had also been stained with anti-CR1 or anti-CR2. B cells from 17 patients with SLE exhibited a mean relative fluorescence for CR1 that was 61% of that found in 17 normal individuals (P less than 0.001). The expression of CR2 by the patients' B cells (n = 14) was 62% of that of the B cells from normal subjects (n = 17) (P less than 0.001). The expression of CR1 correlated with that of CR2 among patients (r = 0.63; P less than 0.01) but not with the expression of CR2 among normal individuals (r = 0.36; P greater than 0.1). The mean CR1 content of the patients' neutrophils was only 59% of the normal mean (P less than 0.001). Thus, abnormalities of complement receptor expression occur on the leukocytes of patients with SLE. These deficiencies may be secondary to interaction of the cells with the products of complement activation, or, in some individuals, the deficiencies may be familial.

Exudation primes human and guinea pig neutrophils for subsequent responsiveness to the chemotactic peptide N-formylmethionylleucylphenylalanine and increases complement component C3bi receptor expression.

Abstract: After circulating in the vascular system a short time, polymorphonuclear leukocytes (PMN) migrate to extravascular sites in response to chemotactic stimuli. Prestimulation of PMN in vitro by secretagogues has been shown to increase their number of N-formylmethionylleucylphenylalanine (fmet-leu-phe) and complement component C3bi (CR3) receptors. We investigated whether the same phenomenon occurred in vivo, comparing characteristics of human skin chamber and guinea pig peritoneal exudate and blood PMN. Exudate PMN of both species contained approximately 28% less of the specific granule marker vitamin B12-binding protein (P less than 0.01) but a similar amount of the azurophil granule marker beta-glucuronidase. The total number of fmet-leu-phe receptors was 5.9 times higher in guinea pig exudate than in blood PMN (P less than 0.01) and 2.9 times higher in human exudate than in blood PMN (P less than 0.02). All exudate PMN and most blood PMN preparations showed a high affinity receptor (Kd approximately 2.3 X 10(-8) M) and a low affinity receptor (approximately 1.5 X 10(-7) M). The upregulation of fmet-leu-phe receptors in exudate PMN correlated with an improved responsiveness to fmet-leu-phe induced membrane depolarization, oxidative metabolism, and chemotaxis. In addition, the concentration of fmet-leu-phe that produced a half-maximal response of chemotaxis, superoxide production, and membrane potential depolarization was 10-fold lower in exudate PMN than in blood PMN. Human exudate PMN had a twofold increased C3bi receptor expression compared with blood PMN. Thus, a preferential loss of specific granules is associated with increased number of high and low affinity fmet-leu-phe receptors and increased C3bi receptor expression not only in vitro, but also in vivo. The data indicate that exudation primes PMN for their subsequent responsiveness to fmet-leu-phe, a modification that may be crucial for efficient antimicrobial host defense.

Interleukin 1-dependent induction of both interleukin 2 secretion and interleukin 2 receptor expression by thymoma cells.

Abstract: The macrophage-derived product, interleukin 1 (IL 1) is thought to play an important regulatory role in the proliferation of T lymphocytes; however, its mechanism of action is unknown We describe in this report a variant subline of EL4 thymoma cells (EL4-61) that displays a high degree of responsiveness to IL 1 We show that recombinant IL 1 can induce both the secretion of interleukin 2 (IL 2) and the expression of IL 2 receptors (IL 2-R) by these cells EL4-61 cells do not constitutively secrete IL 2, nor do they express IL 2-R; but when cultured in the presence of recombinant IL 1, they secrete detectable amounts of IL 2 (5 to 15 U/ml) In the presence of either suboptimal levels of phorbol ester (PMA) or Ionomycin, the addition of IL 1 resulted in up to an 80-fold enhancement in the amount of IL 2 secreted Stimulation with IL 1 alone or in combination with Ionomycin was unable to induce detectable IL 2-R expression by EL4-61 cells However, in the presence of suboptimal concentrations of PMA, IL 1 induced expression of about 3000 high affinity (dissociation constant, Kd of 31 pM) and 50,000 low affinity (Kd of 2800 pM) IL 2-R These IL 2-R were functional, based on their ability to rapidly internalize IL 2 This model system will allow a detailed analysis of the mechanisms involved in the regulation of the immune response by IL 1 and IL 2

Progestin Regulation of Epidermal Growth Factor Receptor in Human Mammary Carcinoma Cells

Abstract: Epidermal growth factor (EGF) receptors are present in human breast cancer and probably mediate the effects of EGF and the autocrine effects of α-transforming growth factors, produced by breast cancer cells. Steroid hormones influence the growth of some human cancers, and both direct and indirect effects on cell proliferation have been proposed. One potential indirect effect of steroids would be to augment sensitivity to other endocrine and autocrine factors by up-regulation of their receptors. We therefore investigated the effects of various steroids on EGF receptor expression in T-47D, MCF-7, and BT 20 human mammary carcinoma cells in culture. Preincubation of T-47D cells for 24 h with a series of androgens, estrogens, glucocorticoids, and progestins resulted in a significant enhancement of specific 125I-EGF binding in the presence of progestins only. Increased binding of EGF was associated with neither a change in cell number nor changes in the specific binding of concanavalin A, insulin, or calcitonin but was accompanied by an increase in lactogenic receptor expression. When assayed at 20°C, increased EGF binding was due to an increase in receptor number (33,380 ± 7,410 sites/cell in control cultures; 67,460 ± 20,330 sites/cell in cultures treated with 1 nm medroxyprogesterone acetate for 24 h; P < 0.05) without a change in receptor affinity. Two- to 3-fold increases in receptor number were also apparent when binding was measured at 4°C, indicating that the effect was due to an increase in expression of receptor at the cell surface rather than progestin effects on internalization and degradation. These data illustrate that the expression of EGF receptor in some breast cancer cells is regulated in part by mechanisms mediated via the progesterone receptor, since the effect was confined to progestins, potency among a series of progestins was correlated with their affinities for progesterone receptor, and sensitivity among the three cell lines studied was related to the presence and concentration of cellular progesterone receptor.

Altered erythrocyte C3b receptor expression, immune complexes, and complement activation in homosexual men in varying risk groups for acquired immune deficiency syndrome.

Abstract: We studied levels of erythrocyte C3b receptors (E-CR1) and correlated them to the level of circulating immune complexes (CIC) and complement activation in patients with or at risk for acquired immunodeficiency syndrome (AIDS). A significant reduction was found in patients with AIDS (185 +/- 93 CR1/cell), AIDS-related complex, and generalized lymphadenopathy, whereas healthy male homosexuals or normal controls had 434 +/- 193 and 509 +/- 140 CR1/cell, respectively (P less than 0.001). Family studies indicate that this defect is acquired. Reduction in E-CR1 was associated with increased levels of CIC when assayed by binding to Raji cells, but not when tested by C1q binding. Complement activation was assessed by levels of C3bi/C3d-g in plasma, measured with a monoclonal antibody specific for a neoantigen in C3d. AIDS patients had increased C3 activation (2.68 +/- 1.67%) when compared with normal controls (0.9 +/- 0.22%) (P less than 0.01). The decreased E-CR1, the presence of CIC, and C3 activation suggest that complement activation by immune complexes may play a role in the clinical expression of the disease.

Quantification and characterization of high-affinity membrane receptors for tumor necrosis factor on human leukemic cell lines.

Abstract: The expression of specific membrane receptors for TNF-alpha was determined on various human leukemic cell lines differing in their sensitivity to the growth-inhibitory activity of TNF-alpha Binding studies with 125I-labelled TNF-alpha indicated specific binding in 8/10 cell lines with approximately 10-fold differences in the quantity of TNF-alpha bound by these distinct cell lines Scatchard analyses of TNF-binding revealed the existence of high-affinity membrane receptors (Kd 15 X 10(-10) M) and approximately 3,000 binding sites/cell on both U937 and K562, representing 2 cell lines with high and low TNF sensitivity, respectively Disuccinimidyl-suberate cross-linking of receptor-bound 125I-TNF-alpha and SDS-PAGE of membrane preparations of either U937 or K562 cells suggest a single receptor protein with an apparent molecular weight of 76 kDa Comparison of the TNF-alpha binding capacity versus in vitro growth inhibition provides evidence that sensitivity to TNF-alpha is determined both at the level of receptor expression and at a post-receptor level IFN-gamma strongly enhanced the TNF-alpha-mediated growth inhibition of 3 sensitive cell lines, but had no effect on 7 other leukemic cell lines with little or no TNF sensitivity No correlation was found between this enhancement of TNF sensitivity and the IFN-gamma-mediated increase in TNF-cell membrane receptors, suggesting that IFN-gamma predominantly exerts its synergistic effect distal to TNF-binding

Presence of preactivated T cells in hemodialyzed patients: their possible role in altered immunity

Abstract: Interleukin 2 (IL-2) and B-cell growth factors I and II (BCGF I and BCGF II) are lymphokines produced by T cells that play a major role in T- and B-cell cooperation. Peripheral blood lymphocytes from 12 uremic patients undergoing intermittent hemodialysis were tested for their capacity to produce IL-2 and BCGFs and to respond to these soluble mediators. IL-2 and BCGF activities were determined by means of two biological assays (proliferation of IL-2-dependent cytotoxic T-cell line CTLL-2 and of anti-human IgM (mu chain)-stimulated normal B cells, respectively) in the supernatants of phytohemagglutinin A-stimulated T-cell cultures. IL-2 activity was significantly decreased in patients as compared to normal controls (mean +/- SEM, 0.28 +/- 0.09 unit per ml) in hemodialyzed patients versus 1.02 +/- 0.16 units per ml in normal controls). This profound abnormality contrasted with the normal activity of the BCGFs that was invariably observed in the same supernatants. A similar dissociation was detected when analyzing the sensitivity of uremic B and T cells to exogenous purified lymphokines. Anti-IgM (mu chain)-stimulated uremic B cells exhibited a normal response to recombinant IL-2 and to chromatography-purified BCGF I and BCGF II. Resting B cells did not show any increased reactivity to these lymphokines. In contrast, whereas in normal controls recombinant IL-2 exclusively induced the proliferation of T cells that had been previously activated by a mitogen, resting T cells from uremic patients were highly responsive to exogenous IL-2. This abnormal response was paralleled by significantly increased proportions of peripheral T cells recognized by the anti-Tac monoclonal antibody that specifically binds to the IL-2 receptor. These data clearly show the existence in hemodialyzed patients of abnormally high proportions of T cells presenting phenotypic and functional signs of preactivation. This increased T-cell IL-2 receptor expression may offer an explanation to the deficient IL-2 activity observed in patients' supernatants (by inducing increased absorption of the lymphokine). The potential relevance of these preactivated T cells to the depressed cell-mediated immunity observed in hemodialyzed patients is outlined.

Transferrin synthesis by inducer T lymphocytes.

Abstract: Transferrin (Tf) is a growth factor that transports iron in plasma. It is essential for proliferation of activated T lymphocytes. Previous studies have suggested that peripheral blood cells are capable of synthesizing Tf. Using in situ hybridization techniques and human Tf complementary DNAs as probes, peripheral blood cells have been examined for sites of Tf messenger RNA (mRNA) transcription. The studies described here demonstrate that Tf is synthesized by a specific subset of T lymphocytes, the T4+ inducer subset. T lymphocyte proliferation is dependent upon the presence of both interleukin 2 (IL-2) and Tf, even though resting cells do not possess receptors for either. The present studies indicate that during T cell activation, induction of IL-2 mRNA transcription and IL-2 receptor expression precede the transcription of Tf mRNA and expression of Tf receptors, respectively. These events in turn precede the initiation of DNA synthesis. Transferrin and its receptor appear to be involved in an autocrine pathway which is functionally linked to the IL-2/IL-2 receptor autocrine loop.

Modulation of interleukin 2 receptor expression on normal human lymphocytes by thymic hormones.

Abstract: The expression of interleukin 2 receptors (IL-2R) is a critical step leading to normal lymphocyte proliferation. Since thymosin fraction 5 (TF5), a thymic hormone preparation, enhances lymphoproliferative responses of human cells, we examined the effects of TF5 on the expression of IL-2R on mitogen-stimulated human lymphocytes. TF5 significantly increased the percentage and antigen density of cells expressing IL-2R after stimulation with an optimal concentration of phytohemagglutinin (PHA) when the cells from the same donor exhibited suboptimal responses to PHA alone. The same effect was observed with a suboptimal PHA concentration and with OKT3 monoclonal antibody stimulation. Thymosin alpha 1, a synthetic polypeptide originally isolated in its native form from TF5, was also able to increase IL-2R expression in response to PHA, suggesting that it is the active species in TF5. The enhancement of IL-2R expression was paralleled by increased proliferative responses. Increased IL-2R expression appears to be the direct effect of thymic hormones, since abrogation of interleukin 2 production by cyclosporin A did not affect TF5-mediated enhancement of PHA-induced IL-2R expression. These results point to a physiological role of thymic hormones in the maintenance of normal levels of IL-2R expression. This immunoregulatory activity of thymic hormones might be relevant in the treatment of conditions where there is decreased IL-2R expression, such as the acquired immune-deficiency syndrome, or in the restoration of normal IL-2R expression to lymphocytes from aged individuals.

The interleukin-2 receptor, its physiology and a new approach to a selective immunosuppressive therapy by anti-interleukin-2 receptor monoclonal antibodies.

Abstract: In this report we have summarized our findings on the IL-2 receptor and our attempts to find an IL-2 receptor targeted immunosuppressive therapy. IL-2 receptors are detectable in two different forms: as monomeric, surface expressed, and as dimeric, presumably non-surface expressed molecules. The dimeric form seems to be non-covalently bound to an as yet undefined 110 KD molecule. The functions of the monomeric versus the dimeric form, as well as that of the noncovalently bound molecule, and their relationship to high and low affinity IL-2 receptors are not yet clear and remain to be elucidated. Upon antigenic or mitogenic stimulation, IL-2 receptors became expressed at the surface of T-lymphocytes. Receptor expression is accompanied by the capacity of the cells to proliferate in response to IL-2. Resting T-cells do not proliferate in response to IL-2. IL-2 dependent proliferation of cells without external stimulation is either due to the presence of a small number of IL-2 receptor bearing cells in the respective population or due to a small number of IL-2 receptors present on the surface of cells. IL-2 itself does not induce IL-2 receptor expression on resting cells but has been shown to up-regulate its own receptor once expressed. In contrast to resting lymphocytes, some leukemic cells and early embryonic thymocytes in the species tested constitutively express IL-2 receptors. The role of such constitutively expressed receptors is not yet clear. As demonstrated in mice, the requirement(s) for induction of IL-2 receptor expression for the helper/inducer subset (Lyt-2+) are different from those of the cytotoxic/suppressor subset (L3T4+). In contrast to Lyt-2+ cells, the accessory cell requirement for L3T4+ cells could not be replaced by cytokines. Whether Lyt-2+ cells require an additional, not yet defined receptor inducing factor (RIF) besides IL-2 in order to become IL-2 receptor positive and to consequently proliferate in response to IL-2, is a matter of controversy. There is evidence that interleukin-1 and some functionally related factors produced by leukemic cells enhance expression and/or function of the IL-2 receptors. IL-2 receptors of the high and low affinity type expressed upon antigenic stimulation are cleared from the cell surface. As demonstrated in this report, the vast majority of them, probably those of low affinity type, are released from the cells continuously. The mechanism of their release and the possible immunoregulatory role of circulating receptors found in the serum of animals is not yet clear.(ABSTRACT TRUNCATED AT 400 WORDS)

The Antiproliferative Effect of Calcitriol on Human Peripheral Blood Mononuclear Cells

Abstract: Activation of lymphocytes leads to the expression of receptors for the calcitropic hormone calcitriol [1,25(OH)2D3], and calcitriol is a potent inhibitor of interleukin– 2 (IL–2) and of lymphocyte proliferation. We used peripheral blood mononuclear cells (PBM) activated in vitro with phytohemagglutinin to study 1) the relationship between 1,25(OH)2D3 receptor expression, IL–2 production, and l,25(OH)2D3–induced inhibition of PBM proliferation in connection with the cell cycle; 2) the effect of 1,25(OH)2D3 on PBM activation and on the expression of activation–related molecules including the IL–2 receptor, and 3) the role of calcium in the antiproliferative effect of the hormone. 1,25(OH)2D3 receptor expression occurred when PBM entered the Gla phase of the cell cycle. The concentrationof the receptor protein reached a peak at G1b and declinedduring the S phase. 1,25(OH)2D3 inhibited cell proliferation by blocking PBM at the Gla–Glb border. The antiproliferative effect of calcitriol was not caused by hormon...

Down-regulation of interleukin 1 (IL 1) receptor expression by IL 1 and fate of internalized 125I-labeled IL 1 beta in a human large granular lymphocyte cell line.

Abstract: The regulation of interleukin 1 (IL 1) receptor expression on a human large granular lymphocyte cell line, YT, and fate of internalized 125I-labeled IL 1 beta (125I-IL 1 beta) were studied. YT cells were selected for this study, because this cell line expresses a large number of specific high-affinity receptor for IL 1, responds biologically to exogenously added IL 1 by expressing high-affinity IL 2 receptors, and does not produce IL 1. YT cells constitutively express approximately 7 X 10(3) IL 1 receptors/cell with a Kd approximately 10(-10) M. Neither IL 2, phorbol myristic acid, nor lipopolysaccharide affected the total binding of 125I-IL 1 beta by YT cells. In contrast, the capacity of YT cells to bind 125I-IL 1 beta when incubated at 37 degrees C for 3 to 16 hr with a low dose of purified IL 1 beta (approximately 6 U/ml) was reduced by greater than 80%. The loss of binding capability gradually recovered by 16 hr after removal of IL 1 beta from cultured YT cells. The apparent loss of IL 1 receptor expression was accompanied by the internalization of 125I-IL 1 beta into cells. Acid treatment of YT cells to remove bound 125I-IL 1 beta at 4 degrees C showed that 50% of the 125I-IL 1 beta bound to cells could no longer be recovered after 30 min at 37 degrees C, and this increased to 80% after 3 hr at 37 degrees C. Fractionation of cell extracts on Percoll gradient additionally showed 125I-IL 1 beta to appear intracellularly after receptor binding on plasma membranes, and to be successively transferred to some membranous organelles (d approximately equal to 1.037) through an intermediate density organelle (d approximately equal to 1.050), and to finally end up in lysosomal cell fractions (d approximately equal to 1.05 to 1.08) after approximately 3 hr at 37 degrees C. Only approximately 5% of internalized 125I-IL 1 beta was released into culture media by 6 hr of incubation at 37 degrees C. However, the radioactivity in the TCA soluble fraction of the culture media increased gradually by 6 hr and a lysosomotropic enzyme, ethylamine, significantly inhibited both the transfer of internalized 125I-IL 1 beta to the lysosomal fraction and the degradation of 125I-IL 1 beta. This study represents the first evidence of autoregulation of IL 1 receptors by IL 1 and internalization of IL 1 molecules after binding to receptors.(ABSTRACT TRUNCATED AT 400 WORDS)

Basis for defective responses of rheumatoid arthritis synovial fluid lymphocytes to anti-CD3 (T3) antibodies.

Abstract: Synovial fluid mononuclear cells (SFMC) from patients with active rheumatoid arthritis characteristically respond poorly to mitogens. In this study, mitogenic antibodies reactive with the CD3(T3) antigen on human T lymphocytes were used to analyze the basis for the deficiency. OKT3-induced proliferation and release of interleukin 1 (IL-1) and interleukin 2 (IL-2) from SFMC were depressed in all patients. Purified IL-1 or recombinant IL-2 restored proliferative responses in SFMC and increased IL-2 receptor density. Exogenous IL-1 also enhanced IL-2 release. Fractionation of SFMC supernatants on phosphocellulose columns revealed the presence of IL-1 and a potent IL-1 inhibitor. The monocyte-derived IL-1 inhibitor blocked IL-1-dependent responses of normal peripheral blood lymphocytes to OKT3, but had no effect on IL-2-dependent events. These results suggest that IL-1 inhibitor(s) in SFMC impair(s) OKT3-induced mitogenesis by interfering with the effects of IL-1 on T lymphocytes. The net result is deficient IL-2 secretion, IL-2 receptor expression, and impaired cellular proliferation. This novel inhibitory circuit provides a rational explanation for the diminished function of synovial fluid T lymphocytes in rheumatoid arthritis patients.

Norepinephrine and thyroid-stimulating hormone induce inositol phosphate accumulation in FRTL-5 cells.

Abstract: 3H-Labeled inositol phosphate accumulation is observed when prelabeled FRTL-5 cells (a rat thyroid cell line) are exposed to norepinephrine (NE) or TSH. The presence of inositol trisphosphate among the products implicates a phosphodiesterase- catalyzed hydrolysis of phosphatidylinositol 4,5- bisphosphate. The response to NE is much greater than that to TSH. This may be explained by the ability of cAMP to inhibit inositol phosphate accumulation in these cells. The stimulation by NE is inhibited by ai-adrenergic receptor antagonists and is markedly potentiated in medium of reduced Ca2+ concentration. After chronic withdrawal of TSH from the growth medium, the magnitude of the response to NE is considerably reduced; however, there is no substantial shift in the dose-response curve. This reflects the dependency of α1-adrenergic receptor expression on TSH in the FRTL-5 cell. In contrast, the characteristics of inositol phosphate accumulation induced by acute treatment with TSH are similar in cells maintaned in...

Cyclosporine and the immune response: basic aspects.

Abstract: Cyclosporine (CsA) is a novel immunosuppressive agent currently used clinically, to prevent rejection of solid organ allografts and to prevent graft-vs.-host disease. Early studies in a variety of animal models exhibited transplantation tolerance after limited treatment with this unique agent. The apparent specific immunological unresponsiveness induced by CsA is thought to be maintained by antigen-specific suppressor T lymphocytes. Studies attempting to dissect the mechanism of action of this unique agent suggested that CsA selectively affected different T lymphocyte populations. Cyclosporine was very effective at inhibiting the production of interleukin-2 (IL-2), a soluble lymphokine known to amplify cytotoxic T cell responses and was also capable of preventing IL-2 receptor expression on the precursor cytotoxic T lymphocyte. In contrast, to the effect on T helper cells and on the precursor cytotoxic T lymphocyte, studies in vitro and in vivo demonstrated that CsA had a sparing effect on suppressor T cell induction. More recent studies have indicated that CsA allows for the amplification of suppressor T lymphocytes independent of interleukin-2 indicating that other cellular and/or soluble factors are important for potentiation of suppressor T lymphocyte activity. However, the molecular action of CsA at the cellular level still remains unresolved. Thus, CsA is not only a useful drug in clinical transplantation but it has become increasingly important as an immunologic probe allowing the dissection of complex cellular interactions involved in the immune response.

Regulation of interferon receptor expression in human blood lymphocytes in vitro and during interferon therapy.

Abstract: Interferons (IFN) elicit antiviral and antineoplastic activities by binding to specific receptors on the cell surface. The binding characteristics of IFN to human lymphocytes were studied using IFN alpha 2 labeled with 125I to high specific activity. The specific binding curves generated were analyzed by the LIGAND program of Munson and Rodbard to determine receptor numbers. The number of receptors in peripheral blood lymphocytes (PBL) and tonsillar B-lymphocytes (TBL) from normal individuals were 505 +/- 293 (n = 10) and 393 +/- 147 (n = 3) respectively. When these cells were preincubated in vitro with unlabeled IFN alpha 2, the receptor number decreased to 82 +/- 45 and 61 +/- 16 respectively. Receptor binding activities recovered gradually over a period of 72 h when the cells were incubated in IFN-free medium. This recovery of receptors could be blocked by the addition of actinomycin D to the incubation medium. A similar decrease in receptor expression was observed in vivo in PBL from patients being treated daily with 5 X 10(6) units/m2 per d of IFN alpha 2 by subcutaneous injection, for acute lymphoblastic leukemia or papilloma virus infections. Receptor numbers in PBL in vivo were further reduced concurrent with the progression of IFN therapy. Thus the reduction in IFN receptor expression observed in vitro can be demonstrated in vivo. These studies indicate that monitoring IFN receptor expression in vivo can provide information regarding the availability of IFN receptors at the cell surface for the mediation of IFN actions during the course of IFN therapy.

Lipopolysaccharide modulates receptors for leukotriene B4, C5a, and formyl-methionyl-leucyl-phenylalanine on rabbit polymorphonuclear leukocytes.

Abstract: Peripheral blood polymorphonuclear leukocytes (PMNL) isolated from rabbits after an iv injection of endotoxin exhibited decreased chemotactic migration in response to leukotriene B4 (LTB4) and C5a, but not N-formyl-methionyl-leucyl-phenylalanine (fMLP), after endotoxin treatment The binding of radiolabeled LTB4, fMLP, and C5a to isolated PMNL was assessed in order to determine whether altered receptor expression could account for the observed functional changes Control PMNL expressed binding sites for fMLP, LTB4, and C5a similar to those previously characterized from human PMNL Control PMNL expressed a single class of 14,600 +/- 2700 receptors for fMLP with a mean dissociation constant (Kd) of 20 +/- 06 nM at 0 degrees C, whereas two subclasses of binding sites were expressed for LTB4: 10,300 +/- 6800 high-affinity and 85,600 +/- 53,000 low-affinity binding sites per PMNL with mean Kd for LTB4 of 075 +/- 043 nM and 70 +/- 58 nM (mean +/- SD, n = 5), respectively Control PMNL bound [125I]-C5a in a dose-dependent and saturable manner at 24 degrees C At saturating concentrations of C5a, PMNL obtained from control rabbits bound 270,000 +/- 50,000 molecules of [125I]-C5a with half-maximal binding occurring at [125I]-C5a concentrations of 55 +/- 19 nM The binding of LTB4 and C5a to PMNL obtained 24 hr after an iv injection of endotoxin was markedly decreased compared with control PMNL PMNL from endotoxin-treated rabbits exhibited 68% fewer high-affinity binding sites per PMNL for LTB4 and a 51% decrease in the amount of [125I]-C5a bound at saturating concentrations compared with control PMNL There was no significant change in the Kd of the high-affinity binding sites for LTB4, no change in the Kd and number of the low-affinity binding sites for LTB4, and a small decrease in the apparent Kd for C5a to 33 +/- 11 nM Even though the pretreatment with iv endotoxin did not alter chemotactic or degranulation responses elicited by fMLP, the endotoxin pretreatment induced an eightfold increase in the receptor density without altering the Kd for fMLP Decreased receptor expression could account in large part for the decreased chemotactic responsiveness towards C5a and LTB4 induced by LPS The finding that a substantial increase in receptors for fMLP need not be accompanied by a comparable functional change suggests that decreased efficiency in receptor coupling to intracellular biochemical events may also result from iv endotoxin

Characterization of C1q receptor expression on human phagocytic cells: effects of PDBu and fMLP.

Abstract: The receptor-mediated binding of C1q to human phagocytic cells was investigated in this study. By using a C1q binding assay, we determined that purified, elutriated monocytes expressed an average of 4.6 X 10(5) C1q receptors (C1qR) per cell, with an equilibrium binding constant (Keq) of 0.91 X 10(7) (M-1). When analyzed in an identical manner, the polymorphonuclear leukocytes (PMN) expressed an average of 4.2 X 10(5) C1qR per cell, with a Keq for C1q of 1 X 10(7) (M-1). Fluorescent flow cytometric analysis showed that C1q was bound by 98% of the monocytes studied. Further, the pattern formed by these cells was consistent with a normal log distribution, indicating that this was a homogeneous population of cells. When PMN were assayed with flow cytometry, however, we found that C1q was bound by an average of only 45% of the PMN analyzed. Further, these PMN were not dispersed in a normal log distribution, indicating some heterogeneity among the cells that bind C1q. We examined the abilities of the chemoattractant N-formylmethionylleucylphenylalanine (fMLP) and the phorbol ester phorbol dibutyrate (PDBu) to modulate expression of C1qR as compared to the receptor for C3b (CR1). Pretreatment of the monocytes and the PMN with either 10(-6)M fMLP or 10 ng/ml of PDBu significantly increased cell surface CR1 expression, as reported previously by other investigators. In contrast, no significant increase in the number of C1qR on the monocytes or the PMN was observed with any of the concentrations of fMLP or PDBu used during pretreatment. However, with certain pretreatment doses of these agents, some reduction was noted in the amount of 125I-C1q bound to the monocytes and the PMN. This study characterizes the binding of C1q to purified monocytes and confirms previously published values for PMN. The distribution of cells expressing C1qR is shown to be significantly different between identically treated populations of monocytes and PMN. Finally, the abilities of fMLP and PDBu to modulate the binding of C1qR are examined. Our results indicate that the control of C1qR expression differs markedly from that of CR1.

Phenotypic analysis of thymocytes that express homing receptors for peripheral lymph nodes.

Abstract: Thymocytes that express high levels of homing receptors for peripheral lymph nodes can be detected with the monoclonal antibody MEL-14. We have shown that in adult mice these rare MEL-14hi thymocytes a) are cortical in location and typically constitute 1 to 3% of the total thymocyte population, b) may be a major source of thymus emigrants, and c) contain a high frequency of precursors of alloreactive cytotoxic T lymphocytes. In this study we have analyzed the phenotype of the MEL-14hi thymocyte subset. Most normal adult MEL-14hi thymocytes are midsize and express the mature phenotype typical of thymus emigrants, medullary thymocytes, and peripheral T cells: they are predominantly PNAlo, H-2K+, Thy-1+, Ly-1hi, and either Lyt-2-/L3T4+ or Lyt-2+/L3T4-. These findings argue strongly for the presence of rare MEL-14hi immunocompetent cortical thymocytes that, aside from their homing receptor expression, are phenotypically indistinguishable from medullary thymocytes. However, a minority (20 to 30%) of MEL-14hi thymocytes are large and phenotypically nonmature: they express intermediate to high levels of PNA binding sites, and are H-2K- to H-2Klo, Thy-1hi, Ly-1+, and either Lyt-2+/L3T4+ or Lyt-2-/L3T4-. Through a technique that selectively labels outer cortical cells, phenotypically nonmature MEL-14hi thymocytes have been shown to be concentrated in the subcapsular blast region of the outer cortex. Although we have no direct evidence of a precursor-product relationship, we consider it likely that the phenotypically nonmature outer cortical MEL-14hi lymphoblasts give rise to phenotypically mature MEL-14hi cells located deeper in the cortex. These results are consistent with our previous proposal that MEL-14hi thymocytes are a major source of thymus emigrants, and indicate that expression of high levels of MEL-14-defined homing receptors may be closely linked to the intrathymic selection process.

Functional role of HLA class I cell-surface molecules in human T-lymphocyte activation and proliferation

Abstract: This investigation addressed the role of major histocompatibility complex-encoded class I molecules in the activation and proliferation of human lymphocytes. We studied the effect of antibodies specific for HLA-A and HLA-B locus gene products on mitogen-stimulated peripheral blood mononuclear cell (PBMC) subpopulations. Three individually derived, well-characterized anti-HLA class I monoclonal antibodies were demonstrated to inhibit the proliferation of human PBMC stimulated by either OKT3 or the calcium ionophore ionomycin. The antibody directed against HLA-A, -B, and -C locus gene products (W6/32) and the antibody directed against HLA-B locus gene products (4E) inhibited proliferation induced by either mitogen by 70-90%. The HLA-A locus-specific antibody (131), though inhibiting ionomycin-induced proliferation by 80-90%, was much less effective when OKT3 was the stimulus. The inhibition affected T4+ and T8+ cells and was not mediated by DR+ accessory cells. The inhibitory effect of these antibodies was associated with a decrease in the level of interleukin 2 activity present in culture supernatants, decreased interleukin 2 receptor expression, and decreased transferrin receptor expression and was not overcome by the addition of exogenous interleukin 2. Our results suggest that HLA class I molecules are directly involved in the early critical events of human lymphocyte activation and proliferation.

Multiple sclerosis: increased expression of interleukin-2 receptors on lymphocytes.

Abstract: The percentage of interleukin-2 receptor-positive peripheral blood lymphocytes in MS patients was significantly higher in acute relapse than in remission or in controls. After stimulation by phytohemagglutinin, the expression of interleukin-2 receptor on peripheral blood lymphocytes of MS patients was within the range of healthy controls, implying no general impairment of receptor expression. These results confirm other evidence that there is a small population of activated T lymphocytes in acute relapse of MS.

The Human Interleukin-2 Receptor: Analysis of Structure and Function

Abstract: Considerable information presently exists regarding the molecular, biochemical, and biological features of the human IL-2 receptor. The IL-2 receptor protein, multiple receptor mRNAs, and a single structural gene have now been identified. The important role of this receptor in normal T-cell growth is well established and its potential participation in B-cell growth and differentiation appreciated. The availability of cloned gene products for both the IL-2 receptor and IL-2 may permit the future development of novel biological agents capable of either augmenting or blunting the T-cell immune response. The intriguing interrelationship of HTLV-I and -II infection and altered IL-2 receptor expression is now being unraveled. However, the structural difference in high and low affinity receptors as well as the mechanism by which signals for T-cell growth are propagated through the high affinity receptor remain dominant, unanswered questions in the field.