CD4-mediated stimulation of human eosinophils: lymphocyte chemoattractant factor and other CD4-binding ligands elicit eosinophil migration.

Lymphocyte chemoattractant factor (LCF) is a tetrameric glycoprotein of 56,000 relative molecular mass produced by activated T lymphocytes. LCF binds to CD4 and has previously been found to stimulate migration of CD4+ lymphocytes and monocytes. Because human eosinophils, like T cells and monocytes, express CD4, we examined functional responses of eosinophils to LCF. Recombinant LCF (rLCF) expressed in COS cells was purified on a CD4 affinity column. Migration of eosinophils was elicited by rLCF at low concentrations: the 50% effective dose (ED50) was 10(-12) to 10(-11) M, concentrations 100- to 1,000-fold lower than the ED50s for the recognized eosinophil chemoattractants C5a and platelet-activating factor. Two other ligands which bound to CD4, human immunodeficiency virus-1 envelope glycoprotein gp120 and monoclonal antibody OKT4, also stimulated eosinophil migration. Monovalent OKT4 Fab competitively inhibited eosinophil responses to rLCF. rLCF did not influence other functional responses of eosinophils tested, including degranulation, superoxide generation, leukotriene C4 production, in vitro survival, or surface expression of the adherence receptor CR3 (CD11b), human histocompatibility leukocyte antigen DR, or interleukin 2 receptor p55 (CD25). We conclude that CD4 on eosinophils is capable of transducing a migratory stimulus and serves as a receptor for a chemoattractant lymphokine LCF. T cell-derived LCF may contribute to recruitment of eosinophils and CD4+ mononuclear cells concomitantly at inflammatory reactions.

Cold urticaria. Recognition and characterization of a neutrophil chemotactic factor which appears in serum during experimental cold challenge.

Sera were obtained from the venous effluents of cold-challenged arms of patients with idiopathic cold urticaria without plasma or serum cryoproteins; these sera exhibited increased neutrophil chemotactic activity without alterations of the complement system. A two- to fourfold augmentation of the base-line neutrophil chemotactic activity of serum from the immersed extremity began within 1 min, peaked at 2 min, and returned to base-line levels within 15 min, whereas there was no change in the serum chemotactic activity in the control arm. The augmented chemotactic activity in the serum specimens from the challenged arm of each patient appeared in a high molecular-weight region, as assessed by the difference in activity recovered after Sephadex G-200 gel filtration of the paired lesional and control specimens. Sequential purification of this high molecular-weight activity by anion- and cation-exchange chromatography revealed a single peak of activity at both steps. The partially purified material continued to exhibit a high molecular weight, being excluded on Sepharose 4B, and had a neutral isoelectric point. The partially purified material showed a preferential chemotactic activity for neutrophilic polymorphonuclear leukocytes, required a gradient for expression of this function, and exhibited a capacity to deactivate this cell type. This active principle, termed high molecular-weight neutrophil chemotactic factor, exhibited a time-course of release that could be superimposed upon that of histamine and the low molecular-weight eosinophil chemotactic factor and may represent another mast cell-derived mediator.

Structural and functional characterization of the human T lymphocyte receptor for insulin-like growth factor I in vitro.

Growth factor receptors for T lymphocytes, such as interleukin 2 and insulin, are present on activated but not resting T lymphocytes. We sought to determine if insulin-like growth factor I (IGF-I) could act as a growth factor for human T cells and to characterize its receptor on resting and activated cells. Recombinant IGF-I induced two separate functions. It was chemotactic for and increased incorporation of tritiated thymidine into both unactivated (resting) and mitogen-activated T cells. High-affinity 125I-IGF-I binding to human T cells was saturable with an apparent Kd of 1.2 +/- .6 X 10(-10) M for binding to activated T cells and 1.2 +/- .9 X 10(-10) for unactivated T cells. The calculated binding for activated cells was 330 +/- 90 and for resting cells 45 +/- 9 high-affinity receptor sites per cell. Affinity cross-linking of 125I-IGF-I to resting or activated T cells revealed a radioligand-receptor complex of 360,000 mol wt when analyzed by SDS-PAGE without reduction and complexes of 270,000 and 135,000 mol wt upon reduction; prior incubation with excess unlabeled IGF-I prevented formation of the 125I-IGF-I receptor complex. Our data suggest that both resting and activated T lymphocytes bear functional IGF-I receptors similar to those found in other tissues. These receptors may mediate T cell growth and chemotaxis.

Interleukin-16.

Interleukin 16 (IL-16) was initially described in 1982 as the first T cell chemoattractant. Through interaction with CD4, IL-16 has now been characterized as a chemoattractant for a variety of CD4+ immune cells. Recent in vivo studies have more fully characterized IL-16 as an immunomodulatory cytokine that contributes to the regulatory process of CD4+ cell recruitment and activation at sites of inflammation in association with asthma and several autoimmune diseases. Since its cloning in 1994, IL-16 structure and function have been studied extensively. This review addresses the current data regarding IL-16 protein and gene structure; the expanding list of cells capable of generating IL-16; the direct interaction of IL-16 with its receptor, CD4; and the functional bioactivities of IL-16 as they relate to inflammation and HIV-1 infection. In addition, potential therapeutic modalities for IL-16 relating to inflammation and immune reconstitution in HIV-1 infection are also discussed.

Identification of chemoattractant activity for lymphocytes in blister fluid of patients with bullous pemphigoid: evidence for the presence of a lymphokine.

Bullous pemphigoid is characterized by the dermal infiltration of lymphocytes, which precedes the striking influx of eosinophils as the lesion evolves into the bullous phase. This finding prompted a search for chemoattractant activity for lymphocytes in the blister fluid of untreated individuals with bullous pemphigoid. We found such activity in the bullous fluids of 6 consecutive patients but not in a patient with pemphigus vulgaris. This lymphocyte chemoattractant activity separates into 4 peaks upon Sephadex G-100 chromatography and the peak of 56,000 daltons was further evaluated. Upon quaternary aminoethyl Sephadex-anion exchange chromatography this peak elutes at 4-8 ms and with preparative isoelectric focusing it demonstrates an isoelectric point of 8.6-9.0. This activity was susceptible to degradation by trypsin and neuraminidase, but was stable upon heating to 56 degrees C for 30 min. Its chemoattractant activity is predominantly chemokinetic by checkerboard analysis. As defined by chromatography, stability, and functional characteristics, this activity is similar to a recently described human lymphocyte chemoattractant lymphokine. This finding suggests that products of activated lymphocytes are present in blister fluids of patients with bullous pemphigoid and may contribute to the early influx of lymphocytes in this disease.

Interleukin-16.

Interleukin-16 (IL-16) is a pro-inflammatory cytokine. It is synthesized as a precursor molecule that is processed by cleavage of a C-terminal 14 kDa peptide, which aggregates into bioactive tetramers. IL-16 requires the expression of CD4 for its functions, which include induction of chemotaxis, interleukin-2 receptor and HLA-DR expression, reversible inhibition of TcR/CD3-dependent activation and induction of a repressor of HIV-1 transcription. It represents a major source of the lymphocyte chemotactic activity early after antigen challenge of atopic asthmatics in which the major cell of origin is the epithelium, although mast cells, CD8 cells, CD4 cells and eosinophils are also sources; and the presence of IL-16 directly correlates with the number of infiltrating CD4+ T cells. Potential therapeutic applications are use of inhibitors of IL-16 in asthma and for IL-16 in selective CD4+ T cell immune reconstitution in HIV-1 infection or following chemotherapy.

Induction of human monocyte motility by lysyl oxidase.

Lysyl oxidase highly purified from calf aorta was found to be a potent chemotactic agent for unstimulated human peripheral blood mononuclear cells, determined in in vitro assays in Boyden chambers. A typical chemotactic bell-shaped curve was observed, with a maximal migratory response of 237% of control occurring at 10(-10) M lysyl oxidase. The chemotactic response was prevented by prior heat inactivation of the enzyme, by treatment of the enzyme with beta-aminopropionitrile or ethylenediamine, which are active site-directed inhibitors of lysyl oxidase, and by a competing, lysine-containing peptide substrate of lysyl oxidase. The chemoattractant response to lysyl oxidases was characterized by both chemokinetic and chemotactic components. These results raise the possibility that extracellular lysyl oxidase may have important roles to play in biology in addition to its established function in the crosslinking of elastin and collagen.

Signaling and functional properties of interleukin-16.

Interleukin-16 is secreted from a variety of immune cells as a peptide of 17 kDa which aggregates into tetrameric form essential for IL-16s direct interaction with and cross linking of its receptor, the CD4 antigen. IL-16 stimulation of CD4+ cells results in the induction of cell motility, and in addition can function as a competence growth factor for CD4+ lymphocytes. These activities suggest that IL-16 could play a role in the accumulation and activation of CD4+ cells recruited to sites of inflammation. Along those lines, IL-16 has been identified at sites of inflammation associated with several different disease states. Its function as a competence growth factor specifically for CD4+ T cells may be useful for immune reconstitution in immunodeficiency diseases such as AIDS.

Biologic activities of HIV-1 envelope glycoprotein: the effects of crosslinking.

We have examined the biologic activities of native and recombinant preparations of human immunodeficiency virus envelope glycoprotein (gp120), both derived from the HIV-1B strain. Antibody to gp120 was used to evaluate the effects of crosslinking gp120 on signalling by the CD4 receptor. Our results indicate that native and recombinant gp120 produce identical effects in our assay systems. Crosslinking gp120 amplified its chemoattractant activity for lymphocytes and monocytes and increased the peak intracellular calcium level, compared with binding of gp120 alone. The induction of inositol trisphosphate (IP3) production, induction of interleukin 2 receptors (IL2R), and inhibition of lymphocyte proliferation following treatment with gp120 were not enhanced by the addition of crosslinking antibody.

Identification of rat mast cell--derived chemoattractant factors for lymphocytes.

Rat peritoneal and pleural mast cells have been demonstrated to release their granule-associated mediators, including chemotactic factors for eosinophilic polymorphonuclear leukocytes, after challenge with rabbit anti-rat F(ab')2 antisera in vitro. The presence of increased numbers of lymphocytes as well as eosinophils 6 to 24 hr after an initial immediate hypersensitivity reaction in the skin led to the present studies, which demonstrate the immunologic release of three chemoattractant factors for lymphocytes. One factor is chemotactic for nylon wool nonadherent rat splenic lymphocytes, a second factor is chemotactic for adherent rat splenic lymphocytes, and the third factor is chemokinetic for both nonadherent and adherent rat splenic lymphocytes. In addition, a high-molecular-weight inhibitor of lymphocyte migration was identified. Immunologic challenge of rat mast cells in vitro results in the release of lymphotactic activity, suggesting a possible explanation for the appearance of a lymphocytic infiltrate in the late phase of the immediate hypersensitivity reaction.

Chemotactic activity of porcine insulin for human T lymphocytes in vitro.

T lymphocytes bear insulin receptors only after activation and entry into the cell cycle. To determine whether cell motility is concomitant with growth factor action in T lymphocytes, we measured the chemotactic activity of porcine insulin (10(-11) to 10(-5) M) for T lymphocytes. We found that the chemotactic response of human T cells activated with phytohemagglutinin (PHA) to porcine insulin was increased over that of resting T cells, with a concomitant two log leftward shift in the dose response. CD4+ and CD8+ subsets responded identically. Checkerboard analysis showed insulin to be chemotactic, as well as chemokinetic. The nature and time course of acquisition of the dose-response shift suggest that chemotaxis may be signaled by insulin acting on high affinity insulin receptors. The chemotactic effect of insulin exemplifies the general chemotactic effect of growth factors for motile target cells, and may be a useful model for the study of chemotactic signaling in T lymphocytes.

Modulation of lymphocyte migration by human lymphokines. III. Characterization of a lymphocyte migration inhibitory factor (LyMIF35K).

The lymphokine that augments the migration of nonsensitized T lymphocytes (LCF) has been observed to be predominantly a chemokinetic factor, suggesting that separate lymphocyte migration inhibitory lymphokine(s) might exist. Utilizing a modified Boyden chamber assay, lymphocyte migration inhibitory activity was identified in the culture supernatants of human nylon wool-nonadherent blood mononuclear cells stimulated with concanavalin A in vitro for 48 hr. Sephadex G-100 gel filtration chromatography of these culture supernatants was shown to contain two regions of noncytotoxic migration inhibitory activity for nonsensitized human blood lymphocytes and rat splenic lymphocytes. The 30-40,000 dalton inhibitory activity was further characterized and noted to be cationic by ion-exchange chromatography and isoelectric focusing (pI = 8.6). Its biologic activity was sensitive to neuraminidase and to heat treatment but not to trypsin. The migration inhibitory activity of this factor (LyMIF35K) was directly proportional to its ability to increase lymphocyte adherence.

Effect of ambient oxygen on cultured endothelial cells from different vascular beds.

By the use of production of neutrophil chemoattractant activity as a marker, we investigated the responsiveness of endothelial cells of four different anatomic origins to altered ambient oxygen tension to determine whether genetic or conditioned variation existed. The ability of bovine aortic, bovine pulmonary arterial, bovine coronary arterial, and human umbilical vein endothelial cells incubated in decreased oxygen concentrations to release neutrophil chemoattractant activity was assessed. Bovine aortic, human umbilical vein, and bovine coronary arterial endothelial cells produce neutrophil chemoattractant activity in response to 10 or 3% ambient oxygen in vitro. In contrast, 0% ambient oxygen is required for appearance of neutrophil chemoattractant activity from pulmonary artery endothelial cells. These studies suggest that there may be genetic or conditioned variations in the response of endothelium from different vascular beds to decreased oxygen tensions. Furthermore, endothelial cells may play a role in neutrophil-mediated tissue injury during ischemia.

Opsonin-independent phagocytosis by human alveolar macrophages: augmentation by human plasma fibronectin.

A trypsin-sensitive membrane recognition unit that mediates phagocytosis of particulate activators of the human alternative complement pathway is present on human alveolar macrophages. Fragmented human plasma fibronectin selected by affinity chromatography with a monoclonal antifibronectin antibody augments this capacity. These data suggest a nonimmune mechanism for the clearance of some microorganisms from the opsonin-deficient microenvironment of the lung in which the alveolar macrophage is the principal resident phagocyte.