Mitsuomi Hirashima

Chemotactic factors associated with leukocyte emigration in immune tissue injury: their separation, characterization, and functional specificity.

Publisher Summary This chapter discusses the separation, characterization, and functional specificity of a number of chemotactic factors, which may satisfy many criteria making them acceptable as inflammatory leukocyte chemotactic factors. They are separated from the sites of inflammatory lesions resulting from immune responses as experimental models. The chapter focuses on three neutrophil chemotactic factors, three macrophage chemotactic factors (MCF-a, b, and c), four lymphocyte chemotactic factors (LCF-a, b, c, and d), and three eosinophil chemotactic factors. It is of special importance that highly purified chemotactic factors among them exhibited cell-type–specific functions for the associated leukocytes in vitro , as well as in vivo. The intradermal injection of these purified chemotactic factors clearly provoked adherence of the associated leukocytes to the vascular endothelium of the venules, followed by selective emigration of the cells into the perivascular locations. Cinemicrophotographic and ultrastructural observations on neutrophil emigration induced by leukoegresin have given one direction to the analysis of morphological sequence in the emigration of other leukocytes in vivo .

A macrophage chemotactic factor sharing common antigenicity with immunoglobulin G from DNP-ascaris extract-induced skin lesion in guinea-pig

SummaryA macrophage chemotactic factor (factora) was extracted from DNP-ascaris extract-induced skin lesions (showing a maximal macrophage reaction) in guinea-pigs and then purified by gel filtration on Sephadex G-100 and chromatography using DEAE-Sephadex and CM-Sephadex. This material was a thermolabile protein (free of nucleic acid) with a molecular weight of about 150,000. It shared common antigenicity with serum IgG, and its chemotactic activity was completely absorbed by anti-IgG and antilight chain antibodies, but not by anti-BSA antibody. In contrast to leucoegresin, this material seemed active for macrophages but not for neutrophils. Less active factorsb andc were thermostable and not absorbed by the antibodies described above. The ratio of chemotactic activity of factorsa,b andc in the skin extracts was roughly 60:25:15.

Induction of an eosinophil chemotactic factor production from T lymphocytes by a B cell lymphoma line

Production of an eosinophil chemotactic factor (ECF) from human mononuclear leukocytes (MNL) was induced by coculture with an irradiated B cell lymphoma line, BALL-1. BALL-1 induced ECF production from OKT4-positive T lymphocytes without evident IL-2 production. Treatment of MNL with anti-IL-2 antibody failed to suppress the BALL-1-induced ECF production, whereas the treatment strongly inhibited IL-2-induced ECF production. Control supernatants of BALL-1 cells alone did not induce ECF production. BALL-1 fixed with periodate-lysine-paraformaldehyde, but not acetone or ethanol, induced evident ECF production. The isoelectric point of BALL-1-induced ECF (m.w. 10-30 kD) was around pI 7, whereas that of the IL-2-induced ECF was around pI 5. A combination of monoclonal antibodies against IL-3, IL-5, and GM-CSF suppressed the activity of the IL-2-induced ECF but not that of the BALL-1-induced ECF. BALL-1-induced ECF suppressed a respiratory burst from an eosinophilic cell line (YY-1) induced by N-formyl-L-methionyl-L-leucyl-L-phenylalanine, whereas the IL-2-induced ECF did not, suggesting that the biological function of these two ECF is different, at least in the effect on respiratory burst of eosinophils. From the present results we propose that one reason for infiltration of eosinophils into the stroma of tumors is that some tumor cells can stimulate OKT4-positive T lymphocytes to produce an ECF, and that eosinophils attracted by this ECF exhibit biological functions which are different from those of eosinophils attracted by other ECF.

Selective regulation of chemotactic lymphokine production by monocytes and macrophage cell line cells.

Effects of lipopolysaccharides (LPS) on the production of chemotactic lymphokines for eosinophils and monocytes (ECF and MCF) from antigen- or mitogen-stimulated T cells were examined. Supernatants from monocytes stimulated with various LPS regulated concanavalin A (Con A)- and purified protein derivative (PPD)-induced ECF or MCF production. The regulation varied with LPS used for monocyte stimulation. The supernatant from monocytes stimulated with LPS from Escherichia coli, J-5 strain, selectively potentiated ECF production, whereas that from Salmonella minesota potentiated MCF production. In contrast, supernatant from monocytes stimulated with LPS from Salmonella typhimurium potentiated both ECF and MCF production, whereas that from Vibrio cholerae failed to potentiate production. The supernatants from monocytes stimulated with lipid A of S. typhimurium, S. minesota, and E. coli, however, failed to potentiate ECF and MCF production. The potentiating activity for each lymphokine was recovered from fractions ranging in molecular weight between 10 and 20 kD. Further purification with isoelectric electrophoresis revealed that the potentiating activity for the production of chemotactic lymphokine for eosinophils has a pI value of about 4-5, and that the activity for MCF production is detected in two fractions of pI about 5-6, and 7-8. Macrophage lineage cell line cells, such as THP-1 and U-937, also release similar factors after differential stimulation.

Production of an eosinophil chemotactic lymphokine by a monocyte-derived factor from patients with hypereosinophilia.

Peripheral OKT4‐positive T lymphocytes from patients with hypereosinophilia spontaneously and selectively produced an eosinophil chemotactic factor (ECF) with chemokinetic activity. The molecular weight of the ECF was about 45,000 to 70,000. A possible mechanism of its spontaneous production by T lymphocytes was analyzed. Culture supernatants of blood monocytes from the patients showed little or no ECF activity, but they had a potency to induce the ECF production from T lymphocytes from normal donors when the cells were stimulated by the supernatants, which suggests that a monocyte‐derived soluble factor (MDF) stimulated T lymphocytes to produce an ECF resembling this spontaneously produced ECF from the patients. MDF seemed to be a synthesized protein by the cells. Gel filtration indicated that molecular weight of MDF ranged between 70,000 and 100,000. MDF activity was stable at 56° for 30 min but labile at 80° for 30 min. MDF failed to show detectable IL‐1 and IL‐2 activity. Furthermore, supernatants of stimulated monocytes by lipopolysacchride or silica particles failed to show ECF‐producing activity, whereas they showed evident lymphocyte‐activation activity. Neither recombinant IL‐1 nor IL‐2 had ECF and ECF‐producing activity. From the present experiments, it was suggested that MDF was at least partly involved in the induction of ECF production by OKT4‐positive T lymphocytes in patients with hypereosinophilia.

A C5-Derived Macrophage Chemotactic Factor from DNP-Ascaris Extract-Induced Skin Lesion in Guinea Pigs

In complement-depleted condition induced by cobra venom factor (CoF), tissue macrophage reaction and macrophage chemotactic activity (MCA) in skin extract from skin lesions induced by DNP-Ascaris extract (DNP-As) were decreased, respectively. MCA in the fraction containing macrophage chemotactic factor b (B fraction) was decreased completely. While MCA in the fraction containing macrophage chemotactic factor a and c (A fraction) was affected only slightly. These results suggested that the decrease of MCA of B-fraction might be associated with depletion of complement system by CoF. Using anti-C5 antibody, it was confirmed that macrophage chemotactic factor-b (MCF-b) (MW 14,000: thermostable) might be derived from C5 fragment in vivo.

Production of fibroblast proliferative cytokines from T lymphocytes stimulated by a B cell lymphoma line and their functional heterogeneity

Human mononuclear leukocytes (MNL) produced several factors with fibroblast proliferation activity (FPA) for HFL-1, a human lung fibroblast cell line, when MNL were cocultured with irradiated BALL-1, a B cell lymphoma line (BCLL), but not with other BCLL. The cellular source of BALL-1-induced FPA seemed to be CD4-positive T lymphocytes. On isoelectric electrophoresis, major activity of BALL-1-induced FPA was detected in the fractions around pH 4-5, and minor activity was present in the fractions around pH 6-7. Major BALL-1-induced FPA consisted of at least 4 different fibroblast proliferation factors (FPFs) according to their molecular weight; 320-600 kDa (P-I), 50-110 kDa (P-II), 22-38 kDa (P-III) and 4.6-11 kDa (P-IV). P-I had affinity to heparin though the rest had little or no affinity. FPA of P-I was suppressed by an antibody against acidic FGF, and FPA of P-III was suppressed by an antibody against IL-6. On the other hand, FPA of P-II and P-IV was suppressed by none of the antibodies against cytokines with FPA, such as FGF, IL-4, IL-6, IFN-gamma, TGF-beta and TNF-alpha. It was thus suggested that P-I was acidic FGF, that P-III was IL-6, and that P-II and P-IV were different cytokines from those described above. Furthermore, it was found that P-II and P-IV failed to exhibit proliferation activity for human umbilical vein endothelial cells (HUVEC).(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of eosinophilia in rats infected with Nippostrongylus brasiliensis. I: Eosinophil chemotactic factor produced spontaneously by mesenteric lymph node cells of infected rats

The possible mechanism of eosinophilia was studied in rats undergoing primary infection with Nippostrongylus brasiliensis (Nb). In vivo studies showed that the kinetics of intestinal tissue eosinophilia was not directly related to those of the intestinal worm burden. Furthermore, Nb worm extract has no or only very weak in vitro eosinophil chemotactic activity, suggesting that parasite-derived eosinophil chemotactic factor (ECF) is, if at all, not a major regulator for intestinal tissue eosinophilia in this Nb rat system. On the other hand, when mesenteric lymph node (MLN) cells obtained various days after infection were cultured, potent ECF activity was detected in the cell-free supernatant from the cultures of MLN cells 15-20 days after infection, at which time marked intestinal tissue eosinophilia was observed in vivo. Production of ECF by MLN cells from Nb-infected rats seems to be spontaneous, since these cultures were performed without adding worm antigen. ECF-producing activity of day-20 MLN cells was suppressed by adding various metabolic inhibitors such as cycloheximide, mitomycin C, or puromycin. After Sephadex G-75 gel filtration, ECF activity produced by day-20 MLN cells was associated with two different molecules.

The mediation of macrophage reaction in inflammation, with special reference to IgG-derived chemotactic factor.

Three different macrophage chemotactic factors were separated from hypersensitivity skin lesions induced by DNP‐ascaris extract or PPD. The amount of each factor seemed to be changeable according to the nature of inflammatory stimuli. The chemotactic activity of the factor a was completely absorbed by anti‐IgG and anti‐light chain antibodies, while that of the factors b and c was not absorbed by the antibodies. The factor a was produced from IgG by neutrophil neutral serine protease resembling elastase but fatling to digest elastin‐orcein; its chemotactic generation was accompanied by release of dialysable peptide(s) from the IgG molecule, suggesting a minor structural change of the IgG molecule. Production of leucoegresin or lymphocyte chemotactic factor by europhile neutral thiol protease from IgG molecule was discussed.

Effects of Eosinophilotropic Cytokines on Differentiation of an Eosinophil Cell Line, YY-1

Differentiation of an eosinophilic cell line, YY-1, was induced by treatment with 0.3 mM butyric acid (BA). The effects of eosinophilotropic cytokines, such as IL-3, IL-5, and GM-CSF on BA-induced differentiation, especially in the chemotactic response to ECF, was examined. Five STO-2-derived ECFs, IL-5, C5a, and fMLP were used as eosinophil chemoattractants. The effects of the cytokines on the chemotactic response of YY-1 greatly differed according to the ECF used. It was thus suggested that heterogeneity in the chemotactic response of eosinophils depends on the cytokines which act on eosinophils during differentiation.