Lymphokine research最新文献

Studies are reviewed in which the role of IFN-gamma in different models of inflammation in mice is examined: LPS-induced generalized Shwartzman reaction, experimental allergic encephalomyelitis (EAE) and experimental cerebral malaria (ECM). The particular role of the cytokine was studied by systemic administration and by blocking the endogenously produced cytokine by the use of neutralizing antibodies. IFN-gamma was found, depending on the model and circumstances, to exert an anti- or a pro-inflammatory effect. In the generalized Shwartzman reaction and ECM this cytokine has a disease promoting role. In EAE, on the contrary, endogenous as well as exogenous IFN-gamma exert a down-regulating effect.

In order to define functional domains involved in the control of TNF-gene transcription, 5'-flanking sequences of the TNF-gene were analysed by TNF-promoter deletion mutants linked to the CAT-gene and by gel retardation assays. To this, three TNF-promoter fragments of different length were fused to the CAT reporter gene and transiently transfected into several human cell lines. We found that a 315 bp long fragment encompassing positions -285 bp to +30 bp with respect to the TNF mRNA cap site is sufficient to function as a PMA inducible promoter/enhancer in all cell lines tested. By further deletion analysis of this clone we could narrow the PMA-inducible element within a 185 bp long sequence (-285 to -110 bp). In addition, we investigated specific interactions between nuclear proteins and TNF promoter sequences using gel-retardation assays. Besides several constitutive binding activities, we could demonstrate in several cell lines a nuclear protein that is induced by PMA and binds to the fragment of the TNF promoter containing the PMA-inducible element.

We have investigated changes in serum interleukin 6 (IL-6) in patients undergoing elective cholecystectomy. Serum IL-6 increased in all patients within 1.5 hour of incision, reaching a maximum between 1.5-4 hours after incision (median 50 U/ml; range 22-79 U/ml). The maximum serum IL-6 correlated with the length of the operation (r = 0.95). Serum C-reactive protein was not detectable until 8-12 hours post-incision, but maximum serum C-reactive protein did not correlate with maximum serum IL-6 concentration or length of operation. There was no consistent increase in plasma interleukin 1 or tumour necrosis factor following surgery. Serum IL-6 is an early marker of tissue damage and may be of value in the study of the metabolic response to injury.

Bone marrow-derived macrophages were used to study the mechanism of action of recombinant gamma interferon associated with multilamellar phospholipid liposomes. Gamma interferon associated with liposomes caused an inhibition of [3H]-thymidine uptake induced by macrophage colony-stimulating factor (CSF-1), and primed macrophages for subsequent induction of tumoricidal activity by bacterial lipopolysaccharide (LPS). The liposomes were equally active whether the gamma interferon was added before, or after vesicle formation. The result suggested that significant biologically active gamma interferon was bound to the outside of the vesicles. Interferon binding to liposomes was confirmed using radiolabelled ligand. The liposomes themselves were found to be biologically active in promoting proliferation and in acting synergistically to prime cytotoxicity. Vesicles that contained both phosphatidylethanolamine and phosphatidyl-serine or succinylated phosphatidylethanolamine were most active. Such vesicles were found to be internalised rapidly by bone marrow-derived macrophages. Thus, encapsulation of ligand, and internalisation into cytoplasm, do not appear to be involved in the action of liposome-associated gamma interferon. On the other hand, the liposomes may contribute in other ways to improving the therapeutic potential of gamma interferon.

The present study examines the effects of IL-4 and TNF-alpha on the CD3-dependent (Ag/MHC-initiated or anti-CD3 mAb-initiated) and CD3-independent (IL-2-initiated) pathways of the initiation of human T-cell activation. Both IL-4 and TNF-alpha significantly augmented the CD3-dependent T-cell proliferation induced by either irradiated OKT3 hybridoma cells or allogeneic B cells. In contrast, the CD3-independent IL-2-initiated T-cell proliferation was enhanced by TNF-alpha and significantly inhibited by IL-4. Although the growth-enhancing effects of both IL-4 and TNF-alpha on the CD3-dependent T-cell proliferation were noticeable regardless of when these cytokines were introduced in culture, the inhibitory effect of IL-4 on the CD3-independent IL-2-initiated T-cell activation was observed only if IL-4 was added at the initiation but not later than 24 hr of "T cells + IL-2" cultures. The growth-enhancing effects of both IL-4 and TNF-alpha on the CD3-dependent T-cell activation were not confined to any one subset of T cells. On the other hand, IL-4 inhibited the IL-2-induced proliferation of CD4+ (helper/inducer) T cells and CD45R+ (virgin) T cells but not that of CD8+ (cytotoxic/suppressor) T cells and CD45R (memory) T cells. When examined for their effects on cytokine production, CD3-dependent production of IL-2 and IFN-gamma was affected by neither cytokine, whereas IL-4 strongly inhibited the production of IFN-gamma by IL-2-stimulated T cells. Consistent with their enhancing and inhibitory effects, respectively, on IL-2-induced T-cell proliferation, TNF-alpha augmented and IL-4 inhibited the development of IL-2-stimulated MHC-unrestricted cytolytic (MUC) T-cell activity directed against tumor cells. When deprived of IL-2, MUC T cells rapidly lose their cytolytic activity, and despite its inhibitory effect on the development of MUC T cells, exposure of IL-2-deprived MUC T cells with decaying cytolytic activity to IL-4 retards the decay in their cytolytic activity. These results suggest the differential regulatory effects of IL-4 and TNF-alpha during human T-cell activation.

Cells of the proerythromyeloid cell line K562 and of the T cell line Jurkat were able to induce an increase in TNF-, IL1 alpha-, and IL1 beta-mRNA expression in human peripheral blood derived monocytes in vitro. The activating principle of these tumor cells was associated with fractions of membrane preparations of distinct molecular mass, 32-38 kD for Jurkat cells and 46-54 kD for K562 cells, respectively. At least part of the activating constituent seemed to be protein in nature. Isolated membrane preparations of both cell types induced production and secretion of TNF. Stimulation of monocytes with the viable tumor cells led to TNF release only when Jurkat cells were used. Viable K562 cells induced enhanced TNFmRNA expression but seemed to absorb soluble TNF from the supernatant.

Antigen presentation by neonatal murine spleen cells and the production of lymphokines and interleukins involved in the stimulation of a T-helper-2 (TH2) cell line (D10-G4.1) were studied as were the effects of ultra violet (UV)-irradiation on this system. Neonatal spleen cells are less capable than adult cells of performing the initial steps of the immune response required for antigen dependent activation of TH2 cells. These steps include soluble antigen processing and presentation and as a result reduced production of IL-4 and IL-1-Inducer Factor ("IL-1-IF") by the T-helper cells and reduced production of IL-1 and IL-2 by the antigen presenting cell population. Spontaneous membrane IL-1 activity is low in the neonate, however, when exposed to "IL-1-IF" they can express adult levels. Ultra-violet (UV) irradiation of the antigen presenting population has a damaging effect on all the above mentioned processes. Antigen processing and presentation, induction of D10 IL-4 production and proliferation, and IL-2 production demonstrate two different age related patterns of UV-irradiation induced damage: a dose dependent inhibition when adult cells are irradiated and an inverse effect in which low doses of irradiation were more inhibitory than higher doses when neonatal cells are irradiated. However, the secretion and membrane expression of IL-1 by both age groups are directly and totally inhibited by the range of UV-irradiation doses used and cannot be reinduced with a supplement of a crude "IL-1-IF". While the capacity to produced IL-1 is totally destroyed by UV-irradiation, the ability to produce IL-2 remains intact and remains responsive to an "IL-2-Inducer" activity during proper antigen presentation. The low responses of neonatal antigen presenting spleen cell populations and the damaging effect of UV on both neonatal and adult responses are not due to the induction of suppressor factors.

Five Monoclonal Antibodies (MAbs) coded respectively #111, 122, 206, 209 and 609 were produced against human recombinant Interleukin-1 beta (rIL-1 beta, amino acids 117-269). Four of these MAbs (#111, 122, 206 and 609) have been identified able to inhibit the biological activity of recombinant and natural Interleukin-1 beta. Competition studies suggested that three non-overlapping epitopes of the molecule were recognized by the MAbs. MAbs #609 and 206 have been selected on the basis of high affinity and used together in a two-site ELISA to detect IL-1 beta. The sensitivity of this ELISA was 1 pg/well (10-20 pg/ml). The assay did not detect rHuIL-1 alpha, rHuIL-2, rHuTNF-alpha, rHuIFN-gamma, rHuIL-6, and natural a- and b-FGF. The immunoassay was then compared to current assay such as co-mitogenic effect on murine thymocytes for detection of IL-1 beta in the intra- and extracellular compartments of IFN-gamma and/or LPS-stimulated human blood monocytes. We confirm that IFN-gamma potentiates IL-1 beta secretion in response to LPS (even with high dose 1 microgram/ml) but not the intracellular precursor of IL-1 beta. This immunoassay could be used also to detect IL-1 beta in plasma, sera and synovial fluids.