Interleukin 1 receptor antagonist inhibits the augmentation of metastasis induced by interleukin 1 or lipopolysaccharide in a human melanoma/nude mouse system.

This study examined the ability of the recombinant human interleukin 1 receptor antagonist (IL-1ra) to block interleukin 1 (IL-1)-mediated experimental metastases from the A375M human melanoma. In vivo, IL-1ra administrated at concentrations > or = 200 times IL-1 significantly inhibited the increase in lung colonies induced by IL-1 in nude mice. The response to IL-1 was significantly inhibited when IL-1ra was administered simultaneously with or 1 to 3 h before IL-1. In vitro, the incubation of IL-1-activated endothelial cells with IL-1ra prevented the increase in adhesion of A375M melanoma cells. At the same experimental conditions, IL-1ra inhibited the augmented expression of the intracellular and vascular cell adhesion molecules 1 and E-selectin induced by IL-1 on endothelial cells. Lipopolysaccharide, an IL-1 inducer, increased the number of lung colonies in nude mice. IL-1ra injected with or 1 h after lipopolysaccharide inhibited this augmentation, suggesting a role for host-produced IL-1 in metastasis formation.

Murine endothelioma cell lines transformed by polyoma middle T oncogene as target for and producers of cytokines.

We studied cytokine-related functional properties of four mouse endotheliomas from different anatomical sites obtained by transformation with middle T oncogene. We examined mRNA expression of IL-6, IL-1 alpha, macrophage-CSF, granulocyte/macrophage-CSF, and two members of an emerging super-family of chemotactic cytokines (JE/monocyte chemoattractant protein-1 (MCP-1) and KC). Exposure to IL-1 augmented or induced cytokine gene transcripts in three endothelioma lines (eEnd.1, sEnd.1, and tEnd) with maximal expression in tEnd.1 cells. Endothelioma cells also responded to TNF-alpha and LPS. Levels of IL-6 and monocyte chemotactic activity (a JE/MCP activity) correlated with mRNA expression. IL-1 also induced production of procoagulant activity and platelet-activating factor in endothelioma cells, with heterogeneity in the levels of response among individuals lines. Murine melanoma B16-F1, human colon carcinoma HT29 cells, CB33MT lymphoblastoid cells, and monocytes adhered to endothelioma monolayers and the adhesive properties of these cell lines were modulated by IL-1 beta, with marked differences among themselves. Murine EC derived from brain capillaries, used as control, shared several properties with bEnd.4 line. Endothelioma lines cause tumors by recruiting host cells. The capacity to produce cytokines that directly or indirectly attract host vascular cells, may play an important role in hemangioma induction in vivo. Murine endothelioma lines, generated by transformation with the polyoma middle T oncogene, retain functional properties of normal endothelium, and may represent an invaluable tool for analysis of the immunobiology and heterogeneity of EC in different tissues.

Expression of monocyte chemotactic protein and interleukin-8 by cytokine-activated human vascular smooth muscle cells.

The present study was designed to investigate the capacity of human vascular smooth muscle cells (SMCs) to produce a cytokine chemotactic for monocytes (monocyte chemotactic protein [MCP]) and by way of comparison, a related polypeptide activator of neutrophils (known as interleukin-8 [IL-8] or neutrophil activating protein-1 [NAP-1]. On exposure to IL-1, SMCs released high levels of chemotactic activity for monocytes, which could be removed by absorption with anti-MCP antibodies. MCP production by activated SMCs was comparable to that of IL-1-stimulated umbilical vein endothelial cells. Activated SMCs released appreciable levels of IL-8, as determined by a specific enzyme-linked immunosorbent assay, but little chemotactic activity for neutrophils. IL-1-treated SMCs expressed high levels of both MCP and IL-8 mRNA transcripts, as assessed by Northern blot analysis. Tumor necrosis factor and bacterial lipopolysaccharide but not IL-6 also induced MCP and IL-8 gene expression in SMCs. Nuclear runoff analysis revealed that IL-1 augmented transcription of the MCP and IL-8 genes. The capacity of SMCs to produce a cytokine (MCP) that recruits and activates circulating mononuclear phagocytes may be of considerable importance in the pathogenesis of vascular diseases (e.g., vasculitis and atherosclerosis) that are characterized by monocyte infiltration of the vessel wall.

Comparative studies on the anorectic activity of d-fenfluramine in mice, rats, and guinea pigs.

The present study compares the anorectic activity of d-fenfluramine and its metabolite d-norfenfluramine in three animal species. d-Fenfluramine and d-norfenfluramine show anorectic activity at increasing doses (ED50) in rats, guinea pigs, and mice, d-norfenfluramine being more active than d-fenfluramine in all three species. Equiactive anorectic activities are reached with different brain levels of d-fenfluramine and d-norfenfluramine, guinea pigs being the most sensitive species, followed by rats then mice. The metabolite most probably plays a major role in the anorectic effect of d-fenfluramine in guinea pigs, contributes to the anorectic activity in rats, but adds little to the action of the parent drug in mice. The different sensitivity to d-fenfluramine and d-norfenfluramine in these three species does not appear to be explained by a number of biochemical parameters, including serotonin uptake or release, receptor subtypes, or 3H-d-fenfluramine binding and uptake.

Role of 5-HT receptors in the effect of d-fenfluramine on gastric emptying and feeding behaviour as examined in the runway test.

In one experiment, the effect of d-fenfluramine (DF) on gastric emptying was studied in rats treated i.p. with metergoline, a non-selective serotonin (5-HT) receptor antagonist, ritanserin, a selective 5-HT2 and 5-HT1C receptor antagonist, and xylamidine, a 5-HT antagonist which has poor access to the brain. Metergoline (1 mg/kg) but not ritanserin (0.5 mg/kg) or xylamidine (3 mg/kg) blocked the effect of 2.5 mg/kg DF studied 2 and 4 h after injection. In a second experiment, we studied the ability of metergoline to antagonise the effect of DF, administered after a meal, on runway performance, food intake and gastric emptying assessed 4 h later. Metergoline at a dose of 1 mg/kg did not antagonise the effect of DF (2.5 mg/kg) on runway performance but completely blocked the effect on gastric emptying. The data clearly show that DF delays gastric emptying by indirectly activating 5-HT1 receptors; this effect is not important for the ability of DF to reduce runway performance and food intake when the drug is injected after a pre-feeding period. While there is evidence that DF hastens the termination of the meal by a 5-HT mechanism, the data suggest that DF may prolong the satiating effect of food during the post-absorptive phase by mechanisms other than 5-HT.

Cytotoxicity of activated monocytes on endothelial cells.

Unstimulated human monocytes did not express appreciable levels of cytotoxicity on normal human umbilical vein endothelial cells (EC) in a 24-48 hr TdR release assay. On activation with IFN-gamma and LPS, monocytes had appreciable cytotoxicity on EC. Monocyte cytotoxicity on EC was not dependent on the presence of contaminating lymphoid cells. Recombinant TNF, IL-1, and IL-6 as well as monocyte supernatants did not exert a cytotoxic effect on EC. Moreover, anti-TNF, anti-IL-1, and anti-IL-6 antibodies, as well as scavengers of reactive oxygen intermediates, did not affect the cytotoxicity of activated monocytes on EC. Antibodies against the beta-chain (CD18) of leukocyte integrins inhibited the adhesion and cytotoxicity of activated monocytes on EC. Pretreatment of EC with IL-1 augmented the adhesion of monocytes on EC. Normal monocytes were not cytotoxic on IL-1-pretreated EC and IL-1 treatment did not increase the susceptibility of EC to activated monocytes. Thus adhesion is necessary but not sufficient for monocyte killing of EC. Anti-alpha L (LFA-1) antibodies markedly reduced monocyte cytotoxicity on EC, although anti-alpha X (p150) antibodies had only a modest effect. Anti-alpha M (Mac-1/CR3) antibodies were intermediate inhibitors of EC killing by activated monocytes. Thus, alpha L, beta 2 (LFA-1), and, to a lesser extent, alpha M, beta 2 (Mac-1/CR3) and alpha X, beta 2 (p 150, 95) integrins are the main adhesive structures involved in the cytotoxic interaction of activated monocytes with EC. Monocyte-mediated damage of EC could play a role as a mechanism of tissue injury under conditions of local or systemic activation of mononuclear phagocytes.