Beate M. Czarnetzki

Pharmacological modulation of IL‐6 and IL‐8 secretion by the H1‐antagonist decarboethoxy‐loratadine and dexamethasone by human mast and basophilic cell lines

Abstract Mast cells and basophils are central effector cells of allergic reactions and are involved in inflammatory diseases. These cell types produce an array of mediators including a broad spectrum of cytokines. In order to examine whether antiallergic drugs modulate the release of these mediators, we have investigated the influence of dexamethasone and decarboethoxy‐loratadine (DEL), the active metabolite of the H1‐blocking agent loratadine, on the release of IL‐6 and IL‐8 by the human mast cell line HMC‐1 and the human basophilic cell line KU812 by ELISA. Dexamethasone (10−6‐10−11 M) or Del (10−5‐10−14 M) were added to the cells either 1 h prior to or simultaneously with PMA and Ca‐ionophore A23187. When preincubated with the cells, DEL dose‐dependently suppressed IL‐6 release by up to 40% and IL‐8 release by up to 50%. Dexamethasone potently suppressed secretion of both cytokines if simultaneously added to the cells with the stimuli by up to 60% and after preincubalion by up to 80%. Since both antihistamines and glucocorticoids are used for treatment of allergic diseases, the findings reported here indicate that these drugs may modulate allergic reactions via inhibition of cytokine release from mast cells and basophils.

Effect of CC chemokines on mediator release from human skin mast cells and basophils.

Chemokines are considered important mediators of various inflammatory processes. In human basophils, different CC chemokines are known to stimulate release of histamine and generation of leukotriene (LT)C4. In the present study, we have evaluated the effect of RANTES (regulated upon activation, normal T cell expressed and secreted), monocyte chemotactic protein (MCP)-1, MCP-2, MCP-3, macrophage inflammatory protein (MIP)-1 alpha and MIP-1 beta on mast cell activation. Whereas all these CC chemokines caused dose-dependent release of histamine from basophils in mixed human leukocyte suspensions, none of them was able to induce release of histamine as well as tryptase or prostaglandin (PG)D2 from human skin mast cells, nor did priming with these substances enhance IgE-mediated mediator release. In addition, all chemokines failed to promote changes in the cytosolic free calcium level in the human mast cell line HMC-1. These results add further evidence for the differences between human mast cells and basophils regarding cytokine-dependent activation.

Glucocorticoid-induced modulation of cytokine secretion from normal and leukemic human myelomonocytic cells.

Since glucocorticoid effects on inflammatory processes may be mediated via modulation of cytokine release, different types of myelomonocytic cells were stimulated in vitro with lipopolysaccharide (50 ng/ml) or phorbol myristate acetate (25 ng/ml) plus the ionophore A23187, 2 x 10(-7) M, and release of interleukin (IL)-1 beta, IL-8 and tumor necrosis factor (TNF)-alpha was measured after 24 h by ELISA. Peripheral blood mononuclear cells from two allergic and two normal human donors released similarly large quantities of IL-8 and lower amounts of IL-1 beta and TNF-alpha. This also held for myelomonocytic cell lines, with THP-1 cells being most active, followed by U-937 and HL-60 cells. All potent glucocorticoids studied caused a dose-dependent inhibition of cytokine release from donor cells, being most marked for IL-1 beta and lowest for IL-8. Inhibition of cytokine release was also noted with U-937 cells, with clear differences in potency between the glucocorticoids, whereas release was enhanced in all experiments with THP-1 cells. These results were confirmed with Northern blot analysis. Modulating effects of glucocorticoids on cytokine release are thus complex, and are particularly dependent on the cell type studied.

Histamine releasability of basophils and skin mast cells in chronic urticaria

In order to clarify the pathogenetic role of basophils and mast cells in chronic urticaria, histamine and leukotriene (LT)C4 release was examined in washed mixed leukocytes (n= 8) and skin mast cells (n= 5) from patients with chronic urticaria and compared with the same cells from normal controls (n= 9). Anti‐IgE‐stimulated basophil histamine release was significantly reduced in urticaria patients (median 2.9%vs 15.1% in normal controls), whereas histamine release to A23187. FMLP, and PAF, as well as anti‐IgE‐induced LTC4 release, showed no differences in both groups. In contrast, anti‐IgE‐stimulated skin mast cells from urticaria patients reacted similarly to those of controls (median histamine release 11.4%vs 14.2% in normal controls). Pretreatment of the cells with interleukin (IL)‐3 upregulated responsiveness of basophil histamine release to anti‐IgE in urticaria patients (median histamine release 14.3%), but pretreatment with the H2‐antagonist cimetidine showed no effect. These data show that reduced basophil histamine releasability in chronic urticaria is not H2 mediated. It is a stimulus, mediator‐, and cell type‐restricted phenomenon that can, at least partially, be reversed in the presence of the cytokine IL‐3.

MHC class II antigen expression is increased in different forms of urticaria.

Urticarial reactions encompass a variety of inflammatory and immunological reactions. In order to clarify specific aspects of these processes, we analyzed the distribution and sequential expression of major histocompatibility complex II (MHC class II) molecules in tissue sections from different types of whealing reactions. Using immunohistochemical techniques and monoclonal antibodies, expression of HLA-DR, HLA-DP, and HLA-DQ was examined on resident and infiltrating cells in different skin cell compartments, comparing early with longer-lasting wheals and lesional with uninvolved skin. Sequential biopsies were studied in cold urticaria (CU). No increase of MHC class II molecule expression was found in early prick test wheals to common inhalant allergens. In CU, however, sequential biopsies demonstrated an up-regulation of MHC class II molecules within 30 min after elicitation. This was more pronounced in longer-lasting urticaria lesions of acute, chronic recurrent and delayed pressure urticaria, with HLA-DR and, to a lesser degree, HLA-DP and HLA-DQ being noted on cell infiltrates, on vascular endothelia and around nerves and sweat glands. Nonelesional skin in these types of urticaria also showed increased MHC class II expression. Longer-lasting urticarial wheals are thus associated with up-regulation of MHC class II molecules on resident and infiltrating cells, suggesting an involvement of these molecules in the pathomechanisms of these types of urticarial lesions.

Cytokine secretion by human mast cells

Abstract Mast cells have been traditionally viewed as effector cells of immediate‐type hypersensitivity reactions. Besides this, mast cell activation and degranulation have been associated with various biologically and clinically important functions. Results of the past few years suggest that mast cells are involved in the development of late‐phase reactions and influence other chronic inflammatory responses through the generation and secretion of various multipotcnlial cylokines.

Thrombin and melittin activate phospholipase C in human HaCaT keratinocytes

Abstract Following the activation of specific receptors, phospholipase C has been shown to cleave the membrane phospholipid phosphatidylinositol bisphosphate into the 2nd messengers inositol 1,4,5‐trisphosphate and di‐acylgiycerol. Both 2nd messengers contribute to the regulation of cellular proliferation. The receptor for bradykinin is coupled to this pathway in keratinocytes, but knowledge about other activators of phospholipase C is limited. Additional mediators and agents were therefore examined regarding their ability to activate phospholipase C in HaCaT keratinocytes. Analysis for 3H‐inositol phosphates was performed by anion‐exchange HPLC. Thrombin and melittin induced a time‐ and dose‐dependent release of inositol 1,4,5‐trisphosphate. Several other mediators examined such as angiotension II, neurotensin, C3a, pituitary adenylate cyclase activating peptide, phenylephrin, and prostaglandin E2, did not induce the formation of inositol phosphates. In view of the mitogenic activity and the increased formation of thrombin after tissue injury, the coupling of the thrombin receptor to phospholipase C in HaCaT keratinocytes suggests a rôle of this protease in epidermal wound healing.

Tumor necrosis factor receptors of the monocyte derived Langerhans cell phenotype "MoLC".

For the generation of dendritic Langerhans cells from progenitors TNF-a is used as a differentiation factor in combination with a growth factor , GM-CSF (1). Growth factors primarily act by suppression of apoptosis (2). During monocyte development they probably regulate the response to TNF-a by inhibiting the cytotoxic signal which is associated with the last 100 aminoacids of the C-terminus of TNF-RI (p55) (3), and activating only the p75 TNF-R.