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Tumor necrosis factor-alpha induces interleukin-1 alpha and interleukin-1 receptor antagonist production by cultured human keratinocytes.

Interleukin-1 (IL-1) may have a significant pro-inflammatory effect in the skin; an imbalance in its production has been linked to cutaneous disease processes. IL-1 receptor antagonist (IL-1ra) is a recently described competitive inhibitor of IL-1 alpha and IL-1 beta that binds to human types I and II IL-1 receptors without apparent cell activation. Human keratinocytes synthesize IL-1ra, IL-1 alpha, and IL-1 beta but fail to secrete these cytokines. This study investigated IL-1ra and IL-1 alpha accumulation by cultured keratinocytes stimulated by tumor necrosis factor-alpha (TNF-alpha), IL-3, IL-4, IL-6, IL-10, interferon-gamma, granulocyte-macrophage colony-stimulating factor, granulocyte colony-stimulating factor, and macrophage colony-stimulating factor and by various extracellular matrix proteins, conditions that these cells may encounter in normal or inflamed skin in vivo. IL-1ra and IL-1 alpha proteins were measured by specific enzyme-linked immunosorbent assay in keratinocyte supernatants and lysates. Only TNF-alpha induced IL-1ra and IL-1 alpha production. TNF-alpha added to culture in amounts of 10 ng/ml or higher, induced a twofold increase in intracellular levels of both IL-ra and IL-1 alpha without secretion at 48 h. The IL-1ra concentration in keratinocyte lysates increased from 9.6 to 17.6 ng/ml after TNF-alpha stimulation, and the IL-1 alpha concentration increased from 1.0 to 3.3 ng/ml. Keratinocytes also exhibited comparable increases in IL-1 alpha and IL-1ra mRNA levels after 12 h in culture with TNF-alpha, as determined by in vitro hybridization to specific cDNA probes. The IL-1 alpha and IL-1ra response to TNF-alpha stimulation showed a varied pattern among different keratinocyte strains over 72 h of culture on plain plastic. In contrast, extracellular matrix proteins (laminin, fibronectin, collagen I and IV, and vitronectin) did not stimulate keratinocyte accumulation of IL-1 alpha or IL-1ra proteins after 72 h in culture. When TNF-alpha was added to cells cultured on these matrices, no change in IL-1 alpha or IL-1ra production was observed above that which could be attributed to TNF-alpha alone. In conclusion, TNF-alpha, but not the extracellular matrix proteins tested, stimulated production of intracellular IL-1 alpha and IL-1ra by keratinocytes. The ratio of IL-1ra to IL-1 alpha after TNF-alpha stimulation of keratinocytes may influence the inflammatory profile in the epidermis.
C L Kutsch, D A Norris, W P Arend

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