IL-6 Bioassay, Propagation Model - MyBio Ireland - Promega

IL-6 Bioassay, Propagation Model


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Interleukin-6 (IL-6, originally termed BCDF/BSF2) was first identified in 1983 alongside molecules with similar but distinct functional properties (IL-4/BCGF1/BSF1 and IL-5/BCGF2). IL-6 is a small (~21kD) pleiotropic cytokine that is secreted by a wide range of cell types: immune (macrophages, dendritic, mast, B and T cells), hepatocytes, synovial fibroblasts, bone marrow, dermal fibroblasts and keratinocytes, mesangial, vascular endothelial and smooth muscle. The IL-6 receptor (IL-6R) is composed of two subunits. IL-6Rα (CD126) is a single chain transmembrane protein that specifically binds the cytokine directly and is expressed on limited cell types including macrophages, neutrophils, CD4+ T cells, podocytes and hepatocytes. The second subunit is a large 130kDa signal transducing chain, glycoprotein 130 (gp130/CD130), that is expressed on most cell types. IL-6 signals in three ways: classical signaling (membrane IL-6Rα and gp130), trans signaling (cytokine associated with soluble IL-6Rα) and trans presentation (dendritic IL-6/IL-6Rα complex presenting to a second cell expressing gp130).Downstream pathway signaling doesn’t occur until the IL-6/IL-6Rα complex associates with gp130 and triggers a conformational change to activate tyrosine kinases JAK1, JAK2 and Tyk2, which in turn activate STAT3. Dimerized STAT3 translocates to the nucleus and activates transcription of a variety of genes, such as proinflammatory cytokines (IL-1β, IL-8), anti-apoptotic proteins (cyclin D1, MYC, Bcl-X) and immunosuppressive proteins (VEGF, IL-10, TGFβ). IL-6 is a member of the IL-6 cytokine family, which includes IL-11, leukemia inhibitory factor (LIF), oncostatin M, ciliary neurotrophic factor (CNTF), cardiotrophin-1 (CT-1) and cardiotrophin-like cytokine (CLC). All of these cytokines signal through gp130 and the STAT3 pathway. IL-6 is transiently secreted following tissue damage or stress (UV irradiation, reactive oxygen species, microbial, viral). It is also one of the cytokines released during bacterial sepsis. Furthermore, IL-6, along with transforming growth factor β (TGFβ), promotes differentiation of CD4+ T cells into Th17 cells and inhibits differentiation of regulatory T cells, thus playing a critical role in autoimmunity. IL-6 has shown to be elevated in patients with rheumatoid arthritis (RA), Crohn’s disease and Castleman’s disease. Tocilizumab, the first successful IL-6R targeted therapy, blocks IL-6 signaling through both membrane and soluble forms of IL-6R. This biologic drug is approved for RA treatment in Japan, Europe and the USA (7). Additional IL-6 blocking therapeutic antibodies include siltuximab and sarilumab. The IL-6 Bioassay, Propagation Model (Cat.# J2992) is a bioluminescent cell-based assay designed to measure IL-6 stimulation or inhibition. The IL-6 Bioassay Cells are provided in Cell Propagation Model (CPM) format, as cryopreserved cells that can be thawed, propagated and banked for long-term use (also offered in a thaw-and-use format; Cat.# JA2501, JA2505).The IL-6 Bioassay consists of a human cell line engineered to express a luciferase reporter driven by a response element (RE). When IL-6 binds, the IL-6R transduces intracellular signals resulting in luminescence (Figure 1). The bioluminescent signal is detected and quantified using Bio-Glo™ Luciferase Assay System(e) (Cat.# G7940, G7941) and a standard luminometer, such as the GloMax(R) Discover System (see Related Products, Section 9.B).

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