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Interleukin-8 and monocyte chemotactic protein-1 gene expression and protein production by human orbital fibroblasts. 1998

V M Elner, and M A Burnstine, and S L Kunkel, and R M Strieter, and S G Elner
Department of Ophthalmology, W. K. Kellogg Eye Center, University of Michigan Medical School, Ann Arbor, USA.

Orbital inflammation is common, but the mechanisms underlying leukocytic infiltration of orbital tissue are poorly understood. We studied resident human orbital fibroblasts (OF) interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) mRNA expression and protein secretion in response to lipopolysaccharide (LPS) or recombinant human cytokines that are present during inflammation. Third-passaged cultured human OF were left unstimulated or incubated with varying concentrations of LPS, recombinant interleukin-1-beta (rIL-1 beta), recombinant tumor necrosis factor-alpha (rTNF-alpha), or recombinant interferon-gamma (rIFN-gamma) for 2, 4, 8, or 24 h. Northern blot analysis and ELISA were performed to determine OFIL-8 and MCP-1 mRNA expression and protein secretion, respectively. Experiments were performed in triplicate and repeated four times on different cell lines. OF lacked constitutive IL-8 or MCP-1 gene expression, but produced substantial dose-dependent increases in steady-state IL-8 and MCP-1 mRNA expression by 2 h of LPS or cytokine stimulation (rIL-1 beta > fTNF-alpha > LPS > rIFN-gamma), maintained at 24 h ELISA for IL-8 and MCP-1 proteins showed significant time- and dose-dependent OF secretion after exposure to recombinant cytokine or LPS (rIL-1 beta > rTNF-alpha > LPS), measured after 4 h of exposure (p < 0.01). This increased in the media over the next 20 h. rIFN-gamma was a potent stimulant of OF MCP-1, significant by 2 h (p < 0.05), but only a weak stimulant of IL-8 at 24 h. OF secreted IL-8 and MCP-1 in response to LPS and proinflammatory cytokines, indicating that these resident cells within the orbit have the capacity to actively participate in the initiation and propagation of orbital inflammation. Strategies aimed at modulating local mediators may be helpful in the management of orbital inflammatory disease.

UI MeSH Term Description Entries
D008070 Lipopolysaccharides Lipid-containing polysaccharides which are endotoxins and important group-specific antigens. They are often derived from the cell wall of gram-negative bacteria and induce immunoglobulin secretion. The lipopolysaccharide molecule consists of three parts: LIPID A, core polysaccharide, and O-specific chains (O ANTIGENS). When derived from Escherichia coli, lipopolysaccharides serve as polyclonal B-cell mitogens commonly used in laboratory immunology. (From Dorland, 28th ed) Lipopolysaccharide,Lipoglycans
D009915 Orbit Bony cavity that holds the eyeball and its associated tissues and appendages. Eye Socket,Eye Sockets,Orbits,Socket, Eye,Sockets, Eye
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D002478 Cells, Cultured Cells propagated in vitro in special media conducive to their growth. Cultured cells are used to study developmental, morphologic, metabolic, physiologic, and genetic processes, among others. Cultured Cells,Cell, Cultured,Cultured Cell
D004305 Dose-Response Relationship, Drug The relationship between the dose of an administered drug and the response of the organism to the drug. Dose Response Relationship, Drug,Dose-Response Relationships, Drug,Drug Dose-Response Relationship,Drug Dose-Response Relationships,Relationship, Drug Dose-Response,Relationships, Drug Dose-Response
D004797 Enzyme-Linked Immunosorbent Assay An immunoassay utilizing an antibody labeled with an enzyme marker such as horseradish peroxidase. While either the enzyme or the antibody is bound to an immunosorbent substrate, they both retain their biologic activity; the change in enzyme activity as a result of the enzyme-antibody-antigen reaction is proportional to the concentration of the antigen and can be measured spectrophotometrically or with the naked eye. Many variations of the method have been developed. ELISA,Assay, Enzyme-Linked Immunosorbent,Assays, Enzyme-Linked Immunosorbent,Enzyme Linked Immunosorbent Assay,Enzyme-Linked Immunosorbent Assays,Immunosorbent Assay, Enzyme-Linked,Immunosorbent Assays, Enzyme-Linked
D005347 Fibroblasts Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules. Fibroblast
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D012333 RNA, Messenger RNA sequences that serve as templates for protein synthesis. Bacterial mRNAs are generally primary transcripts in that they do not require post-transcriptional processing. Eukaryotic mRNA is synthesized in the nucleus and must be exported to the cytoplasm for translation. Most eukaryotic mRNAs have a sequence of polyadenylic acid at the 3' end, referred to as the poly(A) tail. The function of this tail is not known for certain, but it may play a role in the export of mature mRNA from the nucleus as well as in helping stabilize some mRNA molecules by retarding their degradation in the cytoplasm. Messenger RNA,Messenger RNA, Polyadenylated,Poly(A) Tail,Poly(A)+ RNA,Poly(A)+ mRNA,RNA, Messenger, Polyadenylated,RNA, Polyadenylated,mRNA,mRNA, Non-Polyadenylated,mRNA, Polyadenylated,Non-Polyadenylated mRNA,Poly(A) RNA,Polyadenylated mRNA,Non Polyadenylated mRNA,Polyadenylated Messenger RNA,Polyadenylated RNA,RNA, Polyadenylated Messenger,mRNA, Non Polyadenylated
D015152 Blotting, Northern Detection of RNA that has been electrophoretically separated and immobilized by blotting on nitrocellulose or other type of paper or nylon membrane followed by hybridization with labeled NUCLEIC ACID PROBES. Northern Blotting,Blot, Northern,Northern Blot,Blots, Northern,Blottings, Northern,Northern Blots,Northern Blottings

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