Endothelial cell gene expression in inflammation

Activation of endothelium is a critical component of inflammation, allowing the recruitment of leukocytes into the tissues and a shift towards a prothrombotic state 1. Whilst activation of EC by proinflammatory cytokines leads to the expression of many proinflammatory genes (eg adhesion molecules and chemoattractants), this response is matched by a compensatory expression of anti-inflammatory and cytoprotective genes involved in containing the inflammatory response. These serve to attenuate the intensity and/or duration of an inflammatory response and to reduce endothelial injury 2. We are interested in characterising endogenous cytoprotective pathways as potential avenues for therapeutic intervention.

Our group has a long-standing interest in the use of pig models to bridge between rodents and humans for studies of vascular endothelial activation during inflammatory responses. This has involved the generation of cDNA and antibodies to adhesion molecules (E- and P-selectin, ICAM-1, VCAM-1) 3-6, the use of skin models for studying endothelial activation in relation to leukocyte trafficking 7-10, the development of a sub-cutaneous sponge model 11 and the development of molecular imaging of endothelial activation using E-selectin as a target 12, 13.

In line with our interests in porcine models of cardiovascular disease, we have already generated cDNA and monoclonal antibodies to adhesion molecules on porcine endothelium, and studied leukocyte trafficking into skin and subcutaneous sponges. performed a suppressive subtractive hybridisation of mRNA from TNFα-activated versus unstimulated porcine aortic EC (PAEC)14. This resulted in our obtaining separate cDNA libraries of genes that are upregulated and downregulated in PAEC by TNFα. Sequencing of approximately 50 clones from the TNFα-upregulated library has revealed a number of genes known to be regulated by TNFα (eg VCAM-1, IL-8, MCP-1, IP-10) as well as a number of genes not previously known to be TNFαregulated.  Novel genes have been further screened using real-time RT-PCR. We are currently characterising in detail a novel intracellular protein with a putative cytoprotective action.

Reference List

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2. Bach FH, Hancock WW, Ferran C. Protective genes expressed in endothelial cells: a regulatory response to injury. Immunol Today 1997 October;18(10):483-6.
3. Tsang YTM, Haskard DO, Robinson MK. Cloning and expression kinetics of porcine vascular cell adhesion molecule. Biochem Biophys Res Commun 1994;201:805-12.
4. Tsang Y, Stevens PE, Licence ST, Haskard DO, Binns RM, Robinson MK. Porcine E-selectin: cloning and functional characterization. Immunology 1995;85:140-5.
5. Stocker C, Sugars K, Harari O, Landis RC, Morley BJ, Haskard DO. TNFa, IL-4 and IFNg regulate differential expression of P- and E-selectin expression by porcine aortic endothelial cells. J Immunol 2000;164:3309-15.
6. Stocker CJ, Sugars KL, Yarwood H, Delikouras A, Lechler RI, Dorling A, Landis RC, Morley BJ, Haskard DO. Cloning of porcine intercellular adhesion molecule-1 and characterization of its induction on endothelial cells by cytokines. Transplantation 2000 August 27;70(4):579-86.
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10. Harrison AA, Stocker CJ, Chapman PT, Tsang YT, Huehns TY, Gundel RH, Peters AM, Davies KA, George AJ, Robinson MK, Haskard DO. Expression of vascular cell adhesion molecule-1 by vascular endothelial cells in immune and nonimmune inflammatory reactions in the skin. J Immunol 1997 November 1;159(9):4546-54.
11. Woolley ST, Whyte A, Licence ST, Haskard DO, Wooding FBP, Binns RM. Differences in E-selectin expression and leukocyte infiltration induced by inflammatory agents in a novel sub-cutaneous sponge- matrix model. Immunology 1995;84:55-63.
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