Pharmacological strategies to interfere with proinflammatory signal transduction in endothelial cells

Endothelial cells are actively involved in (chronic) inflammatory diseases like rheumatoid arthritis or glomerulonephritis, by playing a key role in ongoing leukocytes recruitment and activation, and angiogenesis. Since endothelial cells line the inner side of the blood vessels, they are well accessible for (non-targeted as well as targeted) drug treatment strategies.
We aimed to study the therapeutic potential of pharmacological interference with inflammatory endothelial cell activation to inhibit the expression of proinflammatory genes involved in leukocyte recruitment.
Using quantitative RT-PCR analysis of inflammatory genes in endothelial cells in vitro we showed that simultaneous inhibition of two main signalling pathways, NF-kappaB and p38 MAPK, resulted in significantly stronger downregulation of inflammatory genes compare to inhibition of each pathway separately. In vivo we studied microvascular endothelial cell responses to acute activation and drug treatment in different mouse organs, and in more detail in different microvascular beds in the kidney. Significant heterogeneity in the expression of endothelial markers and inflammatory molecules on both gene and protein levels was observed. Additionally, using laser microdissection assisted endothelial cell isolation prior to gene expression analysis, we revealed vascular bed-specific pharmacological effects of the anti-inflammatory drug dexamethasone which were masked when analysing drug effects in whole tissue analysis.
Using a targeted drug delivery approach, we showed that we can modify adenovirus based gene delivery systems in such a way that the encoded gene, in our model an inhibitor of NF-kappaB, was selectively delivered in activated endothelium in renal inflammation. The transgene prevented the expression of endothelial activation genes in the diseased organ.
These studies demonstrate that pharmacological intervention with microvascular endothelial cell activation has therapeutic potential. The observation that microvascular beds behave differently depending on their location in the body has important implications for rational drug treatment schedule design for inflammatory diseases.

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	11_thesis.pdf

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