Inflammatory cytokines and fatty acids regulate endothelial cell heparanase expression

Area of Science:

• Vascular Biology
• Molecular Biology
• Biochemistry

Background:

• Heparan sulfates are crucial for basement membrane organization and endothelial barrier function.
• Heparan sulfate proteoglycans are integral components of the extracellular matrix and cell surface.

Purpose of the Study:

• To investigate if endothelial cells secrete heparanase under inflammatory conditions.
• To identify the signaling pathways regulating heparanase expression and secretion.

Main Methods:

• Cultured endothelial cells treated with cytokines, oxidized LDL, and fatty acids.
• Western blot analysis for heparanase mRNA and protein.
• Measurement of heparanase protein in cell media.
• Inhibition studies using pathway-specific inhibitors (NFkappaB, PI3K, MAP kinase, JNK, caspase-8).
• Analysis of heparanase promoter activity (Sp1 site).
• Immunohistochemical analysis of mouse aorta.

Main Results:

• Cytokines (TNFalpha, IL-1beta), oxidized LDL, and fatty acids induced heparanase mRNA and protein.
• TNFalpha and IL-1beta increased heparanase secretion 2-10 fold.
• Vascular endothelial growth factor (VEGF) reduced heparanase secretion.
• NFkappaB, PI3K, MAP kinase, and JNK pathways did not mediate TNFalpha-induced secretion.
• Caspase-8 inhibition completely abolished TNFalpha-induced heparanase secretion.
• Fatty acids induced heparanase via an Sp1 site in the promoter.
• Increased heparanase staining observed in the endothelium of apoE-null mice aorta.

Conclusions:

• Heparanase is an inducible inflammatory gene product in endothelial cells.
• Caspase-8 is a critical mediator of inflammatory heparanase secretion.
• Heparanase may play a significant role in vascular biology and inflammatory processes.