Abstract 733: Regulation and Function of Hyaluronic Acid Synthase-3 in Human Vascular Smooth Muscle Cells
Hyaluronan (HA) is synthesized by three HA-synthase (HAS) isoforms, HAS1, -2 and -3 and regulates a number of fundamental processes such as migration and cell proliferation. However, the functional significance of individual HAS-isoenzymes is unknown. Aim of the present study was to investigate the response of human coronary smooth muscle cells (SMC) to proinflammatory cytokines with respect to HAS-isoform expression and function. Specifically HAS3-mRNA and HA-synthesis were strongly upregulated in quiescent SMC after incubation with IL-1ß (10ng/ml). This effect could be prevented by dexamethasone (100nM) and the NFκB inhibitor Bay11–7082 (10μM). Similar results were obtained in response to TNFalpha. Upregulation of HAS3 mRNA and HA-synthesis occured also in SMC that were co-cultured with activated U937 cells. In contrast PDGF-BB, vasodilatory prostaglandins and thrombin had no effect on HAS3 expression. In response to IL-ß the molecular weight of HA was found to be much smaller compared to PDGF-BB that induced mainly HAS2 as determined by molecular sieve chromatography. HAS3-siRNA was used to analyse the effect of HAS3 on migration, proliferation and morphology of SMC. SMC were transfected with HAS3-siRNA or non-silencing control-siRNA. HAS3-siRNA significantly reduced HAS3-mRNA levels to 50±22% of control cells. Suppression of HAS3 expression caused
decreased cell spreading to 59.4±9.2% (n=4, p<0.05),
reduction of FCS induced migration as determined by a modified boyden chamber assay (57±8%, n=3, p<0.05) and
inhibition of FCS induced DNA synthesis to 63 ± 8% of controls (n=3, p<0,05) as determined by [3H]-thymidine incorporation.
These findings demonstrate in human SMC that intermediate size HA synthesis by HAS3
occurs specifically in response to the proinflammatory cytokines such as IL-1ß,
participates in the regulation of cell shape and spreading,
supports migration and proliferation.
Therefore release of cytokines by inflammatory cells might induce remodelling of HA-matrix in SMC from high MW HA to low molecular weight HA which might in turn support the proliferative and migratory phenotype of SMC in regions of atherosclerotic plaques with increased inflammatory activity.