PDGF stimulates DNA synthesis in human vascular smooth muscle cells via a novel wortmannin-insensitive phosphatidylinositol 3-kinase.

The class 1(A) phosphatidylinositol 3-kinase enzymes consist of a number of heterodimeric complexes of regulatory and catalytic subunits and have been implicated in a number of cellular responses. While platelet-derived growth factor (PDGF)-induced chemotaxis of human vascular smooth muscle cells (HVSMC) is inhibited by both wortmannin and LY294002, DNA synthesis is only inhibited by LY294002. Serum-induced DNA synthesis however is inhibited by LY294002, wortmannin and rapamycin. Similarly PDGF-induced protein kinase B (PKB) activation is inhibited by LY294002 but not by wortmannin or rapamycin. In conclusion PDGF-induced DNA synthesis appears to occur through a phosphatidylinositol 3-kinase (PI3-K)-dependent, but wortmannin-insensitive, PKB/Akt pathway.

Thrombospondin-1 differentially induces chemotaxis and DNA synthesis of human venous smooth muscle cells at the receptor-binding level.

Thrombospondin-1 is a large matricellular protein that acts as a pleiotropic growth factor for human vascular smooth muscle cells, and may play a role in the progression of vascular disease. Although we have previously demonstrated the dependence of both thrombospondin-1-stimulated cell chemotaxis and proliferation on tyrosine kinases, the receptor mechanisms involved remain obscure. This investigation aims to determine the nature of the receptor(s) involved in the cellular responses to thrombospondin-1. Cellular signals were identified by western blotting following cell stimulation, while cellular responses were assessed by measuring DNA synthesis and chemotaxis. These data demonstrate that thrombospondin-1-induced cell chemotaxis can be inhibited by a peptide containing the Arg-Gly-Asp motif, a function-blocking alpha(v)beta(3) antibody, a function-blocking integrin-associated protein (IAP) antibody and pertussis toxin, while thrombospondin-1-stimulated DNA synthesis is inhibited by a function-blocking alpha(3)beta(1) antibody. Similarly the Arg-Gly-Asp-containing peptide inhibits tyrosine phosphorylation of focal adhesion kinase and the p85 regulatory subunit of phosphatidylinositol 3-kinase, but does not significantly affect tyrosine phosphorylation, or activation, of extracellular-regulated kinase. These data suggest that soluble thrombospondin-1 interacts with human vascular smooth muscle cells via two independent and separable receptor-binding sites, to differentially stimulate cell chemotaxis and DNA synthesis.

The antimitogenic action of the sulphated polysaccharide fucoidan differs from heparin in human vascular smooth muscle cells.

The sulphated polysaccharides fucoidan and heparin both inhibit vascular smooth muscle cell (VSMC) proliferation. In this study we compared their actions on mitogenesis and ERK1/ERK2 activation in human VSMC. Although they displaced cell surface [3H]-heparin binding with similar affinity, they exerted clearly distinguishable actions. Fucoidan potently inhibited DNA synthesis stimulated by foetal calf serum, PDGF-BB and thrombospondin-1. Heparin inhibited the mitogenic action of serum and thrombospondin- I (though less potently than fucoidan), but failed to inhibit PDGF-BB-induced DNA synthesis. In parallel studies, fucoidan, but not heparin, inhibited ERK1/ERK2 activation by PDGF-BB. Moreover, fucoidan inhibited serum-induced mitogenesis in "heparin resistant" VSMC, which are refractory to heparin's antimitogenic action. In summary, the structurally different polysaccharides, heparin, fucoidan (and fucans) have distinguishable effects on mitogenesis and ERK1/ERK2 activation, suggesting that different mechanism(s) mediate these actions. The potent antimitogenic action of fucoidan and its efficacy in heparin resistant VSMC emphasise the need to further investigate its mechanism of action in human VSMC and suggest this agent could have therapeutic potential.