Control of endothelial cell proliferation by calcium influx and arachidonic acid metabolism: A pharmacological approach

Abstract

In physiological conditions, endothelial cell proliferation is strictly controlled by several growth factors, among which bFGF and VEGF are the most effective. Both bind to specific tyrosine kinase receptors and trigger intracellular signal cascades. In particular, bFGF stimulates the release of arachidonic acid (AA) and its metabolites in many types of endothelial cells in culture. In bovine aortic endothelial cells, it has been suggested that AA is released by the recruitment of cytosolic phospholipase A2 (cPLA2). AA metabolites are involved in the control of both endothelial cell motility (mostly via the cyclooxygenase pathway) and proliferation (via the lipoxygenase (LOX) cascade). On the other hand, evidence has been provided for a proliferative role of AA-induced calcium influx. By using a pharmacological approach, we have tried to elucidate the contribution to bovine aortic endothelial proliferation of the different pathways leading to production of AA and its metabolites. Two main informations were obtained by our experiments: first, AA release is not entirely due to cPLA2 involvement, but also to DAG lipase recruitment; second, cyclooxygenase derivatives play a role in the control of cell proliferation, and not only of motility. Moreover, by combining proliferation assays and single cell calcium measurements, we show that the blocking effect of carboxyamido-triazole (CAI), an inhibitor of tumor growth and angiogenesis acting on calcium influx-dependent pathways, including AA metabolism, is at least in part due to a direct effect on AA-induced calcium influx.

Arachidonic Acid–Induced Ca2+ Entry Is Involved in Early Steps of Tumor Angiogenesis

“…We previously showed that AA promotes cell proliferation in bovine aortic endothelial cells. This effect is at least partially due to AA itself (11) although a role for eicosanoids of cyclooxygenase and lypooxygenase pathway has been proposed (17). Here, we show that both AA and ETYA induce cell proliferation in BTECs in accordance with a role for the fatty acid both in normal and tumor-derived endothelial cells.…”

“…The dose of antiangiogenic agents is peculiar in determining the efficacy of combination therapies. CAI has been shown to exert its blocking effects starting from 1 Amol/L in normal endothelial cells (5,17,20). Higher doses of CAI have antiangiogenic activity in different systems such as mouse presenting ischemic retinopathy, rat aortic ring culture, or chorioallantoic membrane (18,20,62).…”

“…This is of particular interest considering that CAI is under phase I and phase II clinical trials for the treatment of solid tumors (22,(24)(25)(26)(27). CAI is a nonselective blocker of nonvoltage-gated Ca 2+ channels and it has been shown to inhibit both SOCE and NSOCE (17,(66)(67)(68). In particular, we previously showed that CAI blocks AAinduced NSOCE in normal endothelial cells (17).…”

“…Recently, we showed that AA induces a sustained NSOCE involved in the control of proliferation. It is probably due to more than one type of calcium channel, whose molecular nature is not known in bovine aortic endothelial cells (11,12,17).…”

“…The blockage by CAI results in inhibition of endothelial cell attachment to extracellular matrix substrate, migration, and proliferation in vivo and in vitro (17,(19)(20)(21). Currently, CAI is under investigation as an orally administered tumoristatic and antiangiogenic agent in clinical phase I and phase II trials (22)(23)(24)(25)(26)(27).…”