Oxidized LDLs Induce Massive Apoptosis of Cultured Human Endothelial Cells Through a Calcium-Dependent Pathway

Abstract

Oxidized LDLs are thought to play a central role in atherogenesis. Among their wide variety of biological properties, oxidized LDLs exhibit a cytotoxic effect on cultured vascular cells. Toxic doses of mildly oxidized LDLs elicited massive apoptosis in both primary and immortalized cultures of endothelial cells as shown by characteristic morphological and biochemical changes. Cytoplasmic and nucleic modifications (eg, chromatin condensation and nucleus fragmentation) were visualized by using electron and fluorescence microscopy of intact cells labeled by the fluorescent DNA probe SYTO-11. DNA fragmentation was quantified by ultracentrifugation of chromatin fragments, evaluated in situ by using the TUNEL (Terminal transferase-mediated dUTP-biotin nick end labeling) procedure, and visualized by electrophoresis of radiolabeled DNA fragments showing the characteristic apoptotic ladder. Apoptotic cells became rapidly detached and underwent postapoptotic necrosis that led to cell disintegration. Apoptosis was subsequent to a sustained and delayed peak of cytosolic calcium. Both the calcium peak and apoptosis were blocked by chelating the extracellular calcium with EGTA or by inhibiting the calcium influx by the calcium-channel blockers nifedipine and nisoldipine, thus suggesting that the apoptotic process induced by oxidized LDLs is clearly calcium dependent. Aurintricarboxylic acid, an inhibitor of endonucleases, also blocked the apoptotic process without blocking the calcium peak. These results suggest that toxic doses of mildly oxidized LDLs induce massive apoptosis of endothelial cells through a calcium-dependent mechanism and that this apoptotic process can be prevented by inhibiting the rise of cytosolic calcium or by inhibiting cellular endonucleases by aurintricarboxylic acid.

IL-10 Prevents Apoptosis of Brain Endothelium during Bacteremia

“…Others in the field have also reported apoptosis of various eukaryotic cells upon exposure to borrelia spirochetes (40)(41)(42)(43)(44). Because infection with various species of RF spirochetes can result in cerebrovascular disease in humans (3)(4)(5)(6) and experimental animals (6,7,45,46) and the major virulence factor of borrelia spirochetes, the outer membrane lipoproteins (47), is capable of causing eukaryotic cell apoptosis (41), we hypothesized that bacterial lipoproteins from B. turicatae can cause brain endothelial cell apoptosis, similar to what has been reported for oxidized human lipoproteins (48)(49)(50)(51). We investigated this possibility in mice with relapsing-remitting infection and in vitro.…”

Apoptosis in the vasculature: mechanisms and functional importance

“…In vitro studies had already shown that oxLDL exhibit cytotoxic e ects on cultured endothelial cells, but the type of cell death elicited remained unknown (Hessler et al, 1979). However, several authors recently reported increased apoptotic cell death of bovine aortic endothelial cells exposed to cholesterol oxides in culture (Lizard et al, 1996) and increased apoptosis (and possibly secondary necrosis) of HUVEC after exposure to oxLDL (Dimmeler et al, 1997a;Escargueil-Blanc et al, 1997). OxLDL-induced apoptosis is subsequent to a sustained and delayed peak in cytosolic calcium and is prevented by chelating extracellular calcium or by inhibiting calcium in¯ux (Escargueil-Blanc et al, 1997).…”

“…However, several authors recently reported increased apoptotic cell death of bovine aortic endothelial cells exposed to cholesterol oxides in culture (Lizard et al, 1996) and increased apoptosis (and possibly secondary necrosis) of HUVEC after exposure to oxLDL (Dimmeler et al, 1997a;Escargueil-Blanc et al, 1997). OxLDL-induced apoptosis is subsequent to a sustained and delayed peak in cytosolic calcium and is prevented by chelating extracellular calcium or by inhibiting calcium in¯ux (Escargueil-Blanc et al, 1997). Furthermore, ceramide generation by acid sphingomyelinase and activation of caspase-3 are indispensable for oxLDL-induced apoptosis (Harada-Shiba et al, 1998).…”

Alterations of Extracellular Calcium Elicit Selective Modes of Cell Death and Protease Activation in SH‐SY5Y Human Neuroblastoma Cells

“…, in human epithelial cells (Escargueil-Blanc et al, 1997), and in malignant B cells (Shan et al, 1998). Decreases in Ca2+, may lead to apoptosis.…”