Oxidized low-density lipoprotein induces calpain-dependent cell death and ubiquitination of caspase 3 in HMEC-1 endothelial cells.

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Oxidized low-density lipoprotein induces calpain-dependent cell death and ubiquitination of caspase 3 in HMEC-1 endothelial cells. / Ares, Isabella; Saido, Takaomi C; Andersson, Tommy; Ares, Mikko P S.

In: Biochem J, Vol. 374, No. Pt 2, 2003, p. 403-411.

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T1 - Oxidized low-density lipoprotein induces calpain-dependent cell death and ubiquitination of caspase 3 in HMEC-1 endothelial cells.

AU - Ares, Isabella

AU - Saido, Takaomi C

AU - Andersson, Tommy

AU - Ares, Mikko P S

PY - 2003

Y1 - 2003

N2 - Oxidized low-density lipoprotein (oxLDL) is known to induce apoptosis in endothelial cells, and this is believed to contribute to the progression of atherosclerosis. In the present study we made the novel observation that oxLDL-induced death of HMEC-1 cells is accompanied by activation of calpain. The mu-calpain inhibitor PD 151746 decreased oxLDL-induced cytotoxicity, whereas the general caspase inhibitor BAF (t-butoxycarboryl-Asp-methoxyfluoromethylketone) had no effect. Also, oxLDL provoked calpain-dependent proteolysis of cytoskeletal a-fodrin in the HMEC-1 cells. Our observation of an autoproteolytic cleavage of the 80 kDa subunit of mu-calpain provided further evidence for an oxLDL-indunced stimulation of calpain activity. The Bcl-2 protein Bid was also cleaved during oxLDL-elicited cell death, and this was prevented by calpain inhibitors, but not by inhibitors of cathepsin B and caspases. Treating the HMEC-1 cells with oxLDL did not result in detectable activation of procaspase 3 or cleavage of PARP [poly(ADP-ribose) polymerase], but it did cause polyubiquitination of caspase 3, indicating inactivation and possible degradation of this protease. Despite the lack of caspase 3 activation, oxLDL treatment led to the formation of nucleosomal DNA fragments characteristic of apoptosis. These novel results show that oxLDL initiates a calpain-mediated death-signalling pathway in endothelial cells.

AB - Oxidized low-density lipoprotein (oxLDL) is known to induce apoptosis in endothelial cells, and this is believed to contribute to the progression of atherosclerosis. In the present study we made the novel observation that oxLDL-induced death of HMEC-1 cells is accompanied by activation of calpain. The mu-calpain inhibitor PD 151746 decreased oxLDL-induced cytotoxicity, whereas the general caspase inhibitor BAF (t-butoxycarboryl-Asp-methoxyfluoromethylketone) had no effect. Also, oxLDL provoked calpain-dependent proteolysis of cytoskeletal a-fodrin in the HMEC-1 cells. Our observation of an autoproteolytic cleavage of the 80 kDa subunit of mu-calpain provided further evidence for an oxLDL-indunced stimulation of calpain activity. The Bcl-2 protein Bid was also cleaved during oxLDL-elicited cell death, and this was prevented by calpain inhibitors, but not by inhibitors of cathepsin B and caspases. Treating the HMEC-1 cells with oxLDL did not result in detectable activation of procaspase 3 or cleavage of PARP [poly(ADP-ribose) polymerase], but it did cause polyubiquitination of caspase 3, indicating inactivation and possible degradation of this protease. Despite the lack of caspase 3 activation, oxLDL treatment led to the formation of nucleosomal DNA fragments characteristic of apoptosis. These novel results show that oxLDL initiates a calpain-mediated death-signalling pathway in endothelial cells.

KW - Lipoproteins

KW - Intracellular Fluid: metabolism

KW - Hydrolysis

KW - Human

KW - Growth Inhibitors: toxicity

KW - Enzyme Activation: drug effects

KW - Vascular: cytology

KW - Calcium: metabolism

KW - Apoptosis: drug effects

KW - Endothelium

KW - Caspases: metabolism

KW - Carrier Proteins: metabolism

KW - Calpain: physiology

KW - Cell Line

KW - DNA Fragmentation: physiology

KW - Vascular: physiology

KW - Vascular: enzymology

KW - Endopeptidases: physiology

KW - LDL: toxicity

KW - Microfilament Proteins: metabolism

KW - Oxidation-Reduction

KW - Proto-Oncogene Proteins c-bcl-2: metabolism

KW - Support

KW - Non-U.S. Gov't

KW - Ubiquitin: metabolism

U2 - 10.1042/BJ20021955

DO - 10.1042/BJ20021955

M3 - Article

VL - 374

SP - 403

EP - 411

JO - Biochemical Journal

T2 - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - Pt 2

ER -