ATM protects against oxidative stress induced by oxidized low-density lipoprotein.

Chronic oxidative stress is involved in the pathogenesis of multiple inflammatory diseases, including cardiovascular disease and atherosclerosis. The rare autosomal recessive disorder Ataxia-telangiectasia (A-T) is characterized by progressive cerebellar ataxia secondary to Purkinje cell death, immunodeficiency, and increased cancer incidence. ATM, the protein mutated in A-T, plays a key role in cellular DNA-damage responses. A-T cells show poor cellular anti-oxidant defences and increased oxidant sensitivity compared to normal cells, and ATM functions, in part, as an oxidative stress sensor. The oxidation of low-density lipoprotein (oxLDL) and its uptake by macrophages is an initiating step in the development of atherosclerosis. We demonstrate that oxLDL activates ATM and downstream p21 expression in normal fibroblasts and endothelial cells. In ATM-deficient fibroblasts oxLDL induces DNA double-strand breaks, micronuclei formation and causes chromosome breaks. Furthermore, oxLDL decreases cell viability and inhibits colony formation in A-T fibroblasts more effectively as compared to normal controls. Formation of oxLDL-induced reactive oxygen species is significantly higher in A-T, than normal fibroblasts. Last, pre-treatment of cells with ammonium pyrrolidine dithiocarbamate, a potent antioxidant and inhibitor of transcription factor nuclear factor κB, reduces oxLDL-induced reactive oxygen species formation. Our data indicates that ATM functions in the defence against oxLDL-mediated cytotoxicity.

Hypochlorite-modified high-density lipoprotein promotes induction of HO-1 in endothelial cells via activation of p42/44 MAPK and zinc finger transcription factor Egr-1.

Modification/chlorination of high-density lipoprotein (HDL) by hypochlorous acid (HOCl), formed by the myeloperoxidase-H2O2-chloride system of activated phagocytes, converts an anti-atherogenic lipoprotein into a pro-inflammatory lipoprotein particle. Chlorinated HDL is present in human lesion material, binds to and is internalized by endothelial cells and impairs expression and activity of endothelial nitric oxide synthase. The present study aimed at clarifying whether exposure of endothelial cells to pro-inflammatory HOCl-HDL impacts on expression of heme oxygenase-1, a potential rescue pathway against endothelial dysfunction. Our findings revealed that HDL modified by HOCl, added as reagent or generated enzymatically, induced phosphorylation of p42/44 mitogen-activated protein kinase, expression of transcription factor early growth response-1 (Egr-1) and enhanced expression of heme oxygenase-1 in human endothelial cells. Upregulation of heme oxygenase-1 could be blocked by an inhibitor upstream of p42/44 mitogen-activated protein kinase and/or knockdown of Egr-1 by RNA-interference. Electrophoretic mobility shift assays demonstrated HOCl-HDL-mediated induction of the Egr-1 DNA binding activity. Immunocytochemical and immunoblotting experiments demonstrated HOCl-HDL-induced translocation of Egr-1 to the nucleus. The present study demonstrates a novel compensatory pathway against adverse effects of HOCl-HDL, providing cytoprotection in a number of pathological conditions including cardiovascular disease.