The pro-inflammatory oxidant hypochlorous acid induces Bax-dependent mitochondrial permeabilisation and cell death through AIF-/EndoG-dependent pathways

At sites of chronic inflammation, such as in the inflamed rheumatoid joint, activated neutrophils release hydrogen peroxide (H2O2) and the enzyme myeloperoxidase to catalyse the formation of hypochlorous acid (HOCl). 3-chlorotyrosine, a marker of HOCl in vivo, has been observed in synovial fluid proteins from rheumatoid arthritis patients. However the mechanisms of HOCl-induced cytotxicity are unknown. We determined the molecular mechanisms by which HOCl induced cell death in human mesenchymal progenitor cells (MPCs) differentiated into a chondrocytic phenotype as a model of human cartilage cells and show that HOCl induced rapid Bax conformational change, mitochondrial permeability and release of intra-mitochondrial pro-apoptotic proteins which resulted in nuclear translocation of AIF and EndoG. siRNA-mediated knockdown of Bax substantially prevented mitochondrial permeability, release of intra-mitochondrial pro-apoptotic proteins. Cell death was inhibited by siRNA-mediated knockdown of Bax, AIF or EndoG. Although we observed several biochemical markers of apoptosis, caspase activation was not detected either by western blotting, fluorescence activity assays or by using caspase inhibitors to inhibit cell death. This was further supported by findings that (1) in vitro exposure of recombinant human caspases to HOCl caused significant inhibition of caspase activity and (2) the addition of HOCl to staurosporine-treated MPCs inhibited the activity of cellular caspases. Our results show for the first time that HOCl induced Bax-dependent mitochondrial permeability which led to cell death without caspase activity by processes involving AIF/EndoG-dependent pathways. Our study provides a novel insight into the potential mechanisms of cell death in the inflamed human joint. (c) 2006 Elsevier Inc. All rights reserved.

Regulation of cardiac myocyte cell death.

Cardiac myocyte death, whether through necrotic or apoptotic mechanisms, is a contributing factor to many cardiac pathologies. Although necrosis and apoptosis are the widely accepted forms of cell death, they may utilize the same cell death machinery. The environment within the cell probably dictates the final outcome, producing a spectrum of response between the two extremes. This review examines the probable mechanisms involved in myocyte death. Caspases, the generally accepted executioners of apoptosis, are significant in executing cardiac myocyte death, but other proteases (e.g., calpains, cathepsins) also promote cell death, and these are discussed. The two principal cell death pathways (death receptor- and mitochondrial-mediated) are described in relation to the emerging structural information for the principal proteins, and they are discussed relative to current understanding of myocyte cell death mechanisms. Whereas the mitochondrial pathway is probably a significant factor in myocyte death in both acute and chronic phases of myocardial diseases, the death receptor pathway may prove significant in the longer term. The Bcl-2 family of proteins are key regulators of the mitochondrial death pathway. These proteins are described and their possible functions are discussed. The commitment to cell death is also influenced by protein kinase cascades that are activated in the cell. Whereas certain pathways are cytoprotective (e.g., phosphatidylinositol 3'-kinase), the roles of other kinases are less clear. Since myocyte death is implicated in a number of cardiac pathologies, attenuation of the death pathways may prove important in ameliorating such disease states, and possible therapeutic strategies are explored.