Necrotic Death Mechanisms Research Articles

Serpins Save Cells

Biologically active proteases are held in check in part by a family of peptidase inhibitors known as serpins. Most serpins are secreted proteins, but some are intracellular proteins implicated in regulating lysosomal proteases. Knockout mice have failed to reveal the biological roles of these inhibitors, though--possibly because mice have some 28 members of the intracellular variety. Therefore, Luke et al . turned to the worm Caenorhabditis elegans , which has about a third as many intracellular serpins. Loss of one of these, SRP-6, caused animals to become highly sensitive to hypo-osmotic stress and to die as a result of necrotic cell death. This effect appeared to require inhibition of cysteine peptidases because survival of the knockout worms was improved in mutant animals engineered to express wild-type SRP-6 but not in animals that expressed a mutant serpin that lacked inhibitory activity. Calcium mobilization appeared to be required for cell death as SRP-6 knockout animals lacking the ryanodine receptor, the inositol-1,4,5-trisphosphate receptor, or the Ca 2+ -binding protein calreticulin showed suppression of cell death. Lysosome-like gut granules also appeared to be required, because death was suppressed in animals lacking the Rab-like guanosine triphosphatase Glo-1, which is required for formation of these acidic granules. Animals lacking SRP-6 were also more susceptible than wild-type animals to heat shock, hypoxia, or hyperoxia. The authors argue that the common effects of serpins on these stimuli, which cause distinct biochemical alterations to initiate cell death, indicate that there may be a central necrotic death mechanism that can be regulated by serpins. If so, serpins could act as an antidote to cells undergoing damage by such stresses, which occur, for example, during heart attacks. Such a prosurvival function of intracellular serpins may help explain why increased expression of some serpin family members is associated with a poor prognosis in various human cancers. C. J. Luke, S. C. Pak, Y. S. Askew, T. L. Naviglia, D. J. Askew, S. M. Nobar, A. C. Vetica, O. S. Long, S. C. Watkins, D. B. Stolz, R. J. Barstead, G. L. Moulder, D. Brömme, G. A. Silvermann, An intracellular serpin regulates necrosis by inhibiting the induction and sequelae of lysosomal injury. Cell , 130 , 1108-1119 (2007). [PubMed]

Apoptotic and necrotic death mechanisms are concomitantly activated in the same cell after cerebral ischemia.

Both necrotic and apoptotic cell death mechanisms are activated after cerebral ischemia. However, whether they are concomitantly active in the same cell or in discrete cell populations is not known. We investigated activation of both pathways at the cellular level in mice brains subjected to transient or permanent focal ischemia. Four hours after ischemia, diffuse cathepsin-B spillage into cytoplasm, suggesting lysosomal leakage, was observed within neurons immunoreactive for the active form of caspase-3 (p20). Ischemic neurons with a leaky plasma membrane (positive for propidium iodide) were colabeled with caspase-cleaved actin fragment and exhibited TUNEL-positive nuclei having apoptotic morphology. At 72 hours, up to 27% of cells with caspase activity displayed morphological features suggestive of secondary necrosis. These data, demonstrating an early and concurrent increase in caspase-3 and cathepsin-B activities followed by appearance of caspase-cleavage products, DNA fragmentation, and membrane disintegration, suggest that subroutines of necrotic and apoptotic cell death are concomitantly activated in ischemic neurons and that the dominant cell death phenotype is determined by the relative speed of each process.

Necrotic cell death induced by photodynamic treatment of human lung adenocarcinoma A-549 cells with palladium(II)-tetraphenylporphycene.

In this study we describe photodamaging and photokilling effects of palladium(II)-tetraphenylporphycene (PdTPPo) (previously incorporated into dipalmitoylphosphatidylcholine liposomes) on the human lung adenocarcinoma A-549 cell line. No dark cytotoxicity was found when the drug was applied at 10(-6) M or 5 x 10(-7) M for 1 or 18 h, respectively. After 1-h treatment with 10(-7) M or 5 x 10(-7) M PdTPPo followed by red light irradiation for variable times, dose-dependent lethal effects were observed in A-549 cells. Apoptosis was not found after the above photodynamic treatments or under even milder sublethal conditions. In contrast to HeLa cells subjected to PdTPPo photosensitization where either apoptosis or necrosis were induced, morphological analysis and electrophoretical DNA pattern of A-549 cells always revealed a clearly necrotic death mechanism. However, A-549 cells died by apoptosis after serum and L-glutamine deprivation, indicating that only the photodynamically induced apoptosis was inhibited. Immunofluorescent labeling revealed that microtubules and actin microfilaments were immediately and strongly damaged by photodynamic treatments with PdTPPo. No metaphase arrest and/or mitotic alterations were observed after phototreatments. Present results show that the cell type plays a fundamental role in relation to the apoptotic or necrotic response to photosensitization, and that cytoskeletal components are important targets implicated in cell death processes.