ABSTRACT
Recent studies have revealed that endoplasmic reticulum (ER) disturbance is involved in the pathophysiology of neurodegenerative disorders, contributing to the activation of the ER stress-mediated apoptotic pathway. Therefore, we investigated here the molecular mechanisms underlying the ER-mitochondria axis, focusing on calcium as a potential mediator of cell death signals. Using NT2 cells treated with brefeldin A or tunicamycin, we observed that ER stress induces changes in the mitochondrial function, impairing mitochondrial membrane potential and distressing mitochondrial respiratory chain complex Moreover, stress stimuli at ER level evoked calcium fluxes between ER and mitochondria. Under these conditions, ER stress activated the unfolded protein response by an overexpression of GRP78, and also caspase-4 and-2, both involved upstream of caspase-9. Our findings show that ER and mitochondria interconnection plays a prominent role in the induction of neuronal cell death under particular stress circumstances.
Subject(s)
Caspases/metabolism , Endoplasmic Reticulum/metabolism , Mitochondria/enzymology , Neurons/enzymology , Oxidative Stress , Blotting, Western , Cell Line, Tumor , Endoplasmic Reticulum Chaperone BiP , Enzyme Activation , HumansABSTRACT
Mitochondria likely play a role in Parkinson's disease (PD) neurodegeneration. We modelled PD by creating cytoplasmic hybrid (cybrid) cell lines in which endogenous mitochondrial DNA (mtDNA) from PD or control subject platelets was expressed within human teratocarcinoma (NT2) cells previously depleted of endogenous mtDNA. Complex I activity was reduced in both PD cybrid lines and in the platelet mitochondria used to generate them. Under basal conditions PD cybrids had less ATP, more LDH release, depolarized mitochondria, less mitochondrial cytochrome c, and higher caspase 3 activity. Equivalent MPP+ exposures are more likely to trigger programmed cell death in PD cybrid cells than in control cybrid cells. Our data support a relatively upstream role for mitochondrial dysfunction in idiopathic PD.
Subject(s)
Mitochondria/metabolism , Parkinson Disease/metabolism , 1-Methyl-4-phenylpyridinium/pharmacology , Adenosine Triphosphate/analysis , Aged , Blood Platelets/metabolism , Case-Control Studies , Caspase 3/metabolism , Cell Line, Tumor , Cell Nucleus/metabolism , Cell Survival/drug effects , Citrate (si)-Synthase/analysis , Cytochromes c/analysis , DNA, Mitochondrial/metabolism , Dose-Response Relationship, Drug , Electron Transport Complex IV/analysis , Enzyme Activation/drug effects , Humans , Hybrid Cells , L-Lactate Dehydrogenase/analysis , Membrane Potential, Mitochondrial/drug effects , Middle Aged , Mitochondria/enzymology , Mitochondria/genetics , Mitochondria/ultrastructure , Neurons/pathology , Parkinson Disease/genetics , Teratocarcinoma/pathologyABSTRACT
MPP+ (1-methyl-4-phenylpyridium ion), a complex I - inhibiting metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), causes anatomic-specific neurodegeneration. To evaluate the broader role of mitochondria in MPP+-induced cell death, we exposed neuron-like NT2 human teratocarcinoma cells with mtDNA rho+ and without mtDNA (rho0) to MPP+. MPP+ minimized the ability of both rho+ and rho0 cells to reduce MTT. Only rho+ cells, though, initiated intrinsic pathway-mediated apoptosis. MPP+ also activated calpains in both rho+ and rho0 cell lines. The calpain inhibitor MDL 28710 was able to prevent the MPP+-related MTT reduction change in rho0 but not rho+ cells. We conclude that 1) MPP+-induced apoptosis requires functional mitochondria, 2) MPP+ activates calpains independent of respiratory chain inhibition, and 3) calpain activation mediates some aspects of MPP+ toxicity.
