Role Of Mitochondrial Changes Research Articles

Mitochondrial Dysfunction in Rabies Virus-Infected Human and Canine Brains.

Rabies is a fatal encephalitis caused by the Rabies lyssavirus (RABV). The presence of minimal neuropathological changes observed in rabies indicates that neuronal dysfunction, rather than neuronal death contributes to the fatal outcome. The role of mitochondrial changes has been suggested as a possible mechanism for neuronal dysfunction in rabies. However, these findings are mostly based on studies that have employed experimental models and laboratory-adapted virus. Studies on brain tissues from naturally infected human and animal hosts are lacking. The current study investigated the role of mitochondrial changes in rabies by morphological, biochemical and proteomic analysis of RABV-infected human and canine brains. Morphological analysis showed minimal inflammation with preserved neuronal and disrupted mitochondrial structure in both human and canine brains. Proteomic analysis revealed involvement of mitochondrial processes (oxidative phosphorylation, cristae formation, homeostasis and transport), synaptic proteins and autophagic pathways, with over-expression of subunits of mitochondrial respiratory complexes. Consistent with these findings, human and canine brains displayed elevated activities of complexes I (p < 0.05), IV (p < 0.05) and V (p < 0.05). However, this did not result in elevated ATP production (p < 0.0001), probably due to lowered mitochondrial membrane potential as noted in RABV-infected cells in culture. These could lead to mitochondrial dysfunction and mitophagy as indicated by expression of FKBP8 (p < 0.05) and PINK1 (p < 0.001)/PARKIN (p > 0.05) and ensuing autophagy, as shown by the status of LCIII (p < 0.05), LAMP1 (p < 0.001) and pertinent ultrastructural markers. We propose that altered mitochondrial bioenergetics and cristae architecture probably induce mitophagy, leading to autophagy and consequent neuronal dysfunction in rabies.

Experimental Apoptosis Provides Clues about the Role of Mitochondrial Changes in Neuronal Death

A quantitative morphometric study has been carried out in human neuroblastoma SK-N-BE cells to evaluate the ultrastructural features and the metabolic efficiency of mitochondria involved in the early steps of apoptosis. In mitochondria from control and apoptotic cells cytochrome oxidase (COX) activity was estimated by preferential cytochemistry. Number of mitochondria (numeric density: Nv), volume fraction occupied by mitochondria/microm3 of cytoplasm (volume density: Vv), and average mitochondrial volume (V) were calculated for both COX-positive and -negative organelles. The ratio (R) of the cytochemical precipitate area to the overall area of each mitochondrion was evaluated on COX-positive organelles to estimate the inner mitochondrial membrane fraction actively involved in cellular respiration. Following apoptotic stimulus, the whole mitochondrial population showed a significant increase of Nv and Vv, while V was significantly decreased. In COX-positive organelles higher values of Nv were found, V appeared significantly reduced, and Vv was unchanged. R was increased at a nonsignificant extent in apoptotic cells. COX-positive mitochondria accounted for 21% and 35% of the whole population in control and in apoptotic cells, respectively. These findings document that in the early stages of apoptosis the increased fraction of small mitochondria provides an adequate amount of ATP for progression of the programmed cell death and these more efficient organelles appear to represent a reactive response to the loss of metabolically impaired mitochondria. A better understanding of the mitochondrial role in neuronal apoptosis may suggest potential interventions to prevent the extensive nerve cell death typical of neurodegenerative diseases.

Induction of apoptosis by heat and γ-radiation in a human lymphoid cell line; role of mitochondrial changes and caspase activation

Purpose: The aim of the study was to investigate the molecular mechanisms involved in apoptosis of human promyelocytic cells (HL60) induced by hyperthermia and to compare this to radiation-induced apoptosis as a reference model.Materials and methods: Apoptosis of HL60 cells was induced by heat-treatment (43°C during 1 h) or by γ-radiation (8 Gy) and followed at increasing time periods after treatment with Annexin V binding to phosphatidylserine (PS). The transition of the mitochondrial membrane potential (Δψm) was estimated by the extent of mitochondrial JC-1 uptake. Bcl-2 and Bax protein expression levels were monitored using fluorescent-labelled antibodies. Caspase activation was studied using a fluorochrome-labelled pan-caspase inhibitor (FLICA), which also allowed one to study the kinetics of the apoptotic cascade.Results: After heat-treatment or irradiation of HL60 cells, a decreased Δψm as well as PS membrane expression were detectable after 8 h. Bcl-2 and Bax protein expression levels were decreased and increased, respectively, 1 h after heat-treatment or irradiation. The apoptotic rate of HL60 cells, as measured by the FLICA binding, was faster with heat-treatment as compared to γ-irradiation. Addition of a pan-caspase inhibitor prevented PS externalization after heat-treatment but not after irradiation. The presence of a pan-caspase inhibitor did not influence the decrease of Δψm both after heat-treatment and γ-irradiation. However, the addition of the specific caspase-2 inhibitor zVDVAD-fmk prevented the mitochondrial breakdown after heat-treatment. Inhibition of caspase-2 had no effect on the γ-irradiation induced apoptosis.Conclusion: These results suggest that the commitment to apoptosis in HL60 cells after heat-treatment is started by mitochondrial membrane transition involving the Bcl-2 family members and is mainly executed in a caspase-dependent pathway. The results suggest that caspase-2 plays a key role in the heat-induced apoptosis.