Microcirculatory, mitochondrial, and histological changes following cerebral ischemia in swine.

BACKGROUND: Ischemic brain injury due to stroke and/or cardiac arrest is a major health issue in modern society requiring urgent development of new effective therapies. The aim of this study was to evaluate mitochondrial, microcirculatory, and histological changes in a swine model of global cerebral ischemia. RESULTS: In our model, significant microcirculatory changes, but only negligible histological cell alterations, were observed 3 h after bilateral carotid occlusion, and were more pronounced if the vascular occlusion was combined with systemic hypotension. Analysis of mitochondrial function showed that LEAK respiration (measured in the presence of pyruvate + malate but without ADP) was not affected in any model of global cerebral ischemia in pigs. The OXPHOS capacity with pyruvate + malate as substrates decreased compared with the control levels after bilateral carotid artery occlusion, and bilateral carotid artery occlusion + hypotension by 20% and 79%, respectively, resulting in decreases in the respiratory control index of 14% and 73%, respectively. OXPHOS capacity with succinate as a substrate remained constant after unilateral carotid artery occlusion or bilateral carotid artery occlusion, but decreased by 53% after bilateral carotid artery occlusion and hypotension compared with controls (p < 0.05, n = 3-6). Addition of exogenous cytochrome c to mitochondria isolated from ischemia brains had no effect on respiration in all models used in this study. CONCLUSIONS: We found a decrease in microcirculation and mitochondrial oxidative phosphorylation activity, but insignificant neuronal death, after 3 h ischemia in all our pig models of global cerebral ischemia. Dysfunction of the mitochondrial oxidative phosphorylation system, particularly damage to complex I of the respiratory chain, may be the primary target of the ischemic insult, and occurs before signs of neuronal death can be detected.

Effects of standardized extract of Ginkgo biloba leaves EGb761 on mitochondrial functions: mechanism(s) of action and dependence on the source of mitochondria and respiratory substrate.

In this work, the effects of standardized dry extract of Ginkgo biloba leaves, EGb761, on the respiration of rat heart and liver mitochondria were investigated. We revealed uncoupling of oxidative phosphorylation in rat heart mitochondria by EGb761 which was not observed in liver mitochondria respiring on pyruvate + malate; oxidation of succinate in heart mitochondria was inhibited by EGb761, concentration-dependently, almost completely at 1.00 mg/mL. Uncoupling effect of EGb761 was found to be due to increase in H(+) and K(+) permeability of inner membrane of mitochondria which is most likely to be mediated by the ATP/ADP-translocator and uncoupling proteins. EGb761 depressed State 3 respiration with pyruvate + malate (similarly in heart and liver mitochondria) and succinate (stronger than with pyruvate + malate) but not respiratory chain Complex IV; inhibition of respiration was not restored by uncoupler indicating the inhibitory action of EGb761 on the respiratory complexes preceding to Complex IV and/or on the substrate transport. Moreover, EGb761 rapidly reduced pure cytochrome c. This property of EGb761 together with the observed uncoupling of oxidative phosphorylation and reduction of H2O2 accumulation may be beneficial for the cell in the prevention of apoptosis and protection of cellular functions in pathological situations.

Antibodies bound to Aß oligomers potentiate the neurotoxicity of Aß by activating microglia.

Beta amyloid (Aß) oligomers are thought to contribute to the pathogenesis of Alzheimer's disease. However, clinical trials using Aß immunization were unsuccessful due to strong brain inflammation, the mechanisms of which are poorly understood. In this study we tested whether monoclonal antibodies to oligomeric Aß would prevent the neurotoxicity of Aß oligomers in primary neuronal-glial cultures. However, surprisingly,the antibodies dramatically increased the neurotoxicity of Aß. Antibodies bound to monomeric Aß fragments were non-toxic to cultured neurons, while antibodies to other oligomeric proteins: hamster polyomavirus major capsid protein, human metapneumovirus nucleocapsid protein, and measles virus nucleocapsid protein, strongly potentiated the neurotoxicity of their antigens. The neurotoxicity of antibody-antibody oligomeric antigen complexes was abolished by removal of the Fc region from the antibodies or by removal of microglia from cultures, and was accompanied by inflammatory activation and proliferation of the microglia in culture. In conclusion, we find that immune complexes formed by Aß oligomers or other oligomeric/multimeric antigens and their specific antibodies can cause death and loss of neurons in primary neuronal-glial cultures via Fc-dependent microglial activation. The results suggest that therapies resulting in antibodies to oligomeric Aß or oligomeric brain virus proteins should be used with caution or with suppression of microglial activation.

Influence of ethanol extract of Ginkgo biloba leaves on the isolated rat heart work and mitochondria functions.

In this study, we attempted to elucidate whether the effects of ethanol extract of Ginkgo biloba leaves (GBE) observed previously on isolated rat heart mitochondria may be realized in situ (in case of isolated heart perfused under normal conditions and under ischemia-reperfusion). We found that GBE at low concentrations (0.01, 0.05, and 0.1 µL/mL) does not affect the heart rate and parameters of electrocardiogram (ECG) but produces a small increase in the coronary flow. Higher concentration of GBE (0.2 and 0.3 µL/mL) diminished the heart rate, decreased the coronary flow, and tended to enhance the parameters of ECG. The contractility of isolated rat heart and mitochondrial nicotinamide adenine dinucleotide reduced form fluorescence decreased in a GBE concentration-dependent manner. Mitochondria isolated from hearts pre-perfused with GBE (0.05 µL/mL) for 20 minutes before nonflow global ischemia-reperfusion (45 min/15 min) showed higher respiratory rates with pyruvate + malate in state 2 and state 3, higher respiratory control index, and diminished H2O2 generation compared with untreated group. Higher GBE concentration, 0.4 µL/mL, had no effect on H2O2 generation and did not prevent the ischemia-reperfusion-induced decrease of pyruvate + malate oxidation in state 3 but even enhanced it. However, in the case of nonischemic perfusions, this GBE concentration had no significant effect on these parameters of respiratory functions of isolated heart mitochondria.

Effects of Ginkgo biloba extract on heart and liver mitochondrial functions: mechanism(s) of action.

Though extracts of Ginkgo biloba leaves (GBE) have a wide pharmacological application, little is known about GBE effects on mitochondria. In this work, effects of ethanolic GBE on the respiration of isolated rat heart and liver mitochondria were investigated. We found that GBE stimulates the pyruvate + malate-dependent State 2 respiration of heart mitochondria and decreases mitochondrial membrane potential. Uncoupling effect of GBE was found to be due to its protonophoric action and is likely to be mediated by the ATP/ADP-translocator and uncoupling proteins. The effect of GBE was less in liver than in heart mitochondria. State 3 respiration of heart mitochondria was slightly stimulated at low and depressed at higher GBE concentrations. Inhibition of State 3 respiration of heart mitochondria was not relieved by uncoupler indicating that GBE may inhibit the respiratory chain complexes or the substrate transport. However, Complex IV of the respiratory chain was not inhibited by GBE. H(2)O(2) generation was attenuated by low concentration of GBE probably due to mild uncoupling. The data suggest that mild but not severe uncoupling activity of GBE may be important in providing pharmacological protection of cellular functions in pathological situations.