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Mitochondria are involved in the development of organ failure in critical care diseases. However, the mechanisms underlying mitochondrial dysfunction are not clear yet. Inducible hemoxygenase (HO-1), a member of the heat shock protein family, is upregulated in critical care diseases and considered to confer cytoprotection against oxidative stress. However, one of the products of HO-1 is Fe 2 þ which multiplies the damaging potential of reactive oxygen species catalyzing Fenton reaction. The aim of this study was to clarify the relevance of free iron metabolism to the oxidative damage of the liver in endotoxic shock and its impact on mitochondrial function. Endotoxic shock in rats was induced by injection of lipopolysaccharide (LPS) at a dose of 8 mg/kg (i.v.). We observed that the pro-inflammatory cytokine TNF-a and the liver necrosis marker aspartate aminotransferase were increased in blood, confirming inflammatory response to LPS and damage to liver tissue, respectively. The levels of free iron in the liver were significantly increased at 4 and 8 h after onset of endotoxic shock, which did not coincide with the decrease of transferrin iron levels in the blood, but rather with expression of the inducible form of heme oxygenase (HO-1). The proteins important for sequestering free iron (ferritin) and the export of iron out of the cells (ferroportin) were downregulated facilitating the accumulation of free iron in cells. The temporarily increased concentration of free iron in the liver correlated with the temporary impairment of both mitochondrial function and tissue ATP levels. Addition of exogenous iron ions to mitochondria isolated from control animals resulted in an impairment of mitochondrial respiration similar to that observed in endotoxic shock in vivo. Our data suggest that free iron released by HO-1 causes mitochondrial dysfunction in pathological situations accompanied by endotoxic shock.

Ginsenosides are a special group of triterpenoid saponins attributed to medical effects of ginseng. Therefore, they have been research targets over the last three decades to explain ginseng actions and a wealth of literature has been presented reporting on ginsenosides' effects on the human body. Recently, there is increasing evidence on beneficial effects of ginsenosides to the central nervous system (CNS). Using a wide range of in vitro and in vivo models, researchers have attributed these effects to specific pharmacological actions of ginsenosides on cerebral metabolism, oxidative stress and radical formation, neurotransmitter imbalance and membrane stabilizing effects, and even antiapoptotic effects. Modulating these particular mechanisms by ginsenosides has thus been reported to exert either general stimulatory effects on the brain functions or protecting the CNS against various disease conditions. In this review, we try to address the recently reported ginsenosides' actions on different CNS targets particularly those supporting possible therapeutic efficacies in CNS disorders and neurodegenerative diseases.

Because different proteins compete for the proton gradient across the inner mitochondrial membrane, an efficient mechanism is required for allocation of associated chemical potential to the distinct demands, such as ATP production, thermogenesis, regulation of reactive oxygen species (ROS), etc. Here, we used the superresolution technique dSTORM (direct stochastic optical reconstruction microscopy) to visualize several mitochondrial proteins in primary mouse neurons and test the hypothesis that uncoupling protein 4 (UCP4) and F 0 F 1 -ATP synthase are spatially separated to eliminate competition for the proton motive force. We found that UCP4, F 0 F 1 -ATP synthase, and the mitochondrial marker voltage-dependent anion channel (VDAC) have various expression levels in different mitochondria, supporting the hypothesis of mitochondrial heterogeneity. Our experimental results further revealed that UCP4 is preferentially localized in close vicinity to VDAC, presumably at the inner boundary membrane, whereas F 0 F 1 -ATP synthase is more centrally located at the cristae membrane. The data suggest that UCP4 cannot compete for protons because of its spatial separation from both the proton pumps and the ATP synthase. Thus, mitochondrial morphology precludes UCP4 from acting as an uncoupler of oxidative phosphorylation but is consistent with the view that UCP4 may dissipate the excessive proton gradient, which is usually associated with ROS production. mitochondrial membrane proteins | proton diffusion | direct stochastic optical reconstruction microscopy | uncoupling | reactive oxygen species M itochondria are involved in a wide range of cell functions, including fatty acid oxidation, calcium homeostasis, apoptosis, reactive oxygen species (ROS) signaling, and above all, production of ATP (1, 2). In neurons, these organelles are transported along neuronal processes to provide energy for areas of high energy demand, such as synapses (3). To support their functions, mitochondria exhibit a complex morphology consisting of separate and functionally distinct outer mitochondrial membrane (OMM) and inner mitochondrial membrane (IMM). The latter is structurally organized into two domains: an inner boundary membrane (IBM) and a cristae membrane (CM) (4). The current hypotheses imply that the morphology/topology of the IMM is tightly related to biochemical function, the energy state, and the pathophysiological state of mitochondria (5). Whereas the OMM contains porins [e.g., voltage-dependent anion channel (VDAC)], which mediate its permeability to molecules up to 10 kDa, the IMM topology is highly complex. It is comprised of different transport proteins, the ATP synthase (complex V), and complexes I, III, and IV of the electron transport chain, which are responsible for generating the proton motive force (pmf); pmf represents the driving force for not only ATP synthesis, but also other protein-mediated transport activities (for example, phosphate, pyruvate, and glutamate transport). Uncoupling protein 1 (UCP1; thermogenin), a memb...

Ginsenosides Rb1 and Rg1 are the main active ingredients of Panax ginseng C.A. Meyer (Araliaceae). They appear to exert protection against ischaemia and anoxic damage in animal models, suggesting an antioxidative and cytoprotective role. In our study, primary cultures from embryonic mouse mesencephalon are applied to examine the effects of these two ginsenosides on neuritic growth of dopaminergic cells and their survival affected by 1-methyl-4-phenylpyridinium-iodide (MPP(+)). Ginsenoside Rb1 (at 10 microM) enhanced the survival of dopaminergic neurons by 19% compared to untreated control. MPP(+) (at 1 microM) significantly reduced the number of dopaminergic neurons and severely affected neuronal processes. Both ginsenosides counteracted these degenerations and significantly protected lengths and numbers of neurites of TH(+) cells. Both compounds however could not prevent the cell loss caused by MPP(+). Our study thus indicates partial neurotrophic and neuroprotective actions of ginsenosides Rb1 and Rg1 in dopaminergic cell culture.

Apart from the first family member, uncoupling protein 1 (UCP1), the functions of other UCPs (UCP2-UCP5) are still unknown. In analyzing our own results and those previously published by others, we have assumed that UCP's cellular expression pattern coincides with a specific cell metabolism and changes if the latter is altered. To verify this hypothesis, we analyzed the expression of UCP1-5 in mouse embryonic stem cells before and after their differentiation to neurons. We have shown that only UCP2 is present in undifferentiated stem cells and it disappears simultaneously with the initiation of neuronal differentiation. In contrast, UCP4 is simultaneously up-regulated together with typical neuronal marker proteins TUJ-1 and NeuN during mESC differentiation in vitro as well as during murine brain development in vivo. Notably, several tested cell lines express UCP2, but not UCP4. In line with this finding, neuroblastoma cells that display metabolic features of tumor cells express UCP2, but not UCP4. UCP2's occurrence in cancer, immunological and stem cells indicates that UCP2 is present in cells with highly proliferative potential, which have a glycolytic type of metabolism as a common feature, whereas UCP4 is strongly associated with non-proliferative highly differentiated neuronal cells.

Thymoquinone is the main active constituent of Nigella sativa seeds with antioxidant and antiinflammatory properties. In the present study, primary dopaminergic cultures from mouse mesencephala were used to investigate the neuroprotective effects of thymoquinone against MPP(+) and rotenone toxicities. MPP(+) (10 microm on day 10 in vitro (i.v.) for 48 h) significantly decreased the number of THir by 40% compared with untreated control cultures. Rotenone at both short (20 nm on day 10 i.v. for 48 h) and long-term (1 nm on day 6 i.v. for 6 consecutive days) toxicities reduced the number of THir neurons by 33% and 24%, respectively. Treatment of cultures with thymoquinone (0.01, 0.1, 1, 10 microm on day 8 i.v. for 4 days) rescued about 25% of THir neurons at concentrations of 0.1 microm and 1 microm against MPP(+)-induced cell death. Against rotenone, thymoquinone afforded significant protection in both short- and long-term models. In short-term rotenone toxicity, thymoquinone (from days 8-12 i.v.) saved about 65%, 74% and 79% of THir neurons at concentrations of 0.01, 0.1 and 1 microm, respectively, compared with cell loss induced by rotenone. In long-term rotenone toxicity, concomitant treatment of cultures with thymoquinone significantly rescued about 83-100% of THir neurons compared with rotenone-treated cultures. In conclusion, the current study presents for the first time the potential of thymoquinone to protect primary dopaminergic neurons against MPP(+) and rotenone relevant to Parkinson's disease.