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General anesthesia induces changes in dendritic spine number and synaptic transmission in developing mice. These changes are rather disturbing, as similar changes are seen in animal models of neurodevelopmental disorders. We previously suggested that mTor-dependent upregulation of mitochondrial function may be involved in such changes. To further understand the significance of mitochondrial changes after general anesthesia during neurodevelopment, we exposed young mice to 2.5 % sevoflurane for 2 h followed by injection of rotenone, a mitochondrial complex I inhibitor. In postnatal day 17 (PND17) mice, intraperitoneal injection of rotenone not only blocked sevoflurane-induced increases in mitochondrial function, it also prevented sevoflurane-induced changes in excitatory synaptic transmission. Interestingly, similar changes were not observed in younger, neonatal mice (PND7). We next assessed whether the mitochondrial unfolded protein response (UPRmt) acted as a link between anesthetic exposure and mitochondrial function. Expression of UPRmt proteins, which help maintain protein-folding homeostasis and increase mitochondrial function, was increased 6 h after sevoflurane exposure. Our results show that a single, brief sevoflurane exposure induces age-dependent changes in mitochondrial function that constitute an important mechanism for the increase in excitatory synaptic transmission in late postnatal mice, and also suggest mitochondria and UPRmt as potential targets for preventing anesthesia toxicity.

Cognitive decline is observed in aging and neurodegenerative diseases, including Alzheimer&rsquo, s disease (AD) and dementia. Intracellular energy produced via mitochondrial respiration is used in the regulation of synaptic plasticity and structure, including dendritic spine length and density, as well as for the release of neurotrophic factors involved in learning and memory. To date, a few synthetic agents for improving mitochondrial function have been developed for overcoming cognitive impairment. However, no natural compounds that modulate synaptic plasticity by directly targeting mitochondria have been developed. Here, we demonstrate that a mixture of Schisandra chinensis extract (SCE) and ascorbic acid (AA) improved cognitive function and induced synaptic plasticity-regulating proteins by enhancing mitochondrial respiration. Treatment of embryonic mouse hippocampal mHippoE-14 cells with a 4:1 mixture of SCE and AA increased basal oxygen consumption rate. We found that mice injected with the SCE-AA mixture showed enhanced learning and memory and recognition ability. We further observed that injection of the SCE-AA mixture in mice significantly increased expression of postsynaptic density protein 95 (PSD95), an increase that was correlated with enhanced brain-derived neurotrophic factor (BDNF) expression. These results demonstrate that a mixture of SCE and AA improves mitochondrial function and memory, suggesting that this natural compound mixture could be used to alleviate AD and aging-associated memory decline.

Recent studies revealed that the inhibition of mitochondrial oxidative phosphorylation (OXPHOS) is coupled with the mitochondrial unfolded protein response, thereby stimulating the secretion of non-cell autonomous factors, which may control systemic energy metabolism and longevity. However, the nature and roles of non-cell autonomous factors induced in adipose tissue in response to reduced OXPHOS function remain to be clarified in mammals. CR6-interacting factor 1 (CRIF1) is an essential mitoribosomal protein for the intramitochondrial production of mtDNA-encoded OXPHOS subunits. Deficiency of CRIF1 impairs the proper formation of the OXPHOS complex, resulting in reduced function. To determine which secretory factors are induced in response to reduced mitochondrial OXPHOS function, we analyzed gene expression datasets in Crif1-depleted mouse embryonic fibroblasts. Crif1 deficiency preferentially increased the expression of angiopoietin-like 6 (Angptl6) and did not affect other members of the ANGPTL family. Moreover, treatment with mitochondrial OXPHOS inhibitors increased the expression of Angptl6 in cultured adipocytes. To confirm Angptl6 induction in vivo, we generated a murine model of reduced mitochondrial OXPHOS function using adipose tissue-specific Crif1-deficient mice and verified the upregulation of Angptl6 and fibroblast growth factor 21 (Fgf21) in white adipose tissue. Treatment with recombinant ANGPTL6 protein increased oxygen consumption and Pparα expression through the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway in cultured adipocytes. Furthermore, the ANGPTL6-mediated increase in Pparα expression resulted in increased FGF21 expression, thereby promoting β-oxidation. In conclusion, mitochondrial OXPHOS function governs the expression of ANGPTL6, which is an essential factor for FGF21 production in adipose tissue and cultured adipocytes.

Current therapeutics for Parkinson&rsquo, s disease (PD) are only effective in providing relief of symptoms such as rigidity, tremors and bradykinesia, and do not exert disease-modifying effects by directly modulating mitochondrial function. Here, we investigated auraptene (AUR) as a potent therapeutic reagent that specifically protects neurotoxin-induced reduction of mitochondrial respiration and inhibits reactive oxygen species (ROS) generation. Further, we explored the mechanism and potency of AUR in protecting dopaminergic neurons. Treatment with AUR significantly increased the viability of substantia nigra (SN)-derived SN4741 embryonic dopaminergic neuronal cells and reduced rotenone-induced mitochondrial ROS production. By inducing antioxidant enzymes AUR treatment also increased oxygen consumption rate. These results indicate that AUR exerts a protective effect against rotenone-induced mitochondrial oxidative damage. We further assessed AUR effects in vivo, investigating tyrosine hydroxylase (TH) expression in the striatum and substantia nigra of MPTP-induced PD model mice and behavioral changes after injection of AUR. AUR treatment improved movement, consistent with the observed increase in the number of dopaminergic neurons in the substantia nigra. These results demonstrate that AUR targets dual pathogenic mechanisms, enhancing mitochondrial respiration and attenuating ROS production, suggesting that the preventative potential of this natural compound could lead to improvement in PD-related neurobiological changes.

Paraquat (PQ), an herbicide considered an environmental contributor to the development of Parkinson’s disease (PD), induces dopaminergic neuronal loss through reactive oxygen species (ROS) production and oxidative stress by mitochondrial complex I. Most patients with PQ-induced PD are affected by chronic exposure and require a preventive strategy for modulation of disease progression. To identify drugs that are effective in preventing PD, we screened more than 1000 drugs that are currently used in clinics and in studies employing PQ-treated cells. Of these, chloramphenicol (CP) showed the most powerful inhibitory effect. Pretreatment with CP increased the viability of PQ-treated SN4741 dopaminergic neuronal cells and rat primary cultured dopaminergic neurons compared with control cells treated with PQ only. CP pretreatment also reduced PQ-induced ROS production, implying that mitochondrial complex I is a target of CP. This effect of CP reflected downregulation of the mitochondrial complex I subunit ND1 and diminished PQ recycling, a major mechanism of ROS production, and resulted in the prevention of cell loss. Notably, these effects of CP were not observed in rotenone-pretreated SN4741 cells and Rho-negative cells, in which mitochondrial function is defective. Consistent with these results, CP pretreatment of MPTP-treated PD model mice also ameliorated dopaminergic neuronal cell loss. Our findings indicate that the inhibition of mitochondrial complex I with CP protects dopaminergic neurons and may provide a strategy for preventing neurotoxin-induced PD.

The identification of large numbers of genetic mutations in immature myeloid cells has made it difficult to identify specific targets for acute myeloid leukemia (AML) therapy. Although current pharmacological targets for controlling cancer are focused on identifying genetic mutations, it is hard to develop the specific drugs to achieve complete remission due to complex and variable genetic mutations. To overcome the failure of the genetic mutation theory, the present study targeted mitochondrial metabolism as a strategy for inducing anti‑leukemic activity, based on evidence that AML cells have an abnormally high amount of mitochondria and that somatic mutations can alter metabolic flux in cancer. It was found that L‑deprenyl, which is clinically available for the treatment of Parkinson's disease, exerts anti‑mitochondria activity in KG‑1α cells, as assessed by detection of oxygen consumption rate (OCR) and extracellular acidification (ECAR) using XF analyzer, respectively. Using a luciferase assay for detecting adenosine triphosphate (ATP) content, it was found that suppression of mitochondrial activity led to ATP depletion and was associated with potent cytotoxic activity. L‑deprenyl is known to target monoamine oxidase‑B (MAO‑B) on the outer membrane of mitochondria, therefore, the activity of MAO‑A and ‑B was measured based on the fluorometric detection of H2O2 produced by the enzyme reaction. Notably, MAO‑A and -B activity was low in AML cells and the present findings suggested that the anticancer effect of L‑deprenyl was independent of MAO‑B. Change of mitochondrial respiration‑ and glycolysis‑related gene expression levels were measured by reverse transcription‑quantitative polymerase chain reaction. Consistent with the aforementioned results, treatment with L‑deprenyl reduced the mRNA level of mitochondrial respiration‑ and glycolysis‑related genes. Collectively, the present results identify L‑deprenyl as a novel candidate for the treatment of AML through inhibition of mitochondrial respiration.

Renal cell carcinoma (RCC) progression resulting from the uncontrolled migration and enhanced angiogenesis is an obstacle to effective therapeutic intervention. Tumor metabolism has distinctive feature called Warburg effect, which enhances the aerobic glycolysis rapidly supplying the energy for migration of tumor. To manipulate this metabolic change characteristic of aggressive tumors, we utilized the citrus extract, auraptene, known as a mitochondrial inhibitor, testing its anticancer effects against the RCC4 cell line. We found that auraptene impaired RCC4 cell motility through reduction of mitochondrial respiration and glycolytic pathway-related genes. It also strongly disrupted VEGF-induced angiogenesis in vitro and in vivo. Hypoxia-inducible factor 1a (HIF-1a), a key regulator of cancer metabolism, migration and angiogenesis that is stably expressed in RCCs by virtue of a genetic mutation in the von Hippel-Lindau (VHL) tumor-suppressor protein, was impeded by auraptene, which blocked HIF-1a translation initiation without causing cytotoxicity. We suggest that blockade HIF-1a and reforming energy metabolism with auraptene is an effective approach for suspension RCC progression.

Mitochondrial dysfunction has emerged as a major contributing factor to endothelial dysfunction and vascular disease, but the key mechanisms underlying mitochondrial dysfunction-induced endothelial dysfunction remain to be elucidated. In this study, we aim at determining whether mitochondrial dysfunction in endothelial cells plays a key role in vascular disease, by examining the phenotype of endothelial-specific CR6-interacting factor 1 (CRIF1) knockout mice. We also used siRNA-mediated downregulation of CRIF1 gene in the endothelial cells to study about the in vitro pathophysiological underlying mechanisms.Downregulation of CRIF1 in endothelial cells caused disturbances of mitochondrial oxidative phosphorylation complexes and membrane potential, leading to enhanced mitochondrial reactive oxygen species production. Gene silencing of CRIF1 results in decreased SIRT1 expression along with increased endothelial nitric oxide synthase (eNOS) acetylation, leading to reduced nitric oxide production both in vitro and in vivo. Endothelium-dependent vasorelaxation of aortic rings from CRIF1 knockout (KO) mice was considerably less than in wild-type mice, and it was partially recovered by Sirt1 overexpression in CRIF1 KO mice.Our results show for the first time a relationship between mitochondrial dysfunction and impaired vascular function induced in CRIF1 deficiency conditions and also the possible underlying pathway involved.These findings indicate that CRIF1 plays an important role in maintaining mitochondrial and endothelial function through its effects on the SIRT1-eNOS pathway. Antioxid. Redox Signal. 27, 234-249.

Background Anesthesia during the synaptogenic period induces dendritic spine formation, which may affect neurodevelopment. The authors, therefore, evaluated whether changes in synaptic transmission after dendritic spine formation induced by sevoflurane were associated with long-term behavioral changes. The effects of sevoflurane on mitochondrial function were also assessed to further understand the mechanism behind spinogenesis. Methods Postnatal day 16 to 17 mice were exposed to sevoflurane (2.5% for 2 h), and synaptic transmission was measured in the medial prefrontal cortex 6 h or 5 days later. The expression of postsynaptic proteins and mitochondrial function were measured after anesthesia. Long-term behavioral changes were assessed in adult mice. Results Sevoflurane increased the expression of excitatory postsynaptic proteins in male and female mice (n = 3 to 5 per group). Sevoflurane exposure in male mice transiently increased miniature excitatory postsynaptic current frequency (control: 8.53 ± 2.87; sevoflurane: 11.09 ± 2.58) but decreased miniature inhibitory postsynaptic current frequency (control: 10.18 ± 4.66; sevoflurane: 6.88 ± 2.15). Unexpectedly, sevoflurane increased miniature inhibitory postsynaptic current frequency (control: 1.81 ± 1.11; sevoflurane: 3.56 ± 1.74) in female mice (neurons, n = 10 to 21 per group). Sevoflurane also increased mitochondrial respiration in male mice (n = 5 to 8 per group). However, such changes from anesthesia during the critical period did not induce long-term behavioral consequences. Values are presented as mean ± SD. Conclusions Sevoflurane exposure during the critical period induces mitochondrial hyperactivity and transient imbalance of excitatory/inhibitory synaptic transmission, without long-lasting behavioral consequences. Further studies are needed to confirm sexual differences and to define the role of mitochondrial activity during anesthesia-induced spine formation.

There are two causes of Parkinson's disease (PD): environmental insults and genetic mutations of PD-associated genes. Environmental insults and genetic mutations lead to mitochondrial dysfunction, and a combination of mitochondrial dysfunction and increased oxidative stress in dopaminergic neurons is thought to contribute to the pathogenesis of PD. Among the PD-associated genes, DJ-1 acts as a redox sensor for oxidative stress and has been also proposed to maintain mitochondrial complex I activity. To understand molecular functions of DJ-1 in the cell, we have generated DJ-1 null cells from the DJ-1(-/-) mouse embryos. Using these null cells, we investigated the susceptibility to an environmental toxin, paraquat, which is known to inhibit mitochondrial complex I. Interestingly, we found that DJ-1 null cells showed a resistance to paraquat-induced apoptosis, including reduced poly (ADP-ribose) polymerase and procaspase-3. Also DJ-1 null cells generated less superoxide than SN4741 cells by paraquat treatment. Consistent with the reduced paraquat sensitivity, DJ-1 null cells showed reduced complex I activity, which was partially rescued by ectopic DJ-I expression. In summary, our results suggest that DJ-1 is critical to maintain mitochondrial complex I and complex I could be a key target in interaction of paraquat toxicity and DJ-1 for giving rise to PD.

Context: The oncogenic BRAFV600E mutation results in an active structural conformation characterized by greatly elevated ERK activity. However, additional cellular effects caused by subcellular action of BRAFV600E remain to be identified. Objective: To explore these effects, differences in the subcellular localization of wild-type and mutant BRAF in thyroid cancer were investigated. Results: A significant proportion of endogenous and exogenous BRAFV600E, but not wild-type BRAF, was detected in the mitochondrial fraction, similar to other BRAF mutants including BRAFV600D, BRAFV600K, BRAFV600R, and BRAFG469A, which showed elevated kinase activity and mitochondrial localization. Induced expression of BRAFV600E suppressed the apoptotic responses against staurosporine and TNFα/cycloheximide. Interestingly, the mitochondrial localization and antiapoptotic activities of BRAFV600E were unaffected by sorafenib and U0126 suppression of MAPK kinase (MEK) and ERK activities. Similarly, although the RAF inhibitor sorafenib effectively inhibited MEK/ERK activation, it did not block the mitochondrial localization of BRAFV600E. In addition, inducible expression of BRAFV600E increased the glucose uptake rate and decreased O2 consumption, suggesting that BRAFV600E reduces mitochondrial oxidative phosphorylation, a signature feature of cancer cells. Again, these metabolic alterations resulted by BRAFV600E expression were not affected by the treatment of thyroid cells by sorafenib. Therefore, RAF and MEK inhibitors are unable to block the antiapoptotic activity of BRAFV600E or correct the high glucose uptake rate and glycolytic activity and suppressed mitochondrial oxidative phosphorylation induced by BRAFV600E. Conclusions: The mitochondrial localization observed in oncogenic BRAF mutants might be related to their altered responses to apoptotic stimuli and characteristic metabolic phenotypes found in thyroid cancer. The inability of MEK and RAF inhibitors, U0126 and sorafenib, respectively, to block the mitochondrial localization of BRAFV600E has additional therapeutic implications for BRAFV600E-positive thyroid cancers.

Among the primary brain tumors, glioblastoma multiforme (GBM) has a radical proliferation ability that complicates the therapeutic modulation of cancer progression. The majority of GBM patients have a low survival rate (

Impaired mitochondrial oxidative phosphorylation (OXPHOS) has been proposed as an etiological mechanism underlying insulin resistance. However, the initiating organ of OXPHOS dysfunction during the development of systemic insulin resistance has yet to be identified. To determine whether adipose OXPHOS deficiency plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with OXPHOS–deficient adipose tissue was examined. Crif1 is a protein required for the intramitochondrial production of mtDNA–encoded OXPHOS subunits; therefore, Crif1 haploinsufficient deficiency in mice results in a mild, but specific, failure of OXPHOS capacity in vivo. Although adipose-specific Crif1-haploinsufficient mice showed normal growth and development, they became insulin-resistant. Crif1-silenced adipocytes showed higher expression of chemokines, the expression of which is dependent upon stress kinases and antioxidant. Accordingly, examination of adipose tissue from Crif1-haploinsufficient mice revealed increased secretion of MCP1 and TNFα, as well as marked infiltration by macrophages. These findings indicate that the OXPHOS status of adipose tissue determines its metabolic and inflammatory responses, and may cause systemic inflammation and insulin resistance., Author Summary Type 2 diabetes is one of the most challenging health problems in the 21st century. Although insulin resistance is regarded as a fundamental defect that precedes the development of type 2 diabetes, the nature and cause of insulin resistance remain unknown. Adipose tissue is an important organ that determines whole-body energy metabolism, and its dysfunction is a critical element in the development of systemic insulin resistance. Adipose mitochondrial function is suppressed in the insulin-resistant state, and increased adipose mitochondrial biogenesis is associated with the reversal of insulin resistance by a PPARγ agonist. However, despite these important observations, little is known about how mitochondrial respiratory dysfunction in white adipose tissue (WAT) causes insulin resistance. To determine whether adipose deficiency of mitochondrial respiratory capacity plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with mitochondrial OXPHOS (oxidative phosphorylation)–deficient adipose tissue was examined. Crif1 is a protein required for the translation of mtDNA–encoded OXPHOS subunits. Interestingly, mice haploinsufficient for Crif1 in adipose tissue showed reduced OXPHOS capacity and developed marked insulin resistance.

Adipocyte differentiation requires the coordinated activities of several nuclear transcription factors. Recently, mitochondria biogenesis was reported to occur during adipocyte differentiation and following treatment with thiazolidinediones in vitro and in vivo. Crif1 is a translational factor for mitochondrial DNA (mtDNA) and is important for transcription of the mitochondrial oxidative phosphorylation (OXPHOS) complex. To investigate the role of OXPHOS in adipogenesis, we analyzed adipocyte differentiation following disruption of Crif1 in vitro and in vivo. The adipose-specific Crif1 knockout mouse had a lower body weight and less fat mass than wild-type mice. Furthermore, adipocytes were smaller and had a dysplastic morphology in the adipose-specific Crif1 knockout mouse. 3T3-L1 adipocytes or adipose-derived stem cells (ADSCs) that lacked Crif1 expressed lower levels of mtDNA-encoded OXPHOS subunits, and adipocyte differentiation was disrupted. Rosiglitazone treatment did not induce adipogenesis or mitochondria biogenesis in Crif1 knockout ADSCs. These results show that mitochondrial OXPHOS and Crif1 are required for rosiglitazone- and hormone-induced adipogenesis.