Mitochondrial effects of dexamethasone imply both membrane and cytosolic-initiated pathways in HepG2 cells

Glucocorticoid treatment is often linked to increased whole-body energy expenditure and hypermetabolism. Glucocorticoids affect mitochondrial energy production, notably in the liver, where they lead to mitochondrial uncoupling reducing the efficacy of oxidative phosphorylation. However, the signaling pathways involved in these phenomena are poorly understood. Here we treated HepG2 cells with dexamethasone for different times and, by using different combinations of inhibitors, we showed that dexamethasone treatment leads to recruitment of two main signaling pathways. The first one involves a G-protein coupled membrane glucocorticoid binding site and rapidly decreases complexes I and II activities while complex III activity is upregulated in a p38MAPK dependent mechanism. The second one implies the classical cytosolic glucocorticoid receptor and triggers long-term transcriptional increases of respiration rates and of complex IV activity and quantity. We concluded that mitochondria are the target of multiple dexamethasone-induced regulatory pathways that are set up gradually after the beginning of hormone exposure and that durably influence mitochondrial oxidative phosphorylation

Iron deficiency without anemia is responsible for decreased left ventricular function and reduced mitochondrial complex I activity in a mouse model

BACKGROUND: Iron deficiency (ID), with or without anemia, is frequent in heart failure patients, and iron supplementation improves patient condition. However, the link between ID (independently of anemia) and cardiac function is poorly understood, but could be explained by an impaired mitochondrial metabolism. Our aim was to explore this hypothesis in a mouse model. METHODS AND RESULTS: We developed a mouse model of ID without anemia, using a blood withdrawal followed by 3-weeks low iron diet. ID was confirmed by low spleen, liver and heart iron contents and the repression of HAMP gene coding for hepcidin. ID was corrected by a single ferric carboxymaltose (FCM) injection (ID + FCM mice). Hemoglobin levels were similar in ID, ID + FCM and control mice. ID mice had impaired physical performances and left ventricular function (echocardiography). Mitochondrial complex I activity of cardiomyocytes was significantly decreased in ID mice, but not complexes II, III and IV activities. ID + FCM mice had improved physical performance, cardiac function and complex I activity compared to ID mice. Using BN-PAGE, we did not observe complex I disassembly, but a reduced quantity of the whole enzyme complex I in ID mice, that was restored in ID + FCM mice. CONCLUSIONS: ID, independently of anemia, is responsible for a decreased left ventricular function, through a reduction in mitochondrial complex I activity, probably secondary to a decrease in complex I quantity. These abnormalities are reversed after iron treatment, and may explain, at least in part, the benefit of iron supplementation in heart failure patients with ID

Estrogen-related receptor α and PGC-1-related coactivator constitute a novel complex mediating the biogenesis of functional mitochondria

Mitochondrial biogenesis, which depends on nuclear as well as mitochondrial genes, occurs in response to increased cellular ATP demand. The nuclear transcriptional factors, estrogen-related receptor α (ERRα) and nuclear respiratory factors 1 and 2, are associated with the coordination of the transcriptional machinery governing mitochondrial biogenesis, whereas coactivators of the peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1) family serve as mediators between the environment and this machinery. In the context of proliferating cells, PGC-1-related coactivator (PRC) is a member of the PGC-1 family, which is known to act in partnership with nuclear respiratory factors, but no functional interference between PRC and ERRα has been described so far. We explored three thyroid cell lines, FTC-133, XTC.UC1 and RO 82 W-1, each characterized by a different mitochondrial content, and studied their behavior towards PRC and ERRα in terms of respiratory efficiency. Overexpression of PRC and ERRα led to increased respiratory chain capacity and mitochondrial mass. The inhibition of ERRα decreased cell growth and respiratory chain capacity in all three cell lines. However, the inhibition of PRC and ERRα produced a greater effect in the oxidative cell model, decreasing the mitochondrial mass and the phosphorylating respiration, whereas the nonphosphorylating respiration remained unchanged. We therefore hypothesize that the ERRα–PRC complex plays a role in arresting the cell cycle through the regulation of oxidative phosphorylation in oxidative cells, and through some other pathway in glycolytic cells

Phenotypic spectrum of MFN2 mutations in the Spanish population

INTRODUCTION: The most common form of axonal Charcot-Marie-Tooth (CMT) disease is type 2A, caused by mutations in the mitochondrial GTPase mitofusin 2 (MFN2). OBJECTIVE: The objective of our study is to establish the incidence of MFN2 mutations in a cohort of Spanish patients with axonal CMT neuropathy. MATERIAL AND METHODS: Eighty-five families with suspected axonal CMT were studied. All MFN2 exons were studied through direct sequencing. A bioenergetics study in fibroblasts was conducted using a skin biopsy taken from a patient with an Arg468His mutation. RESULTS: Twenty-four patients from 14 different families were identified with nine different MFN2 mutations (Arg94Trp, Arg94Gln, Ile203Met, Asn252Lys, Gln276His, Gly296Arg, Met376Val, Arg364Gln and Arg468His). All mutations were found in the heterozygous state and four of these mutations had not been described previously. MFN2 mutations were responsible for CMT2 in 16% +/- 7% of the families studied and in 30.8 +/- 14.2% (12/39) of families with known dominant inheritance. The bioenergetic studies in fibroblasts show typical results of MFN2 patients with a mitochondrial coupling defect (ATP/O) and an increase of the respiration rate linked to complex II. CONCLUSION: It is concluded that mutations in MFN2 are the most frequent cause of CMT2 in this region. The Arg468His mutation was the most prevalent (6/14 families), and our study confirms that it is pathological, presenting as a neuropathy in a mild to moderate degree. This study also demonstrates the value of MFN2 studies in cases of congenital axonal neuropathy, especially in cases of dominant inheritance, severe clinical symptoms or additional symptoms such as optic atrophy

Pro-oxidant effect of ALA is implicated in mitochondrial dysfunction of HepG2 cells

Heme biosynthesis begins in the mitochondrion with the formation of delta-aminolevulinic acid (ALA). In acute intermittent porphyria, hereditary tyrosinemia type I and lead poisoning patients, ALA is accumulated in plasma and in organs, especially the liver. These diseases are also associated with neuromuscular dysfunction and increased incidence of hepatocellular carcinoma. Many studies suggest that this damage may originate from ALA-induced oxidative stress following its accumulation. Using the MnSOD as an oxidative stress marker, we showed here that ALA treatment of cultured cells induced ROS production, increasing with ALA concentration. The mitochondrial energetic function of ALA-treated HepG2 cells was further explored. Mitochondrial respiration and ATP content were reduced compared to control cells. For the 300 μM treatment, ALA induced a mitochondrial mass decrease and a mitochondrial network imbalance although neither necrosis nor apoptosis were observed. The up regulation of PGC-1, Tfam and ND5 genes was also found; these genes encode mitochondrial proteins involved in mitochondrial biogenesis activation and OXPHOS function. We propose that ALA may constitute an internal bioenergetic signal, which initiates a coordinated upregulation of respiratory genes, which ultimately drives mitochondrial metabolic adaptation within cells. The addition of an antioxidant, Manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), resulted in improvement of maximal respiratory chain capacity with 300 μM ALA. Our results suggest that mitochondria, an ALA-production site, are more sensitive to pro-oxidant effect of ALA, and may be directly involved in pathophysiology of patients with inherited or acquired porphyria

3D propagation of the shock-induced vibrations through the whole lower-limb during running

Shock-induced vibrations to the feet have been related to the feel of comfort, the biomechanical control of performance, and the risk of fatigue or injury. Up to recently, the complexity of measuring the human biodynamic response to vibration exposure implied to focus most of the research on the axial acceleration at the tibia. Using wireless three-dimensional accelerometers, this paper investigates the propagation of shock-induced vibrations through the whole lower-limb during running in the temporal and the spectral domains. Results indicated that the vibrations were not consistent across the lower-limb, showing various spatial and spectral distributions of energy. The amount of energy was not constantly decreasing from the distal to the proximal extremity of the runner’s lower-limb, especially regarding the lateral epicondyle of the femur. Vibrations in the transversal plane of the segments were substantial compared to the longitudinal axis regarding the distal extremity of the tibia, and the lateral epicondyle of the femur. Further, the spectral content was wider at the distal than at the proximal end of the lower-limb. Finally, to get a thorough understanding of the risks incurred by the runners, the need to account for shock-induced vibrations up to 50 Hz has been stressed when investigating three-dimensional vibrations. The overall study raises attention on the substantial importance of the transverse components of the acceleration, and their potential relation to shear fatigue and injury during running

Estrogenic regulation of claudin 5 and tight junction protein 1 gene expression in zebrafish: A role on blood-brain barrier?

The blood-brain barrier (BBB) is a physical interface between the blood and the brain parenchyma, playing key roles in brain homeostasis. In mammals, the BBB is established thanks to tight junctions between cerebral endothelial cells, involving claudin, occludin, and zonula occludens proteins. Estrogens have been documented to modulate BBB permeability. Interestingly, in the brain of zebrafish, the estrogen-synthesizing activity is strong due to the high expression of Aromatase B protein, encoded by the cyp19a1b gene, in radial glial cells (neural stem cells). Given the roles of estrogens in BBB function, we investigated their impact on the expression of genes involved in BBB tight junctions. We treated zebrafish embryos and adult males with 17β-estradiol and observed an increased cerebral expression of tight junction and claudin 5 genes in adult males only. In females, treatment with the nuclear estrogen receptor antagonist (ICI182,780 ) had no impact. Interestingly, telencephalic injuries performed in males decreased tight junction gene expression that was partially reversed with 17β-estradiol. This was further confirmed by extravasation experiments of Evans blue showing that estrogenic treatment limits BBB leakage. We also highlighted the intimate links between endothelial cells and neural stem cells, suggesting that cholesterol and peripheral steroids could be taken up by endothelial cells and used as precursors for estrogen synthesis by neural stem cells. Together, our results show that zebrafish provides an alternative model to further investigate the role of steroids on the expression of genes involved in BBB integrity, both in constitutive and regenerative physiological conditions. The link we described between capillaries endothelial cells and steroidogenic neural cells encourages the use of this model in understanding the mechanisms by which peripheral steroids get into neural tissue and modulate neurogenic activity

Comparison of spheroids formed by rat glioma stem cells and neural stem cells reveals differences in glucose metabolism and promising therapeutic applications

Cancer stem cells (CSCs) are thought to be partially responsible for cancer resistance to current therapies and tumor recurrence. Dichloroacetate (DCA), a compound capable of shifting metabolism from glycolysis to glucose oxidation, via an inhibition of pyruvate dehydrogenase kinase was used. We show that DCA is able to shift the pyruvate metabolism in rat glioma CSCs but has no effect in rat neural stem cells. DCA forces CSCs into oxidative phosphorylation but does not trigger the production of reactive oxygen species and consecutive anti-cancer apoptosis. However, DCA, associated with etoposide or irradiation, induced a Bax-dependent apoptosis in CSCs in vitro and decreased their proliferation in vivo. The former phenomenon is related to DCA-induced Foxo3 and p53 expression, resulting in the overexpression of BH3-only proteins (Bad, Noxa, and Puma), which in turn facilitates Bax-dependent apoptosis. Our results demonstrate that a small drug available for clinical studies potentiates the induction of apoptosis in glioma CSCs

AK2 deficiency compromises the mitochondrial energy metabolism required for differentiation of human neutrophil and lymphoid lineages

Reticular dysgenesis is a human severe combined immunodeficiency that is primarily characterized by profound neutropenia and lymphopenia. The condition is caused by mutations in the adenylate kinase 2 (AK2) gene, resulting in the loss of mitochondrial AK2 protein expression. AK2 regulates the homeostasis of mitochondrial adenine nucleotides (ADP, ATP and AMP) by catalyzing the transfer of high-energy phosphate. Our present results demonstrate that AK2-knocked-down progenitor cells have poor proliferative and survival capacities and are blocked in their differentiation toward lymphoid and granulocyte lineages. We also observed that AK2 deficiency impaired mitochondrial function in general and oxidative phosphorylation in particular - showing that AK2 is critical in the control of energy metabolism. Loss of AK2 disrupts this regulation and leads to a profound block in lymphoid and myeloid cell differentiation

Effects of the cannabinoid CB1 antagonist rimonabant on hepatic mitochondrial function in rats fed a high-fat diet

The aim of this study was to investigate the effect of rimonabant treatment on hepatic mitochondrial function in rats fed a high-fat diet. Sprague-Dawley rats fed a high-fat diet (35% lard) for 13 wk were treated with rimonabant (10 mg·kg−1·day−1) during the last 3 wk and matched with pair-fed controls. Oxygen consumption with various substrates, mitochondrial enzyme activities on isolated liver mitochondria, and mitochondrial DNA quantity were determined. Body weight and fat mass were decreased in rats treated with rimonabant compared with pair-fed controls. Moreover, the serum adiponectin level was increased with rimonabant. Hepatic triglyceride content was increased, while serum triglycerides were decreased. An increase of mitochondrial respiration was observed in rats treated with rimonabant. The increase of mitochondrial respiration with palmitoyl-CoA compared with respiration with palmitoyl-l-carnitine stating that the entry of fatty acids into mitochondria via carnitine palmitoyltransferase I was increased in rats treated with rimonabant. Moreover, rimonabant treatment led to a reduction in the enzymatic activity of ATP synthase, whereas the quantity of mitochondrial DNA and the activity of citrate synthase remained unchanged. To summarize, rimonabant treatment leads to an improvement of hepatic mitochondrial function by increasing substrate oxidation and fatty acid entry into mitochondria for the β-oxidation pathway and by increasing proton leak. However, this increase of mitochondrial oxidation is regulated by a decrease of ATP synthase activity in order to have only ATP required for the cell function