João S. Teodoro

SIRT1 is required for AMPK activation and the beneficial effects of resveratrol on mitochondrial function

Resveratrol induces mitochondrial biogenesis and protects against metabolic decline, but whether SIRT1 mediates these benefits is the subject of debate. To circumvent the developmental defects of germline SIRT1 knockouts, we have developed an inducible system that permits whole-body deletion of SIRT1 in adult mice. Mice treated with a moderate dose of resveratrol showed increased mitochondrial biogenesis and function, AMPK activation, and increased NAD(+) levels in skeletal muscle, whereas SIRT1 knockouts displayed none of these benefits. A mouse overexpressing SIRT1 mimicked these effects. A high dose of resveratrol activated AMPK in a SIRT1-independent manner, demonstrating that resveratrol dosage is a critical factor. Importantly, at both doses of resveratrol no improvements in mitochondrial function were observed in animals lacking SIRT1. Together these data indicate that SIRT1 plays an essential role in the ability of moderate doses of resveratrol to stimulate AMPK and improve mitochondrial function both in vitro and in vivo.

Declining NAD+ Induces a Pseudohypoxic State Disrupting Nuclear-Mitochondrial Communication during Aging

Ever since eukaryotes subsumed the bacterial ancestor of mitochondria, the nuclear and mitochondrial genomes have had to closely coordinate their activities, as each encode different subunits of the oxidative phosphorylation (OXPHOS) system. Mitochondrial dysfunction is a hallmark of aging, but its causes are debated. We show that, during aging, there is a specific loss of mitochondrial, but not nuclear, encoded OXPHOS subunits. We trace the cause to an alternate PGC-1α/β-independent pathway of nuclear-mitochondrial communication that is induced by a decline in nuclear NAD(+) and the accumulation of HIF-1α under normoxic conditions, with parallels to Warburg reprogramming. Deleting SIRT1 accelerates this process, whereas raising NAD(+) levels in old mice restores mitochondrial function to that of a young mouse in a SIRT1-dependent manner. Thus, a pseudohypoxic state that disrupts PGC-1α/β-independent nuclear-mitochondrial communication contributes to the decline in mitochondrial function with age, a process that is apparently reversible.

Role of oxidative stress in the pathogenesis of nonalcoholic steatohepatitis

The worldwide rising prevalence of obesity and insulin resistance is associated with a parallel increase in nonalcoholic fatty liver disease (NAFLD). NAFLD is characterized by excess accumulation of triglyceride in the hepatocyte due to increased inflow of free fatty acids and/or de novo lipogenesis caused by various drugs and multiple defects in energy metabolism. Accumulation of lipids in the hepatocyte impairs the oxidative capacity of the mitochondria, increasing the reduced state of the electron transport chain (ETC) complexes and stimulating peroxisomal and microsomal pathways of fat oxidation. The consequent increased generation of reactive oxygen species (ROS) and reactive aldehydic derivatives causes oxidative stress and cell death, via ATP, NAD, and glutathione depletion and DNA, lipid, and protein damage. Oxidative stress also triggers production of inflammatory cytokines, causing inflammation and a fibrogenic response. This ultimately results in the development of nonalcoholic steatohepatitis (NASH), which can result in end-stage liver disease. The current therapeutic strategies for NASH treatment are mostly directed toward correction of the risk factors. Stimulation of mitochondrial function may also prevent NASH development, protecting the cell against the increased flux of reduced substrates to the ETC and ROS generation.

Assessment of the toxicity of silver nanoparticles in vitro: A mitochondrial perspective

The major toxicological concern associated with nanomaterials is the fact that some manufactured nanomaterials are redox active, and some particles transport across cell membranes, especially into mitochondria. Thus, evaluation of their toxicity upon acute exposure is essential. In this work, we evaluated the toxicity of silver nanoparticles (40 and 80 nm) and their effects in rat liver mitochondria bioenergetics. Wistar rat liver mitochondria demonstrate alterations in respiration and membrane potential capacities in the presence of either 40 or 80 nm silver nanoparticles. Our data demonstrated a statistically significant decrease in mitochondrial membrane potential, ADP-induced depolarization, and respiratory control ratio (RCR) upon exposure to silver nanoparticles. Our results show that silver nanoparticles cause impairment of mitochondrial function, due mainly to alterations of mitochondrial membrane permeability. This results in an uncoupling effect on the oxidative phosphorylation system. Thus, mitochondrial toxicity may have a central role in the toxicity resulting from exposure to silver nanoparticles.

Berberine protects against high fat diet-induced dysfunction in muscle mitochondria by inducing SIRT1-dependent mitochondrial biogenesis

Berberine (BBR) has recently been shown to improve insulin sensitivity in rodent models of insulin resistance. Although this effect was explained partly through an observed activation of AMP-activated protein kinase (AMPK), the upstream and downstream mediators of this phenotype were not explored. Here, we show that BBR supplementation reverts mitochondrial dysfunction induced by High Fat Diet (HFD) and hyperglycemia in skeletal muscle, in part due to an increase in mitochondrial biogenesis. Furthermore, we observe that the prevention of mitochondrial dysfunction by BBR, the increase in mitochondrial biogenesis, as well as BBR-induced AMPK activation, are blocked in cells in which SIRT1 has been knocked-down. Taken together, these data reveal an important role for SIRT1 and mitochondrial biogenesis in the preventive effects of BBR on diet-induced insulin resistance.

Differential alterations in mitochondrial function induced by a choline-deficient diet: understanding fatty liver disease progression.

Non-alcoholic fatty liver disease (NAFLD) is an increasingly reported pathology, characterized by fat accumulation within the hepatocyte. Growing evidences suggest specific effects on mitochondrial metabolism, but it is still unclear the relationship between fatty liver progression and mitochondrial function. In the present work we have investigated the impact of fatty liver on mitochondrial bioenergetic functions and susceptibility to mitochondrial permeability transition (MPT) induction in animals fed a choline-deficient diet (CDD) for 4, 8, 12 or 16 weeks. Mitochondria isolated from CDD animals always exhibited higher state 4 respiration. Mitochondrial membrane potential was decreased in CDD animals at 4 and 16 weeks. At 12 weeks, oxidative phosphorylation was more efficient in CDD animals, suggesting a possible early response trying to revert the deleterious effect of increased triglyceride storage in the liver. However, mitochondrial dysfunction was evident in CDD animals at 16 weeks as indicated by decreased RCR and ADP/O, with a corresponding decrease in respiratory chain enzymes activities. Such loss of respiratory efficiency was associated with accumulation of protein oxidation products, in tissue and mitochondrial fraction. Additionally, although no differences in ATPase activity, the lag phase was increased in mitochondria from CDD animals at 16 weeks, associated with decreased content of the adenine nucleotide translocator. Increased susceptibility to calcium-induced MPT was evident in CDD animals at all time points. These results suggest a dynamic mechanism for the development of NALFD associated with altered mitochondrial function.

Hepatic FXR: key regulator of whole-body energy metabolism

The farnesoid X receptor (FXR) is a nuclear receptor whose activation leads to alterations in pathways involved in energy metabolism. For example, it serves as a bile acid receptor in tissues such as the liver, and as an energy metabolism regulator in liver, muscle and adipose tissue. However, the effects of FXR activation are not exclusive to the tissue where it is present, because receptor crosstalk affects tissues throughout the body. It has been demonstrated that FXR regulates the metabolism of not just bile acids, but also of fats and hydrocarbon metabolites. FXR is currently under study as a therapeutic target for the treatment of diseases of excess, such as diabetes. Here we review the effects of FXR activation in the response of an organism to excess energy.

Berberine reverts hepatic mitochondrial dysfunction in high-fat fed rats: A possible role for SirT3 activation

Berberine is an isoquinoline alkaloid with anti-diabetic properties. Despite the central role of liver and thus hepatic mitochondria in whole-body metabolism, berberine effects on hepatic mitochondrial function in an obesity model are still unknown. Here, we demonstrate that berberine treatment recovers mitochondrial efficiency when altered by a high-fat feeding. Mitochondria isolated from the liver of high-fat fed rats exhibited decreased capacity to accumulate calcium and impaired oxidative phosphorylation (OXPHOS) capacity, as shown by impaired mitochondrial membrane potential, oxygen consumption and cellular ATP levels. Interestingly, the recovery of mitochondrial function by berberine was associated with an increased activity of the mitochondrial sirtuin 3 (SirT3). In conclusion, berberine potent protective effects against metabolic syndrome may rely on increasing mitochondrial SirT3 activity, normalizing mitochondrial function and preventing a state of energetic deficit caused by impaired OXPHOS.

The NAD ratio redox paradox: why does too much reductive power cause oxidative stress?

Abstract The reductive power provided by nicotinamide adenine dinucleotides is invaluable for several cellular processes. It drives metabolic reactions, enzymatic activity, regulates genetic expression and allows for the maintenance of a normal cell redox status. Therefore, the balance between the oxidized (NAD+) and the reduced (NADH) forms is critical for the cell’s proper function and ultimately, for its survival. Being intimately associated with the cells’ metabolism, it is expected that alterations to the NAD+/NADH ratio are to be found in situations of metabolic diseases, as is the case of diabetes. NAD+ is a necessary cofactor for several enzymes’ activity, many of which are related to metabolism. Therefore, a decrease in the NAD+/NADH ratio causes these enzymes to decrease in activity (reductive stress), resulting in an altered metabolic situation that might be the first insult toward several pathologies, such as diabetes. Here, we review the importance of nicotinamide adenine dinucleotides in the liver cell and its fluctuations in a state of type 2 diabetes mellitus.

Prevention of I/R injury in fatty livers by ischemic preconditioning is associated with increased mitochondrial tolerance: the key role of ATPsynthase and mitochondrial permeability transition

Ischemia/reperfusion (I/R) injury is a commonly encountered clinical problem and occurs probably as a consequence of irreversible mitochondrial injury. The increased susceptibility of fatty livers to ischemic injury is associated with depletion of adenosine triphosphate (ATP) content, which is preserved by preconditioning. Mitochondria being the main ATP production source for the cell, we aimed to evaluate whether ischemic preconditioning (IPC) of fatty livers prevents the impairment in mitochondrial function induced by I/R. Lean and steatotic animals were subjected to 90 min of hepatic warm ischemia and 12 h of reperfusion. IPC effect was tested in fatty livers. After reperfusion, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured. Mitochondrial membrane potential, mitochondrial respiration and susceptibility to mitochondrial permeability transition (MPT) were evaluated, as well as ATPase activity and adenine nucleotides. IPC of fatty livers decreased serum AST and ALT levels. Fatty animals subjected to I/R exhibited decreased mitochondrial membrane potential and a delay in the repolarization after a phosphorylation cycle, associated with increased state 4 respiration. Increased tolerance to MPT induction, preservation of F1Fo‐ATPsynthase activity and mitochondrial bioenergetics were observed in ischemic preconditioned fatty livers. Thus, IPC is an endogenous protecting mechanism that preserves mitochondrial function and bioenergetics in fatty livers.