Ana Teresa Varela

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.

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.

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.

Enhancement of brown fat thermogenesis using chenodeoxycholic acid in mice

Objective:Besides their role in lipid absorption, bile acids (BAs) can act as signalling molecules. Cholic acid was shown to counteract obesity and associated metabolic disorders in high-fat-diet (cHF)-fed mice while enhancing energy expenditure through induction of mitochondrial uncoupling protein 1 (UCP1) and activation of non-shivering thermogenesis in brown adipose tissue (BAT). In this study, the effects of another natural BA, chenodeoxycholic acid (CDCA), on dietary obesity, UCP1 in both interscapular BAT and in white adipose tissue (brite cells in WAT), were characterized in dietary-obese mice.Research design:To induce obesity and associated metabolic disorders, male 2-month-old C57BL/6J mice were fed cHF (35% lipid wt wt−1, mainly corn oil) for 4 months. Mice were then fed either (i) for 8 weeks with cHF or with cHF with two different doses (0.5%, 1%; wt wt−1) of CDCA (8-week reversion); or (ii) for 3 weeks with cHF or with cHF with 1% CDCA, or pair-fed (PF) to match calorie intake of the CDCA mice fed ad libitum; mice on standard chow diet were also used (3-week reversion).Results:In the 8-week reversion, the CDCA intervention resulted in a dose-dependent reduction of obesity, dyslipidaemia and glucose intolerance, which could be largely explained by a transient decrease in food intake. The 3-week reversion revealed mild CDCA-dependent and food intake-independent induction of UCP1-mediated thermogenesis in interscapular BAT, negligible increase of UCP1 in subcutaneous WAT and a shift from carbohydrate to lipid oxidation.Conclusions:CDCA could reverse obesity in cHF-fed mice, mainly in response to the reduction in food intake, an effect probably occuring but neglected in previous studies using cholic acid. Nevertheless, CDCA-dependent and food intake-independent induction of UCP1 in BAT (but not in WAT) could contribute to the reduction in adiposity and to the stabilization of the lean phenotype.

Sirtuin 1 in rat orthotopic liver transplantation: An IGL-1 preservation solution approach

AIM To investigate the possible involvement of Sirtuin 1 (SIRT1) in rat orthotopic liver transplantation (OLT), when Institute Georges Lopez 1 (IGL-1) preservation solution is enriched with trimetazidine (TMZ). METHODS Male Sprague-Dawley rats were used as donors and recipients. Livers were stored in IGL-1 preservation solution for 8h at 4 °C, and then underwent OLT according to Kamadas cuff technique without arterialization. In another group, livers were stored in IGL-1 preservation solution supplemented with TMZ, at 10(-6) mol/L, for 8 h at 4 °C and then underwent OLT. Rats were sacrificed 24 h after reperfusion, and liver and plasma samples were collected. Liver injury (transaminase levels), mitochondrial damage (glutamate dehydrogenase activity) oxidative stress (malondialdehyde levels), and nicotinamide adenine dinucleotide (NAD(+)), the co-factor necessary for SIRT1 activity, were determined by biochemical methods. SIRT1 and its substrates (ac-FoxO1, ac-p53), the precursor of NAD(+), nicotinamide phosphoribosyltransferase (NAMPT), as well as the phosphorylation of adenosine monophosphate activated protein kinase (AMPK), p-mTOR, p-p70S6K (direct substrate of mTOR), autophagy parameters (beclin-1, LC3B) and MAP kinases (p-p38 and p-ERK) were determined by Western blot. RESULTS Liver grafts preserved in IGL-1 solution enriched with TMZ presented reduced liver injury and mitochondrial damage compared with those preserved in IGL-1 solution alone. In addition, livers preserved in IGL-1 + TMZ presented reduced levels of oxidative stress. This was consistent with enhanced SIRT1 protein expression and elevated SIRT1 activity, as indicated by decreased acetylation of p53 and FoxO1. The elevated SIRT1 activity in presence of TMZ can be attributed to the enhanced NAMPT protein and NAD(+)/NADH levels. Up-regulation of SIRT1 was consistent with activation of AMPK and inhibition of phosphorylation of mTOR and its direct substrate (p-p70S6K). As a consequence, autophagy mediators (beclin-1 and LC3B) were over-expressed. Furthermore, MAP kinases were regulated in livers preserved with IGL-1 + TMZ, as they were characterized by enhanced p-ERK and decreased p-p38 protein expression. CONCLUSION Our study shows that IGL-1 preservation solution enriched with TMZ protects liver grafts from the IRI associated with OLT, through SIRT1 up-regulation.

Dibenzofuran-induced mitochondrial dysfunction: Interaction with ANT carrier

Exposure to environmental pollutants such as dibenzofurans and furans is linked to the pathophysiology of several diseases. Dibenzofuran (DBF) is listed as a pollutant of concern due to its persistence in the environment, bioaccumulation and toxicity to humans, being associated with the development of lung diseases and cancers, due to its extremely toxic properties such as carcinogenic and teratogenic. Mitochondria play a key role in cellular homeostasis and keeping a proper energy supply for eukaryotic cells is essential in the fulfillment of the tissues energy-demand. Therefore, interference with mitochondrial function leads to cell death and organ failure. In this work, the effects of DBF on isolated rat liver mitochondria were analyzed. DBF exposure caused a markedly increase in the lag phase that follows depolarization induced by ADP, indicating an effect in the phosphorylative system. This was associated with a dose-dependent decrease in ATPase activity. Moreover, DBF also increased the threshold to the induction of the mitochondrial permeability transition (MPT) by calcium. Pretreatment of mitochondria with DBF also increased the concentration of carboxyatractyloside (CAT) necessary to abolish ADP phosphorylation and to induce the MPT, suggesting that DBF may interfere with mitochondria through an effect on the adenine nucleotide translocase (ANT). By co-immunoprecipitating ANT and Cyclophilin D (CypD) following MPT induction, we observed that in the presence of DBF, the ratio CypD/ANT was decreased. This demonstrates that DBF interferes with the ANT and so prevents CypD binding to the ANT, causing decreased phosphorylative capacity and inhibiting the MPT, which is also reflected by an increase in calcium retention capacity. Clarifying the role of pollutants in some mechanisms of toxicity, such as unbalance of bioenergetics status and mitochondrial function, may help to explain the progressive and chronic evolution of diseases derived from exposure to environmental pollutants.

High-fat and obesogenic diets: current and future strategies to fight obesity and diabetes

Obesity, diabetes and their associated diseases are some of the greatest challenges that the world health care systems already face and with prospects of overburdening their capacities and funding. Due to decreased energetic expenditure and increased caloric intake, particularly in saturated fats, the number of people afflicted by said conditions is increasing by the day. Due to the failure, to this day, to effectively and ubiquity prevent and revert these diseases, the research into new compounds and therapeutic strategies is vital. In this review, we explain the most common dietary models of obesity and diabetes and the novel avenues of research we believe will be taken in the next few years in obesity and diabetes research. We primarily focus on the role of mitochondria and how the modulation of mitochondrial function and number as well as several promising therapeutic strategies involving metabolic regulators can positively affect the obese and diabetic status.

Fatty liver and ischemia/reperfusion: are there drugs able to mitigate injury?

Fatty livers are more prone to damage caused by ischemia/reperfusion (I/R). Impaired microcirculation, Kupffer cell dysfunction, increased adhesion of leukocytes, impaired mitochondrial function and ATP depletion are probable causes for fatty liver susceptibility. Therefore, hepatic steatosis is a major risk factor for liver surgery and success of transplantation of fatty donor organs. The mechanisms involved in I/R injury are complex and there is no general consensus regarding the sources of ROS generation, nitric oxide (NO) action, the role of tumor necrosis factor-α (TNF-α), and transcription factors, such as nuclear factor kappa B (NFκB). Impairment of mitochondrial function is one of the most important alterations that occur in I/R injury, resulting in the alteration of energy metabolism. Ischemic preconditioning (IPC) and post conditioning (IPost) are adaptive mechanisms against I/R insults that induce intracellular protective responses associated with the preservation of mitochondrial function.There are several pharmacological drugs and natural derivatives presenting metabolic and/or antioxidant effects that can directly or indirectly protect the liver against I/R injury. While the precise targets and mechanisms are still not totally understood, the mitochondrion presents itself as a major player on mediating these protective events. As so, compounds that are able to improve mitochondrial function and hepatic energetic balance might prove viable candidates when developing new pharmacological approaches that can minimize injury to steatotic livers subjected to I/R events.

Mitochondrial bioenergetics and posthepatectomy liver dysfunction

Liver regeneration requires an enormous energy supply. Experimental evidence suggests that mitochondrial function is of paramount importance for liver regeneration. However, this has not been investigated in the clinical setting. We aimed to: (i) evaluate changes in mitochondrial function during hepatectomy, especially after hepatic pedicle clamping; and (ii) correlate these changes with postoperative hepatocellular function and clinical outcome.

PPARα Agonist WY-14643 Induces SIRT1 Activity in Rat Fatty Liver Ischemia-Reperfusion Injury

Ischemia-reperfusion injury (IRI) remains a frequent complication in surgery, especially in case of steatotic livers that present decreased tolerance towards IRI. Apart from its major role in metabolism, activation of peroxisome proliferator-activated receptor α (PPARα) has been related with positive effects on IRI. In addition, the deacetylase enzyme sirtuin 1 (SIRT1) has recently emerged as a promising target for preventing IRI, through its interaction with stress-related mechanisms, such as endoplasmic reticulum stress (ERS). Taking this into account, this study aims to explore whether PPARα agonist WY-14643 could protect steatotic livers against IRI through sirtuins and ERS signaling pathway. Obese Zucker rats were pretreated or not pretreated with WY-14643 (10 mg/kg intravenously) and then submitted to partial (70%) hepatic ischemia (1 hour) followed by 24 hours of reperfusion. Liver injury (ALT levels), lipid peroxidation (MDA), SIRT1 activity, and the protein expression of SIRT1 and SIRT3 and ERS parameters (IRE1α, peIF2, caspase 12, and CHOP) were evaluated. Treatment with WY-14643 reduced liver injury in fatty livers, enhanced SIRT1 activity, and prevented ERS. Together, our results indicated that PPARα agonist WY-14643 may exert its protective effect in fatty livers, at least in part, via SIRT1 induction and ERS prevention.