In vivo chronic exposure to inorganic mercury worsens hypercholesterolemia, oxidative stress and atherosclerosis in the LDL receptor knockout mice.

Heavy metal exposure leads to multiple system dysfunctions. The mechanisms are likely multifactorial and involve inflammation and oxidative stress. The aim of this study was to evaluate markers and risk factors for atherosclerosis in the LDL receptor knockout mouse model chronically exposed to inorganic mercury (Hg) in the drinking water. Results revealed that Hg exposed mice present increased plasma levels of cholesterol, without alterations in glucose. As a major source and target of oxidants, we evaluated mitochondrial function. We found that liver mitochondria from Hg treated mice show worse respiratory control, lower oxidative phosphorylation efficiency and increased H2O2 release. In addition, Hg induced mitochondrial membrane permeability transition. Erythrocytes from Hg treated mice showed a 50% reduction in their ability to take up oxygen, lower levels of reduced glutathione (GSH) and of antioxidant enzymes (SOD, catalase and GPx). The Hg treatment disturbed immune system cells counting and function. While lymphocytes were reduced, monocytes, eosinophils and neutrophils were increased. Peritoneal macrophages from Hg treated mice showed increased phagocytic activity. Hg exposed mice tissues present metal impregnation and parenchymal architecture alterations. In agreement, increased systemic markers of liver and kidney dysfunction were observed. Plasma, liver and kidney oxidative damage indicators (MDA and carbonyl) were increased while GSH and thiol groups were diminished by Hg exposure. Importantly, atherosclerotic lesion size in the aorta root of Hg exposed mice were larger than in controls. In conclusion, in vivo chronic exposure to Hg worsens the hypercholesterolemia, impairs mitochondrial bioenergetics and redox function, alters immune cells profile and function, causes several tissues oxidative damage and accelerates atherosclerosis development.

CETP expression ameliorates endothelial function in female mice through estrogen receptor-α and endothelial nitric oxide synthase pathway.

Endothelial dysfunction is an early manifestation of atherosclerosis. The cholesteryl ester transfer protein (CETP) has been considered proatherogenic by reducing plasma HDL levels. However, CETP may exhibit cell- or tissue-specific effects. We have previously reported that male mice expressing the human CETP gene show impaired endothelium-mediated vascular relaxation associated with oxidative stress. Although sexual dimorphisms on the metabolic role of CETP have been proposed, possible sex differences in the vascular effects of CETP were not previously studied. Thus, here we investigated the endothelial function of female CETP transgenic mice as compared with nontransgenic controls (NTg). Aortas from CETP females presented preserved endothelium-dependent relaxation to acetylcholine and an endothelium-dependent reduction of phenylephrine-induced contraction. eNOS phosphorylation (Ser1177) and calcium-induced NO levels were enhanced, whereas reactive oxygen species (ROS) production and NOX2 and SOD2 expression were reduced in the CETP female aortas. Furthermore, CETP females exhibited increased aortic relaxation to 17ß-estradiol (E2) and upregulation of heat shock protein 90 (HSP90) and caveolin-1, proteins that stabilize estrogen receptor (ER) in the caveolae. Indeed, CETP females showed an increased E2-induced relaxation in a manner sensitive to estrogen receptor-α (ERα) and HSP90 inhibitors methylpiperidinopyrazole (MPP) and geldanamycin, respectively. MPP also impaired the relaxation response to acetylcholine in CETP but not in NTg females. Altogether, the study indicates that CETP expression ameliorates the anticontractile endothelial effect and relaxation to E2 in females. This was associated with less ROS production, and increased eNOS-NO and E2-ERα pathways. These results highlight the need for considering the sex-specific effects of CETP on cardiovascular risk.NEW & NOTEWORTHY Here we demonstrated that CETP expression has a sex-specific impact on the endothelium function. Contrary to what was described for males, CETP-expressing females present preserved endothelium-dependent relaxation to acetylcholine and improved relaxation response to 17ß-estradiol. This was associated with less ROS production, increased eNOS-derived NO, and increased expression of proteins that stabilize estrogen receptor-α (ERα), thus increasing E2-ERα signaling sensitivity. These results highlight the need for considering the sex-specific effects of CETP on cardiovascular risk.

Lack of mitochondrial NADP(H)-transhydrogenase expression in macrophages exacerbates atherosclerosis in hypercholesterolemic mice.

The atherosclerosis prone LDL receptor knockout mice (Ldlr-/-, C57BL/6J background) carry a deletion of the NADP(H)-transhydrogenase gene (Nnt) encoding the mitochondrial enzyme that catalyzes NADPH synthesis. Here we hypothesize that both increased NADPH consumption (due to increased steroidogenesis) and decreased NADPH generation (due to Nnt deficiency) in Ldlr-/- mice contribute to establish a macrophage oxidative stress and increase atherosclerosis development. Thus, we compared peritoneal macrophages and liver mitochondria from three C57BL/6J mice lines: Ldlr and Nnt double mutant, single Nnt mutant and wild-type. We found increased oxidants production in both mitochondria and macrophages according to a gradient: double mutant > single mutant > wild-type. We also observed a parallel up-regulation of mitochondrial biogenesis (PGC1a, TFAM and respiratory complexes levels) and inflammatory (iNOS, IL6 and IL1b) markers in single and double mutant macrophages. When exposed to modified LDL, the single and double mutant cells exhibited significant increases in lipid accumulation leading to foam cell formation, the hallmark of atherosclerosis. Nnt deficiency cells showed up-regulation of CD36 and down-regulation of ABCA1 transporters what may explain lipid accumulation in macrophages. Finally, Nnt wild-type bone marrow transplantation into LDLr-/- mice resulted in reduced diet-induced atherosclerosis. Therefore, Nnt plays a critical role in the maintenance of macrophage redox, inflammatory and cholesterol homeostasis, which is relevant for delaying the atherogenesis process.

Cholesteryl Ester Transfer Protein and Lipid Metabolism and Cardiovascular Diseases.

In this chapter, we present the major advances in CETP research since the detection, isolation, and characterization of its activity in the plasma of humans and several species. Since CETP is a major modulator of HDL plasma levels, the clinical importance of CETP activity was recognized very early. We describe the participation of CETP in reverse cholesterol transport, conflicting results in animal and human genetic studies, possible new functions of CETP, and the results of the main clinical trials on CETP inhibition. Despite major setbacks in clinical trials, the hypothesis that CETP inhibitors are anti-atherogenic in humans is still being tested.

Coenzyme Q10 protects against ß-cell toxicity induced by pravastatin treatment of hypercholesterolemia.

New onset of diabetes is associated with the use of statins. We have recently demonstrated that pravastatin-treated hypercholesterolemic LDL receptor knockout (LDLr-/- ) mice exhibit reductions in insulin secretion and increased islet cell death and oxidative stress. Here, we hypothesized that these diabetogenic effects of pravastatin could be counteracted by treatment with the antioxidant coenzyme Q 10 (CoQ 10 ), an intermediate generated in the cholesterol synthesis pathway. LDLr -/- mice were treated with pravastatin and/or CoQ 10 for 2 months. Pravastatin treatment resulted in a 75% decrease of liver CoQ 10 content. Dietary CoQ 10 supplementation of pravastatin-treated mice reversed fasting hyperglycemia, improved glucose tolerance (20%) and insulin sensitivity (>2-fold), and fully restored islet glucose-stimulated insulin secretion impaired by pravastatin (40%). Pravastatin had no effect on insulin secretion of wild-type mice. In vitro, insulin-secreting INS1E cells cotreated with CoQ 10 were protected from cell death and oxidative stress induced by pravastatin. Simvastatin and atorvastatin were more potent in inducing dose-dependent INS1E cell death (10-15-fold), which were also attenuated by CoQ 10 cotreatment. Together, these results demonstrate that statins impair ß-cell redox balance, function and viability. However, CoQ 10 supplementation can protect the statins detrimental effects on the endocrine pancreas.

Diabetogenic effect of pravastatin is associated with insulin resistance and myotoxicity in hypercholesterolemic mice.

BACKGROUND: HMG-CoA reductase inhibitors (statins) are cholesterol-lowering drugs widely used to treat hypercholesterolemia and prevent cardiovascular disease. Statins are generally well tolerated, but adverse reactions may occur, particularly myopathy and new onset of diabetes. The exact mechanism of statin-induced myopathy and diabetes has not been fully elucidated. We have previously shown that treatment of hypercholesterolemic (LDLr-/-) mice with pravastatin for 2 months decreased pancreatic islet insulin secretion and increased oxidative stress and cell death, but no glucose intolerance was observed. The purpose of the current work was to study long-term pravastatin effects on glucose homeostasis, insulin sensitivity, muscle protein turnover and cell viability. METHODS: LDLr-/- mice were treated with pravastatin for 3, 6 and 10 months. Glucose tolerance, insulin resistance and glucose-stimulated insulin secretion were evaluated. The rates of protein synthesis and degradation were determined in gastrocnemius muscle after 10 months of treatment. Insulin signalling, oxidative stress and cell death were analysed in vitro using C2C12 myotubes. RESULTS: After 6 and 10 months of treatment, these mice became glucose intolerant, and after 10 months, they exhibited marked insulin resistance. Reduced islet glucose-stimulated insulin secretion was observed after the 3rd month of treatment. Mice treated for 10 months showed significantly decreased body weight and increased muscle protein degradation. In addition, muscle chymotrypsin-like proteasomal activity and lysosomal cathepsin were markedly elevated. C2C12 myotubes exposed to increasing concentrations of pravastatin presented dose-dependent impairment of insulin-induced Akt phosphorylation, increased apoptotic markers (Bax protein and cleaved caspase-3) and augmented superoxide anion production. CONCLUSIONS: In addition to reduced insulin secretion, long-term pravastatin treatment induces insulin resistance and muscle wasting. These results suggest that the diabetogenic effect of statins is linked to the appearance of myotoxicity induced by oxidative stress, impaired insulin signalling, proteolysis and apoptosis.

Contribution to mitochondrial research in Brazil: 10th anniversary of the mitomeeting.

This commentary introduces the subject, the context and the history of the Brazilian annually held meeting on Mitochondrial Research by the occasion of its 10th anniversary. Mitomeetings gather people interested in all aspects of mitochondrial biology in diverse species, including protists, animals, plants, and fungi.

Mangifera indica L. extract (Vimang®) reduces plasma and liver cholesterol and leucocyte oxidative stress in hypercholesterolemic LDL receptor deficient mice.

Cardiovascular diseases are major causes of death worldwide. Beyond the classical cholesterol risk factor, other conditions such as oxidative stress are well documented to promote atherosclerosis. The Mangifera indica L. extract (Vimang®) was reported to present antioxidant and hypocholesterolemic properties. Thus, here we evaluate the effects of Vimang treatment on risk factors of the atherosclerosis prone model of familial hypercholesterolemia, the LDL receptor knockout mice. Mice were treated with Vimang during 2 weeks and were fed a cholesterol-enriched diet during the second week. The Vimang treated mice presented significantly reduced levels of plasma (15%) and liver (20%) cholesterol, increased plasma total antioxidant capacity (10%) and decreased reactive oxygen species (ROS) production by spleen mononuclear cells (50%), P < 0.05 for all. In spite of these benefits, the average size of aortic atherosclerotic lesions stablished in this short experimental period did not change significantly in Vimang treated mice. Therefore, in this study we demonstrated that Vimang has protective effects on systemic and tissue-specific risk factors, but it is not sufficient to promote a reduction in the initial steps of atherosclerosis development. In addition, we disclosed a new antioxidant target of Vimang, the spleen mononuclear cells that might be relevant for more advanced stages of atherosclerosis.

Facilitation of Ca2+ -induced opening of the mitochondrial permeability transition pore either by nicotinamide nucleotide transhydrogenase deficiency or statins treatment.

Mitochondrial redox imbalance and high Ca2+ uptake induce the opening of the permeability transition pore (PTP) that leads to disruption of energy-linked mitochondrial functions and triggers cell death in many disease states. In this review, we discuss the major results from our studies investigating the consequences of NAD(P)-transhydrogenase (NNT) deficiency, and of statins treatment for mitochondrial functions and susceptibility to Ca2+ -induced PTP. We highlight the aggravation of high fat diet-induced fatty liver disease in the context of NNT deficiency and the role of antioxidants in the prevention of statins toxicity to mitochondria.

Aerobic exercise training protects against endothelial dysfunction by increasing nitric oxide and hydrogen peroxide production in LDL receptor-deficient mice.

BACKGROUND: Endothelial dysfunction associated with hypercholesterolemia is an early event in atherosclerosis characterized by redox imbalance associated with high superoxide production and reduced nitric oxide (NO) and hydrogen peroxide (H2O2) production. Aerobic exercise training (AET) has been demonstrated to ameliorate atherosclerotic lesions and oxidative stress in advanced atherosclerosis. However, whether AET protects against the early mechanisms of endothelial dysfunction in familial hypercholesterolemia remains unclear. This study investigated the effects of AET on endothelial dysfunction and vascular redox status in the aortas of LDL receptor knockout mice (LDLr(-/-)), a genetic model of familial hypercholesterolemia. METHODS: Twelve-week-old C57BL/6J (WT) and LDLr(-/-) mice were divided into sedentary and exercised (AET on a treadmill 1 h/5 × per week) groups for 4 weeks. Changes in lipid profiles, endothelial function, and aortic NO, H2O2 and superoxide production were examined. RESULTS: Total cholesterol and triglycerides were increased in sedentary and exercised LDLr(-/-) mice. Endothelium-dependent relaxation induced by acetylcholine was impaired in aortas of sedentary LDLr(-/-) mice but not in the exercised group. Inhibition of NO synthase (NOS) activity or H2O2 decomposition by catalase abolished the differences in the acetylcholine response between the animals. No changes were noted in the relaxation response induced by NO donor sodium nitroprusside or H2O2. Neuronal NOS expression and endothelial NOS phosphorylation (Ser1177), as well as NO and H2O2 production, were reduced in aortas of sedentary LDLr(-/-) mice and restored by AET. Incubation with apocynin increased acetylcholine-induced relaxation in sedentary, but not exercised LDLr(-/-) mice, suggesting a minor participation of NADPH oxidase in the endothelium-dependent relaxation after AET. Consistent with these findings, Nox2 expression and superoxide production were reduced in the aortas of exercised compared to sedentary LDLr(-/-) mice. Furthermore, the aortas of sedentary LDLr(-/-) mice showed reduced expression of superoxide dismutase (SOD) isoforms and minor participation of Cu/Zn-dependent SODs in acetylcholine-induced, endothelium-dependent relaxation, abnormalities that were partially attenuated in exercised LDLr(-/-) mice. CONCLUSION: The data gathered by this study suggest AET as a potential non-pharmacological therapy in the prevention of very early endothelial dysfunction and redox imbalance in familial hypercholesterolemia via increases in NO bioavailability and H2O2 production.

Cholesteryl Ester Transfer Protein (CETP) expression does not affect glucose homeostasis and insulin secretion: studies in human CETP transgenic mice.

BACKGROUND: Cholesteryl ester transfer protein (CETP) is a plasma protein that mediates the exchange of triglycerides for esterified cholesterol between HDL and apoB-lipoproteins. Previous studies suggest that CETP may modify glucose metabolism in patients or cultured cells. In this study, we tested if stable CETP expression would impair glucose metabolism. METHODS: We used human CETP transgenic mice and non-transgenic littermate controls (NTg), fed with control or high fat diet, as well as in dyslipidemic background and aging conditions. Assays included glucose and insulin tolerance tests, isolated islets insulin secretion, tissue glucose uptake and adipose tissue GLUT mRNA expression. RESULTS: CETP expression did not modify glucose or insulin tolerance in all tested conditions such as chow and high fat diet, adult and aged mice, normo and dyslipidemic backgrounds. Fasting and fed state plasma levels of insulin were not differ in CETP and NTg mice. Direct measurements of isolated pancreatic islet insulin secretion rates induced by glucose (11, 16.7 or 22 mM), KCl (40 mM), and leucine (10 mM) were similar in NTg and CETP mice, indicating that CETP expression did not affect ß-cell function in vivo and ex vivo. Glucose uptake by insulin target tissues, measured in vivo using (3)H-2-deoxyglucose, showed that CETP expression had no effect on the glucose uptake in liver, muscle, perigonadal, perirenal, subcutaneous and brown adipose tissues. Accordingly, GLUT1 and GLUT4 mRNA in adipose tissue were not affected by CETP. CONCLUSIONS: In summary, by comparing the in vivo all-or-nothing CETP expressing mouse models, we demonstrated that CETP per se has no impact on the glucose tolerance and tissue uptake, global insulin sensitivity and beta cell insulin secretion rates.

Impaired compensatory beta-cell function and growth in response to high-fat diet in LDL receptor knockout mice.

In this study, we investigated the effect of low density lipoprotein receptor (LDLr) deficiency on gap junctional connexin 36 (Cx36) islet content and on the functional and growth response of pancreatic beta-cells in C57BL/6 mice fed a high-fat (HF) diet. After 60 days on regular or HF diet, the metabolic state and morphometric islet parameters of wild-type (WT) and LDLr-/- mice were assessed. HF diet-fed WT animals became obese and hypercholesterolaemic as well as hyperglycaemic, hyperinsulinaemic, glucose intolerant and insulin resistant, characterizing them as prediabetic. Also they showed a significant decrease in beta-cell secretory response to glucose. Overall, LDLr-/- mice displayed greater susceptibility to HF diet as judged by their marked cholesterolaemia, intolerance to glucose and pronounced decrease in glucose-stimulated insulin secretion. HF diet induced similarly in WT and LDLr-/- mice, a significant decrease in Cx36 beta-cell content as revealed by immunoblotting. Prediabetic WT mice displayed marked increase in beta-cell mass mainly due to beta-cell hypertrophy/replication. Nevertheless, HF diet-fed LDLr-/- mice showed no significant changes in beta-cell mass, but lower islet-duct association (neogenesis) and higher beta-cell apoptosis index were seen as compared to controls. The higher metabolic susceptibility to HF diet of LDLr-/- mice may be explained by a deficiency in insulin secretory response to glucose associated with lack of compensatory beta-cell expansion.

Food restriction by intermittent fasting induces diabetes and obesity and aggravates spontaneous atherosclerosis development in hypercholesterolaemic mice.

Different regimens of food restriction have been associated with protection against obesity, diabetes and CVD. In the present study, we hypothesised that food restriction would bring benefits to atherosclerosis- and diabetes-prone hypercholesterolaemic LDL-receptor knockout mice. For this purpose, 2-month-old mice were submitted to an intermittent fasting (IF) regimen (fasting every other day) over a 3-month period, which resulted in an overall 20 % reduction in food intake. Contrary to our expectation, epididymal and carcass fat depots and adipocyte size were significantly enlarged by 15, 72 and 68 %, respectively, in the IF mice compared with the ad libitum-fed mice. Accordingly, plasma levels of leptin were 50 % higher in the IF mice than in the ad libitum-fed mice. In addition, the IF mice showed increased plasma levels of total cholesterol (37 %), VLDL-cholesterol (195 %) and LDL-cholesterol (50 %). As expected, in wild-type mice, the IF regimen decreased plasma cholesterol levels and epididymal fat mass. Glucose homeostasis was also disturbed by the IF regimen in LDL-receptor knockout mice. Elevated levels of glycaemia (40 %), insulinaemia (50 %), glucose intolerance and insulin resistance were observed in the IF mice. Systemic inflammatory markers, TNF-α and C-reactive protein, were significantly increased and spontaneous atherosclerosis development were markedly increased (3-fold) in the IF mice. In conclusion, the IF regimen induced obesity and diabetes and worsened the development of spontaneous atherosclerosis in LDL-receptor knockout mice. Although being efficient in a wild-type background, this type of food restriction is not beneficial in the context of genetic hypercholesterolaemia.

Activation of the mitochondrial ATP-sensitive K+ channel reduces apoptosis of spleen mononuclear cells induced by hyperlipidemia.

BACKGROUND: We have previously demonstrated that increased rates of superoxide generation by extra-mitochondrial enzymes induce the activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) in the livers of hypertriglyceridemic (HTG) mice. The resulting mild uncoupling mediated by mitoK(ATP) protects mitochondria against oxidative damage. In this study, we investigate whether immune cells from HTG mice also present increased mitoK(ATP) activity and evaluate the influence of this trait on cell redox state and viability. METHODS: Oxygen consumption (Clark-type electrode), reactive oxygen species production (dihydroethidium and H2-DCF-DA probes) and cell death (annexin V, cytocrome c release and Trypan blue exclusion) were determined in spleen mononuclear cells. RESULTS: HTG mice mononuclear cells displayed increased mitoK(ATP) activity, as evidenced by higher resting respiration rates that were sensitive to mitoK(ATP) antagonists. Whole cell superoxide production and apoptosis rates were increased in HTG cells. Inhibition of mitoK(ATP) further increased the production of reactive oxygen species and apoptosis in these cells. Incubation with HTG serum induced apoptosis more strongly in WT cells than in HTG mononuclear cells. Cytochrome c release into the cytosol and caspase 8 activity were both increased in HTG cells, indicating that cell death signaling starts upstream of the mitochondria but does involve this organelle. Accordingly, a reduced number of blood circulating lymphocytes was found in HTG mice. CONCLUSIONS: These results demonstrate that spleen mononuclear cells from hyperlipidemic mice have more active mitoK(ATP) channels, which downregulate mitochondrial superoxide generation. The increased apoptosis rate observed in these cells is exacerbated by closing the mitoK(ATP) channels. Thus, mitoK(ATP) opening acts as a protective mechanism that reduces cell death induced by hyperlipidemia.

CETP Expression in Bone-Marrow-Derived Cells Reduces the Inflammatory Features of Atherosclerosis in Hypercholesterolemic Mice.

CETP activity reduces plasma HDL-cholesterol concentrations, a correlate of an increased risk of atherosclerotic events. However, our recent findings suggest that CETP expression in macrophages promotes an intracellular antioxidant state, reduces free cholesterol accumulation and phagocytosis, and attenuates pro-inflammatory gene expression. To determine whether CETP expression in macrophages affects atherosclerosis development, we transplanted bone marrow from transgenic mice expressing simian CETP or non-expressing littermates into hypercholesterolemic LDL-receptor-deficient mice. The CETP expression did not change the lipid-stained lesion areas but decreased the macrophage content (CD68), neutrophil accumulation (LY6G), and TNF-α aorta content of young male transplanted mice and decreased LY6G, TNF-α, iNOS, and nitrotyrosine (3-NT) in aged female transplanted mice. These findings suggest that CETP expression in bone-marrow-derived cells reduces the inflammatory features of atherosclerosis. These novel mechanistic observations may help to explain the failure of CETP inhibitors in reducing atherosclerotic events in humans.

Enhanced insulin secretion and glucose tolerance in rats exhibiting low plasma free fatty acid levels and hypertriglyceridaemia due to congenital albumin deficiency.

Congenitally analbuminaemic individuals and rats (NARs) exhibit several metabolic abnormalities, including hypertriglyceridaemia and plasma free fatty acid deficiency. Our aim was to study glucose homeostasis and insulin secretion in NARs. Plasma concentrations of lipids, glucose and insulin and secretion of insulin from the pancreatic islets were measured in female NARs and control animals (Sprague-Dawley rats; SDRs). Glucose homeostasis tests were also performed. Plasma glucose levels were similar between NARs and SDRs, irrespective of feeding status. However, fed insulinaemia was ∼37% higher (P 0.05) in NARs than in SDRs. The NARs displayed a markedly increased glucose tolerance, i.e. the integrated glycaemic response was one-third that of the control animals. Enhanced glucose tolerance was associated with threefold higher insulinaemia at peak glycaemia after a glucose load than in the control animals. Similar peripheral insulin sensitivity was observed between groups. Isolated pancreatic islets from NARs secreted significantly more insulin than islets from SDRs in response to a wide range of glucose concentrations (2.8-33.3 mm). Despite having similar liver glycogen contents in the fully fed state, NARs had ∼40% (P 0.05) lower glycogen contents than SDRs after 6 h fasting. The injection of a gluconeogenic substrate, pyruvate, elicited a faster rise in glycaemia in NARs compared with SDRs. Overall, NARs displayed enhanced glucose tolerance, insulin secretion and gluconeogenic flux. The higher glucose tolerance in NARs compared with SDRs is attributed to enhanced islet responsiveness to secretagogues, while peripheral insulin sensitivity seems not to be involved in this alteration. We propose that the enhanced glucose metabolism is a chronic compensatory adaptation to decreased free fatty acid availability in NARs.

Mitochondrial Adaptive Responses to Hypertriglyceridemia and Bioactive Lipids.

Significance: Altered plasma triglyceride metabolism and changes in dietary fatty acid types and levels are major contributors to the development of metabolic and cardiovascular diseases such as fatty liver disease, obesity, diabetes, and atherosclerosis. Lipid accumulation in visceral adipose tissue and ectopically in other organs, as well as lipid-induced redox imbalance, is connected to mitochondrial dysfunction in a range of oxidative stress-associated metabolic and degenerative disorders. Recent Advances: Successful mitochondrial adaptive responses in the context of hypertriglyceridemia and dietary bioactive polyunsaturated fatty acids contribute to increase body energy expenditure and reduce oxidative stress, thus allowing several cell types to cope with metabolic challenges and stresses. These responses include mitochondrial redox signaling, mild uncoupling, and changes in network dynamic behavior. Critical Issues: Mitochondrial bioenergetics and redox changes in a lipid overload context are relatively well characterized. However, the turning point between adaptive and maladaptive mitochondrial responses remains a critical issue to be elucidated. In addition, the relationship between changes in fusion/fission machinery and mitochondrial function is less well understood. Future Directions: The effective mitochondrial responses described here support the research for new drug design and diet or nutraceutical formulations targeting mitochondrial mild uncoupling and effective quality control as putative strategies for cardiometabolic diseases. Antioxid. Redox Signal. 36, 953-968.

Dichloroacetate reactivates pyruvate-supported peroxide removal by liver mitochondria and prevents NAFLD aggravation in NAD(P)+ transhydrogenase-null mice consuming a high-fat diet.

The mechanisms by which a high-fat diet (HFD) promotes non-alcoholic fatty liver disease (NAFLD) appear to involve liver mitochondrial dysfunction and redox imbalance. The functional loss of the enzyme NAD(P)+ transhydrogenase, a main source of mitochondrial NADPH, results in impaired mitochondrial peroxide removal, pyruvate dehydrogenase inhibition by phosphorylation, and progression of NAFLD in HFD-fed mice. The present study aimed to investigate whether pharmacological reactivation of pyruvate dehydrogenase by dichloroacetate attenuates the mitochondrial redox dysfunction and the development of NAFLD in NAD(P)+ transhydrogenase-null (Nnt-/-) mice fed an HFD (60% of total calories from fat). For this purpose, Nnt-/- mice and their congenic controls (Nnt+/+) were fed chow or an HFD for 20 weeks and received sodium dichloroacetate or NaCl in the final 12 weeks via drinking water. The results showed that HFD reduced the ability of isolated liver mitochondria from Nnt-/- mice to remove peroxide, which was prevented by the dichloroacetate treatment. HFD-fed mice of both Nnt genotypes exhibited increased body and liver mass, as well as a higher content of hepatic triglycerides, but dichloroacetate treatment attenuated these abnormalities only in Nnt-/- mice. Notably, dichloroacetate treatment decreased liver pyruvate dehydrogenase phosphorylation levels and prevented the aggravation of NAFLD in HFD-fed Nnt-/- mice. Conversely, dichloroacetate treatment elicited moderate hepatocyte ballooning in chow-fed mice, suggesting potentially toxic effects. We conclude that the protection against HFD-induced NAFLD by dichloroacetate is associated with its role in reactivating pyruvate dehydrogenase and reestablishing the pyruvate-supported liver mitochondrial capacity to handle peroxide in Nnt-/- mice.

Mild Mitochondrial Uncoupling Decreases Experimental Atherosclerosis, A Proof of Concept.

AIM: Atherosclerosis is responsible for high morbidity and mortality rates around the world. Local arterial oxidative stress is involved in all phases of atherosclerosis development. Mitochondria is a relevant source of the oxidants, particularly under certain risky conditions, such as hypercholesterolemia. The aim of this study was to test whether lowering the production of mitochondrial oxidants by induction of a mild uncoupling can reduce atherosclerosis in hypercholesterolemic LDL receptor knockout mice. METHODS: The mice were chronically treated with very low doses of DNP (2,4-dinitrophenol) and metabolic, inflammatory and redox state markers and atherosclerotic lesion sizes were determined. RESULTS: The DNP treatment did not change the classical atherosclerotic risk markers, such as plasma lipids, glucose homeostasis, and fat mass, as well as systemic inflammatory markers. However, the DNP treatment diminished the production of mitochondrial oxidants, systemic and tissue oxidative damage markers, peritoneal macrophages and aortic rings oxidants generation. Most importantly, development of spontaneous and diet-induced atherosclerosis (lipid and macrophage content) were significantly decreased in the DNP-treated mice. In vitro, DNP treated peritoneal macrophages showed decreased H2O2 production, increased anti-inflammatory cytokines gene expression and secretion, increased phagocytic activity, and decreased LDL-cholesterol uptake. CONCLUSIONS: These findings are a proof of concept that activation of mild mitochondrial uncoupling is sufficient to delay the development of atherosclerosis under the conditions of hypercholesterolemia and oxidative stress. These results promote future approaches targeting mitochondria for the prevention or treatment of atherosclerosis.

A systematic review and meta-analyses on the effects of atorvastatin on blood pressure and heart rate.

AIMS: Considering the inconsistencies in the literature on the atorvastatin effect on blood pressure (BP), we performed these meta-analyses. METHODS AND RESULTS: Through a search of the Excerpta Medica Database (EMBASE), PubMed, and Web of Science databases, 1412 articles were identified, from which 33 randomized clinical trials (RCT) and 44 pre-clinical were selected. Populations from RCT were stratified according to baseline BP and lipid levels. We performed meta-analyses of the effect of atorvastatin on systolic (SBP), diastolic and mean BP; heart rate (HR); HR variability, and baroreflex. Atorvastatin reduced SBP in the overall population (P = 0.05 vs. placebo; P = 0.03 vs. baseline), in normotensive and hyperlipidaemic (P = 0.04 vs. placebo; P = 0.0001 vs. baseline) and in hypertensive and hyperlipidaemic (P = 0.02 vs. placebo; P = 0.008 vs. baseline) individuals in parallel RCT, but it did not affect SBP in normotensive and normolipidaemic individuals (P = 0.51 vs. placebo; P = 0.4 vs. baseline). Although an effect of atorvastatin was detected in hyperlipidaemic individuals, the meta-regression coefficient for the association of low density lipoprotein (LDL)-cholesterol reduction with SBP reduction in the overall population demonstrated that SBP reduction is not dependent on the changes in LDL-cholesterol. A meta-analysis of preclinical reports demonstrated that SBP was reduced in atorvastatin-treated hypertensive and normolipidaemic rats (spontaneously hypertensive rats: P < 0.00001), but not in normotensive and normolipidaemic rats (control rats: P = 0.97). Atorvastatin also reduced the HR in spontaneously hypertensive rat. CONCLUSION: Atorvastatin lowers BP independent of LDL-cholesterol levels. Additional studies are needed to estimate the involvement of the autonomic nervous system in the BP-lowering effect of atorvastatin.