Phylogeography and Pleistocene refugia of the adder (Vipera berus) as inferred from mitochondrial DNA sequence data.

In order to contribute to the debate about southern glacial refugia used by temperate species and more northern refugia used by boreal or cold-temperate species, we examined the phylogeography of a widespread snake species (Vipera berus) inhabiting Europe up to the Arctic Circle. The analysis of the mitochondrial DNA (mtDNA) sequence variation in 1043 bp of the cytochrome b gene and in 918 bp of the noncoding control region was performed with phylogenetic approaches. Our results suggest that both the duplicated control region and cytochrome b evolve at a similar rate in this species. Phylogenetic analysis showed that V. berus is divided into three major mitochondrial lineages, probably resulting from an Italian, a Balkan and a Northern (from France to Russia) refugial area in Eastern Europe, near the Carpathian Mountains. In addition, the Northern clade presents an important substructure, suggesting two sequential colonization events in Europe. First, the continent was colonized from the three main refugial areas mentioned above during the Lower-Mid Pleistocene. Second, recolonization of most of Europe most likely originated from several refugia located outside of the Mediterranean peninsulas (Carpathian region, east of the Carpathians, France and possibly Hungary) during the Mid-Late Pleistocene, while populations within the Italian and Balkan Peninsulas fluctuated only slightly in distribution range, with larger lowland populations during glacial times and with refugial mountain populations during interglacials, as in the present time. The phylogeographical structure revealed in our study suggests complex recolonization dynamics of the European continent by V. berus, characterized by latitudinal as well as altitudinal range shifts, driven by both climatic changes and competition with related species.

Selective ion changes during spontaneous mitochondrial transients in intact astrocytes.

The bioenergetic status of cells is tightly regulated by the activity of cytosolic enzymes and mitochondrial ATP production. To adapt their metabolism to cellular energy needs, mitochondria have been shown to exhibit changes in their ionic composition as the result of changes in cytosolic ion concentrations. Individual mitochondria also exhibit spontaneous changes in their electrical potential without altering those of neighboring mitochondria. We recently reported that individual mitochondria of intact astrocytes exhibit spontaneous transient increases in their Na(+) concentration. Here, we investigated whether the concentration of other ionic species were involved during mitochondrial transients. By combining fluorescence imaging methods, we performed a multiparameter study of spontaneous mitochondrial transients in intact resting astrocytes. We show that mitochondria exhibit coincident changes in their Na(+) concentration, electrical potential, matrix pH and mitochondrial reactive oxygen species production during a mitochondrial transient without involving detectable changes in their Ca(2+) concentration. Using widefield and total internal reflection fluorescence imaging, we found evidence for localized transient decreases in the free Mg(2+) concentration accompanying mitochondrial Na(+) spikes that could indicate an associated local and transient enrichment in the ATP concentration. Therefore, we propose a sequential model for mitochondrial transients involving a localized ATP microdomain that triggers a Na(+)-mediated mitochondrial depolarization, transiently enhancing the activity of the mitochondrial respiratory chain. Our work provides a model describing ionic changes that could support a bidirectional cytosol-to-mitochondria ionic communication.

A nuclear defect in the 4p16 region predisposes to multiple mitochondrial DNA deletions in families with Wolfram syndrome.

Wolfram syndrome is a progressive neurodegenerative disorder transmitted in an autosomal recessive mode. We report two Wolfram syndrome families harboring multiple deletions of mitochondrial DNA. The deletions reached percentages as high as 85-90% in affected tissues such as the central nervous system of one patient, while in other tissues from the same patient and from other members of the family, the percentages of deleted mitochondrial DNA genomes were only 1-10%. Recently, a Wolfram syndrome gene has been linked to markers on 4p16. In both families linkage between the disease locus and 4p16 markers gave a maximum multipoint lod score of 3.79 at theta = 0 (Pi<0.03) with respect to D4S431. In these families, the syndrome was caused by mutations in this nucleus-encoded gene which deleteriously interacts with the mitochondrial genome. This is the first evidence of the implication of both genomes in a recessive disease.

A nuclear defect in the 4p16 region predisposes to multiple mitochondrial DNA deletions in families with Wolfram syndrome.

Wolfram syndrome is a progressive neurodegenerative disorder transmitted in an autosomal recessive mode. We report two Wolfram syndrome families harboring multiple deletions of mitochondrial DNA. The deletions reached percentages as high as 85-90% in affected tissues such as the central nervous system of one patient, while in other tissues from the same patient and from other members of the family, the percentages of deleted mitochondrial DNA genomes were only 1-10%. Recently, a Wolfram syndrome gene has been linked to markers on 4p16. In both families linkage between the disease locus and 4p16 markers gave a maximum multipoint lod score of 3.79 at theta = 0 (Pi<0.03) with respect to D4S431. In these families, the syndrome was caused by mutations in this nucleus-encoded gene which deleteriously interacts with the mitochondrial genome. This is the first evidence of the implication of both genomes in a recessive disease.

Arginase II Promotes Macrophage Inflammatory Responses Through Mitochondrial Reactive Oxygen Species, Contributing to Insulin Resistance and Atherogenesis.

BACKGROUND: Macrophage-mediated chronic inflammation is mechanistically linked to insulin resistance and atherosclerosis. Although arginase I is considered antiinflammatory, the role of arginase II (Arg-II) in macrophage function remains elusive. This study characterizes the role of Arg-II in macrophage inflammatory responses and its impact on obesity-linked type II diabetes mellitus and atherosclerosis. METHODS AND RESULTS: In human monocytes, silencing Arg-II decreases the monocytes' adhesion to endothelial cells and their production of proinflammatory mediators stimulated by oxidized low-density lipoprotein or lipopolysaccharides, as evaluated by real-time quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Macrophages differentiated from bone marrow cells of Arg-II-deficient (Arg-II(-/-)) mice express lower levels of lipopolysaccharide-induced proinflammatory mediators than do macrophages of wild-type mice. Importantly, reintroducing Arg-II cDNA into Arg-II(-/-) macrophages restores the inflammatory responses, with concomitant enhancement of mitochondrial reactive oxygen species. Scavenging of reactive oxygen species by N-acetylcysteine prevents the Arg-II-mediated inflammatory responses. Moreover, high-fat diet-induced infiltration of macrophages in various organs and expression of proinflammatory cytokines in adipose tissue are blunted in Arg-II(-/-) mice. Accordingly, Arg-II(-/-) mice reveal lower fasting blood glucose and improved glucose tolerance and insulin sensitivity. Furthermore, apolipoprotein E (ApoE)-deficient mice with Arg-II deficiency (ApoE(-/-)Arg-II(-/-)) display reduced lesion size with characteristics of stable plaques, such as decreased macrophage inflammation and necrotic core. In vivo adoptive transfer experiments reveal that fewer donor ApoE(-/-)Arg-II(-/-) than ApoE(-/-)Arg-II(+/+) monocytes infiltrate into the plaque of ApoE(-/-)Arg-II(+/+) mice. Conversely, recipient ApoE(-/-)Arg-II(-/-) mice accumulate fewer donor monocytes than do recipient ApoE(-/-)Arg-II(+/+) animals. CONCLUSIONS: Arg-II promotes macrophage proinflammatory responses through mitochondrial reactive oxygen species, contributing to insulin resistance and atherogenesis. Targeting Arg-II represents a potential therapeutic strategy in type II diabetes mellitus and atherosclerosis. (J Am Heart Assoc. 2012;1:e000992 doi: 10.1161/JAHA.112.000992.).

Time-Dependent Specificity of Immunopathologic (C4d-CD68) and Histologic Criteria of Antibody-Mediated Rejection for Donor-Specific Antibodies and Allograft Dysfunction in Heart Transplantation.

BACKGROUND: In heart transplantation, antibody-mediated rejection (AMR) is diagnosed and graded on the basis of immunopathologic (C4d-CD68) and histopathologic criteria found on endomyocardial biopsies (EMB). Because some pathologic AMR (pAMR) grades may be associated with clinical AMR, and because humoral responses may be affected by the intensity of immunosuppression during the first posttransplantation year, we investigated the incidence and positive predictive values (PPV) of C4d-CD68 and pAMR grades for clinical AMR as a function of time. METHODS: All 564 EMB from 40 adult heart recipients were graded for pAMR during the first posttransplantation year. Clinical AMR was diagnosed by simultaneous occurrence of pAMR on EMB, donor specific antibodies and allograft dysfunction. RESULTS: One patient demonstrated clinical AMR at postoperative day 7 and one at 6 months (1-year incidence 5%). C4d-CD68 was found on 4,7% EMB with a "decrescendo" pattern over time (7% during the first 4 months vs. 1.2% during the last 8 months; P < 0.05). Histopathologic criteria of AMR occurred on 10.3% EMB with no particular time pattern. Only the infrequent (1.4%) pAMR2 grade (simultaneous histopathologic and immunopathologic markers) was predictive for clinical AMR, particularly after the initial postoperative period (first 4 months and last 8 months PPV = 33%-100%; P < 0.05). CONCLUSION: In the first posttransplantation year, AMR immunopathologic and histopathologic markers were relatively frequent, but only their simultaneous occurrence (pAMR2) was predictive of clinical AMR. Furthermore, posttransplantation time may modulate the occurrence of C4d-CD68 on EMB and thus the incidence of pAMR2 and its relevance to the diagnosis of clinical AMR.

Cannabinoid 1 receptor promotes cardiac dysfunction, oxidative stress, inflammation, and fibrosis in diabetic cardiomyopathy.

Endocannabinoids and cannabinoid 1 (CB(1)) receptors have been implicated in cardiac dysfunction, inflammation, and cell death associated with various forms of shock, heart failure, and atherosclerosis, in addition to their recognized role in the development of various cardiovascular risk factors in obesity/metabolic syndrome and diabetes. In this study, we explored the role of CB(1) receptors in myocardial dysfunction, inflammation, oxidative/nitrative stress, cell death, and interrelated signaling pathways, using a mouse model of type 1 diabetic cardiomyopathy. Diabetic cardiomyopathy was characterized by increased myocardial endocannabinoid anandamide levels, oxidative/nitrative stress, activation of p38/Jun NH(2)-terminal kinase (JNK) mitogen-activated protein kinases (MAPKs), enhanced inflammation (tumor necrosis factor-α, interleukin-1β, cyclooxygenase 2, intracellular adhesion molecule 1, and vascular cell adhesion molecule 1), increased expression of CB(1), advanced glycation end product (AGE) and angiotensin II type 1 receptors (receptor for advanced glycation end product [RAGE], angiotensin II receptor type 1 [AT(1)R]), p47(phox) NADPH oxidase subunit, β-myosin heavy chain isozyme switch, accumulation of AGE, fibrosis, and decreased expression of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (SERCA2a). Pharmacological inhibition or genetic deletion of CB(1) receptors attenuated the diabetes-induced cardiac dysfunction and the above-mentioned pathological alterations. Activation of CB(1) receptors by endocannabinoids may play an important role in the pathogenesis of diabetic cardiomyopathy by facilitating MAPK activation, AT(1)R expression/signaling, AGE accumulation, oxidative/nitrative stress, inflammation, and fibrosis. Conversely, CB(1) receptor inhibition may be beneficial in the treatment of diabetic cardiovascular complications.

Subclinical Thyroid Dysfunction and the Risk of Heart Failure in Older Persons at High Cardiovascular Risk.

Context: Subclinical thyroid dysfunction is common in older people. However, its clinical importance is uncertain. Objective: Our objective was to determine the extent to which subclinical hyperthyroidism and hypothyroidism influence the risk of heart failure and cardiovascular diseases in older people. Setting and Design: The Prospective Study of Pravastatin in the Elderly at Risk (PROSPER) is an prospective cohort study. Patients: Patients included men and women aged 70-82 yr (n = 5316) with known cardiovascular risk factors or previous cardiovascular disease. Main Outcome Measures: Incidence rate of heart failure hospitalization, atrial fibrillation, and cardiovascular events and mortality according to baseline thyroid status were evaluated. Euthyroid participants (TSH =0.45-4.5 mIU/liter) were compared with those with subclinical hyperthyroidism (TSH <0.45 mIU/liter) and those with subclinical hypothyroidism (TSH ≥4.5 mIU/liter, both with normal free T(4)). Results: Subclinical hyperthyroidism was present in 71 participants and subclinical hypothyroidism in 199 participants. Over 3.2 yr follow-up, the rate of heart failure was higher for subclinical hyperthyroidism compared with euthyroidism [age- and sex-adjusted hazard ratio (HR) = 2.93, 95% confidence interval (CI) = 1.37-6.24, P = 0.005; multivariate-adjusted HR = 3.27, 95% CI = 1.52-7.02, P = 0.002). Subclinical hypothyroidism (only at threshold >10 mIU/liter) was associated with heart failure (age- and sex-adjusted HR = 3.01, 95% CI = 1.12-8.11, P = 0.029; multivariate HR = 2.28, 95% CI = 0.84-6.23). There were no strong evidence of an association between subclinical thyroid dysfunction and cardiovascular events or mortality, except in those with TSH below 0.1 or over 10 mIU/liter and not taking pravastatin. Conclusion: Older people at high cardiovascular risk with low or very high TSH along with normal free T(4) appear at increased risk of incident heart failure.

The effect of n-3 enriched nutrition therapy on post-operative cognitive dysfunction after cardiac surgery

Background: Postoperative cognitive dysfunction (POCD) occurs frequently after cardiac surgery. Some data suggest that inflammation plays a key role in the development of POCD. N-3 fatty acids have been shown to have a beneficial effect on inflammation. We hypothesised that perioperative n-3 enriched nutrition therapy would reduce the incidence of POCD in this group of patients. Methods: Randomized, double blind placebo controlled trial in patients aged 65 or older undergoing elective cardiac surgery with cardiopulmonary bypass. 2x 250 mL placebo (Ensure Plus™, Abbott Nutrition) or n-3 enriched nutrition therapy (ProSure™ Abbott Nutrition) were administered for ten days starting 5 days prior to surgery. Cognition was assessed preoperatively and 7 days after surgery with the Consortium to Establish a Registry for Alzheimer's Disease - Neuropsychological Assessment Battery (CERAD-NAB) [1]. Results: 16 patients were included. Mean age was 72 } 5.3 for placebo and 75 } 4.8 for ProSure™ respectively. CRP and IL-6 did not differ significantly between groups preoperatively and on postoperative days 1, 3, and 7. Preoperative CERAD total scores were 86 } 10 and 81 } 9 (p = n.s.) for Placebo and ProSure™, respectively. Postoperative scores were 88 } 12, and 77 } 19 (p = n.s.) The change in score was not different between the two groups (Placebo: +3 } 5; ProSure: -5 } 11). Conclusion: In this very small sample no effect of preoperatively started n-3 enriched nutritional supplements on inflammation or cognitive functions were detected. However, there is a large likelihood of a type II error and more patients need to be included to assess possible beneficial effects of this intervention in elderly patients undergoing elective cardiac surgery. 1 Chandler MJ, et al. Neurology. 2005;65:102-6.

Mitochondrial fusion is increased by the nuclear coactivator PGC-1beta

Background There is no evidence to date on whether transcriptional regulators are able to shift the balance between mitochondrial fusion and fission events through selective control of gene expression. Methodology/Principal Findings Here, we demonstrate that reduced mitochondrial size observed in knock-out mice for the transcriptional regulator PGC-1β is associated with a selective reduction in Mitofusin 2 (Mfn2) expression, a mitochondrial fusion protein. This decrease in Mfn2 is specific since expression of the remaining components of mitochondrial fusion and fission machinery were not affected. Furthermore, PGC-1β increases mitochondrial fusion and elongates mitochondrial tubules. This PGC-1β-induced elongation specifically requires Mfn2 as this process is absent in Mfn2-ablated cells. Finally, we show that PGC-1β increases Mfn2 promoter activity and transcription by coactivating the nuclear receptor Estrogen Related Receptor α (ERRα). Conclusions/Significance Taken together, our data reveal a novel mechanism by which mammalian cells control mitochondrial fusion. In addition, we describe a novel role of PGC-1β in mitochondrial physiology, namely the control of mitochondrial fusion mainly through Mfn2.

In situ fluorescence imaging of glutamate-evoked mitochondrial Na+ responses in astrocytes.

Astrocytes can experience large intracellular Na+ changes following the activation of the Na+-coupled glutamate transport. The present study investigated whether cytosolic Na+ changes are transmitted to mitochondria, which could therefore influence their function and contribute to the overall intracellular Na+ regulation. Mitochondrial Na+ (Na+(mit)) changes were monitored using the Na+-sensitive fluorescent probe CoroNa Red (CR) in intact primary cortical astrocytes, as opposed to the classical isolated mitochondria preparation. The mitochondrial localization and Na+ sensitivity of the dye were first verified and indicated that it can be safely used as a selective Na+(mit) indicator. We found by simultaneously monitoring cytosolic and mitochondrial Na+ using sodium-binding benzofuran isophthalate and CR, respectively, that glutamate-evoked cytosolic Na+ elevations are transmitted to mitochondria. The resting Na+(mit) concentration was estimated at 19.0 +/- 0.8 mM, reaching 30.1 +/- 1.2 mM during 200 microM glutamate application. Blockers of conductances potentially mediating Na+ entry (calcium uniporter, monovalent cation conductances, K+(ATP) channels) were not able to prevent the Na+(mit) response to glutamate. However, Ca2+ and its exchange with Na+ appear to play an important role in mediating mitochondrial Na+ entry as chelating intracellular Ca2+ with BAPTA or inhibiting Na+/Ca2+ exchanger with CGP-37157 diminished the Na+(mit) response. Moreover, intracellular Ca2+ increase achieved by photoactivation of caged Ca2+ also induced a Na+(mit) elevation. Inhibition of mitochondrial Na/H antiporter using ethylisopropyl-amiloride caused a steady increase in Na+(mit) without increasing cytosolic Na+, indicating that Na+ extrusion from mitochondria is mediated by these exchangers. Thus, mitochondria in intact astrocytes are equipped to efficiently sense cellular Na+ signals and to dynamically regulate their Na+ content.

Mitochondrial DNA of Nellore and European x Nellore crossing cattle of high performance

The objective of this work was to evaluate, through a polymorphism in the ND5 gene of the bovine mitochondrial DNA, the frequency of Bos taurus indicus mtDNA individuals in a sample of Nellore purebred origin animals (n = 69) and crossbred animals originated from crosses of European sires and Nellore purebred origin females (n = 275). Only 2.26% (8/354) of the animals presented Bos taurus indicus mtDNA. The high frequency of Bos taurus taurus mtDNA in these animals can be a consequence of selection, once the animals studied are originated from selected lineages of high performance for meat production.

Subclinical thyroid dysfunction and the risk for fractures: a systematic review and meta-analysis.

BACKGROUND: Data on the association between subclinical thyroid dysfunction and fractures conflict. PURPOSE: To assess the risk for hip and nonspine fractures associated with subclinical thyroid dysfunction among prospective cohorts. DATA SOURCES: Search of MEDLINE and EMBASE (1946 to 16 March 2014) and reference lists of retrieved articles without language restriction. STUDY SELECTION: Two physicians screened and identified prospective cohorts that measured thyroid function and followed participants to assess fracture outcomes. DATA EXTRACTION: One reviewer extracted data using a standardized protocol, and another verified data. Both reviewers independently assessed methodological quality of the studies. DATA SYNTHESIS: The 7 population-based cohorts of heterogeneous quality included 50,245 participants with 1966 hip and 3281 nonspine fractures. In random-effects models that included the 5 higher-quality studies, the pooled adjusted hazard ratios (HRs) of participants with subclinical hyperthyroidism versus euthyrodism were 1.38 (95% CI, 0.92 to 2.07) for hip fractures and 1.20 (CI, 0.83 to 1.72) for nonspine fractures without statistical heterogeneity (P = 0.82 and 0.52, respectively; I2= 0%). Pooled estimates for the 7 cohorts were 1.26 (CI, 0.96 to 1.65) for hip fractures and 1.16 (CI, 0.95 to 1.42) for nonspine fractures. When thyroxine recipients were excluded, the HRs for participants with subclinical hyperthyroidism were 2.16 (CI, 0.87 to 5.37) for hip fractures and 1.43 (CI, 0.73 to 2.78) for nonspine fractures. For participants with subclinical hypothyroidism, HRs from higher-quality studies were 1.12 (CI, 0.83 to 1.51) for hip fractures and 1.04 (CI, 0.76 to 1.42) for nonspine fractures (P for heterogeneity = 0.69 and 0.88, respectively; I2 = 0%). LIMITATIONS: Selective reporting cannot be excluded. Adjustment for potential common confounders varied and was not adequately done across all studies. CONCLUSION: Subclinical hyperthyroidism might be associated with an increased risk for hip and nonspine fractures, but additional large, high-quality studies are needed. PRIMARY FUNDING SOURCE: Swiss National Science Foundation.

Mitochondrial fatty acid oxidation in obesity

Significance: Current lifestyles with high-energy diets and little exercise are triggering an alarming growth in obesity. Excess of adiposity is leading to severe increases in associated pathologies, such as insulin resistance, type 2 diabetes, atherosclerosis, cancer, arthritis, asthma, and hypertension. This, together with the lack of efficient obesity drugs, is the driving force behind much research. Recent Advances: Traditional anti-obesity strategies focused on reducing food intake and increasing physical activity. However, recent results suggest that enhancing cellular energy expenditure may be an attractive alternative therapy. Critical Issues: This review evaluates recent discoveries regarding mitochondrial fatty acid oxidation (FAO) and its potential as a therapy for obesity. We focus on the still controversial beneficial effects of increased FAO in liver and muscle, recent studies on how to potentiate adipose tissue energy expenditure, and the different hypotheses involving FAO and the reactive oxygen species production in the hypothalamic control of food intake. Future Directions: The present review aims to provide an overview of novel anti-obesity strategies that target mitochondrial FAO and that will definitively be of high interest in the future research to fight against obesity-related disorders.

How to interprete subclinical thyroid dysfunction in the prevention and management of heart failure events? Individual participant data analysis from six prospective cohorts

Background: Guidelines of the Diagnosis and Management of Heart Failure (HF) recommend investigating exacerbating conditions, such as thyroid dysfunction, but without specifying impact of different TSH levels. Limited prospective data exist regarding the association between subclinical thyroid dysfunction and HF events. Methods: We performed a pooled analysis of individual participant data using all available prospective cohorts with thyroid function tests and subsequent follow-up of HF events. Individual data on 25,390 participants with 216,247 person-years of follow-up were supplied from 6 prospective cohorts in the United States and Europe. Euthyroidism was defined as TSH 0.45-4.49 mIU/L, subclinical hypothyroidism as TSH 4.5-19.9 mIU/L and subclinical hyperthyroidism as TSH <0.45 mIU/L, both with normal free thyroxine levels. HF events were defined as acute HF events, hospitalization or death related to HF events. Results: Among 25,390 participants, 2068 had subclinical hypothyroidism (8.1%) and 648 subclinical hyperthyroidism (2.6%). In age- and gender-adjusted analyses, risks of HF events were increased with both higher and lower TSH levels (P for quadratic pattern<0.01): hazard ratio (HR) was 1.01 (95% confidence interval [CI] 0.81-1.26) for TSH 4.5-6.9 mIU/L, 1.65 (CI 0.84-3.23) for TSH 7.0-9.9 mIU/L, 1.86 (CI 1.27-2.72) for TSH 10.0-19.9 mIUL/L (P for trend <0.01), and was 1.31 (CI 0.88-1.95) for TSH 0.10-0.44 mIU/L and 1.94 (CI 1.01-3.72) for TSH <0.10 mIU/L (P for trend=0.047). Risks remained similar after adjustment for cardiovascular risk factors. Conclusion: Risks of HF events were increased with both higher and lower TSH levels, particularly for TSH ≥10 mIU/L and for TSH <0.10 mIU/L. Our findings might help to interpret TSH levels in the prevention and investigation of HF.