Altered hepatic lipid droplet morphology and lipid metabolism in fasted Plin2-null mice.

Perilipin 2 (Plin2) binds to the surface of hepatic lipid droplets (LDs) with expression levels that correlate with triacylglyceride (TAG) content. We investigated if Plin2 is important for hepatic LD storage in fasted or high-fat diet-induced obese Plin2+/+ and Plin2-/- mice. Plin2-/- mice had comparable body weights, metabolic phenotype, glucose tolerance, and circulating TAG and total cholesterol levels compared with Plin2+/+ mice, regardless of the dietary regime. Both fasted and high-fat fed Plin2-/- mice stored reduced levels of hepatic TAG compared with Plin2+/+ mice. Fasted Plin2-/- mice stored fewer but larger hepatic LDs compared with Plin2+/+ mice. Detailed hepatic lipid analysis showed substantial reductions in accumulated TAG species in fasted Plin2-/- mice compared with Plin2+/+ mice, whereas cholesteryl esters and phosphatidylcholines were increased. RNA-Seq revealed minor differences in hepatic gene expression between fed Plin2+/+ and Plin2-/- mice, in contrast to marked differences in gene expression between fasted Plin2+/+ and Plin2-/- mice. Our findings demonstrate that Plin2 is required to regulate hepatic LD size and storage of neutral lipid species in the fasted state, while its role in obesity-induced steatosis is less clear.

Mechanisms Behind NAFLD: a System Genetics Perspective.

PURPOSE OF REVIEW: To summarize the key factors contributing to the onset and progress of nonalcoholic fatty liver disease (NAFLD) and put them in a system genetics context. We particularly focus on how genetic regulation of hepatic lipids contributes to NAFLD. RECENT FINDINGS: NAFLD is characterized by excessive accumulation of fat in the liver. This can progress to steatohepatitis (inflammation and hepatocyte injury) and eventually, cirrhosis. The severity of NAFLD is determined by a combination of factors including obesity, insulin resistance, and lipotoxic lipids, along with genetic susceptibility. Numerous studies have been conducted on large human cohorts and mouse panels, to identify key determinants in the genome, transcriptome, proteome, lipidome, microbiome and different environmental conditions contributing to NAFLD. We review common factors contributing to NAFLD and put them in a systems genetics context. In particular, we describe how genetic regulation of liver lipids contributes to NAFLD. The combination of an unhealthy lifestyle and genetic predisposition increases the likelihood of accumulating lipotoxic specie lipids that may be one of the driving forces behind developing severe forms of NAFLD.

Conserved multi-tissue transcriptomic adaptations to exercise training in humans and mice.

Physical activity is associated with beneficial adaptations in human and rodent metabolism. We studied over 50 complex traits before and after exercise intervention in middle-aged men and a panel of 100 diverse strains of female mice. Candidate gene analyses in three brain regions, muscle, liver, heart, and adipose tissue of mice indicate genetic drivers of clinically relevant traits, including volitional exercise volume, muscle metabolism, adiposity, and hepatic lipids. Although ∼33% of genes differentially expressed in skeletal muscle following the exercise intervention are similar in mice and humans independent of BMI, responsiveness of adipose tissue to exercise-stimulated weight loss appears controlled by species and underlying genotype. We leveraged genetic diversity to generate prediction models of metabolic trait responsiveness to volitional activity offering a framework for advancing personalized exercise prescription. The human and mouse data are publicly available via a user-friendly Web-based application to enhance data mining and hypothesis development.

Consumption of salmon fishmeal increases hepatic cholesterol content in obese C57BL/6 J mice.

PURPOSE: By-products from farmed fish contain large amounts of proteins and may be used for human consumption. The purpose of this study was to investigate cardiometabolic effects and metabolic tolerance in mice consuming fishmeal from salmon by-products, salmon filet or beef. METHODS: Female C57BL/6J mice were fed chow, as a healthy reference group, or a high-fat diet for 10 weeks to induce obesity and glucose intolerance. Obese mice were subsequently given isocaloric diets containing 50% of the dietary protein from salmon fishmeal, salmon filet or beef for 10 weeks. Mice were subjected to metabolic phenotyping, which included measurements of body composition, energy metabolism in metabolic cages and glucose tolerance. Lipid content and markers of hepatic toxicity were determined in plasma and liver. Hepatic gene and protein expression was determined with RNA sequencing and immunoblotting. RESULTS: Mice fed fishmeal, salmon filet or beef had similar food intake, energy consumption, body weight gain, adiposity, glucose tolerance and circulating levels of lipids and hepatic toxicity markers, such as p-ALT and p-AST. Fishmeal increased hepatic cholesterol levels by 35-36% as compared to salmon filet (p = 0.0001) and beef (p = 0.005). This was accompanied by repressed expression of genes involved in steroid and cholesterol metabolism and reduced levels of circulating Pcsk9. CONCLUSION: Salmon fishmeal was well tolerated, but increased hepatic cholesterol content. The high cholesterol content in fishmeal may be responsible for the effects on hepatic cholesterol metabolism. Before introducing fishmeal from salmon by-products as a dietary component, it may be advantageous to reduce the cholesterol content in fishmeal.

Potential Mechanisms for How Long-Term Physical Activity May Reduce Insulin Resistance.

Insulin became available for the treatment of patients with diabetes 100 years ago, and soon thereafter it became evident that the biological response to its actions differed markedly between individuals. This prompted extensive research into insulin action and resistance (IR), resulting in the universally agreed fact that IR is a core finding in patients with type 2 diabetes mellitus (T2DM). T2DM is the most prevalent form of diabetes, reaching epidemic proportions worldwide. Physical activity (PA) has the potential of improving IR and is, therefore, a cornerstone in the prevention and treatment of T2DM. Whereas most research has focused on the acute effects of PA, less is known about the effects of long-term PA on IR. Here, we describe a model of potential mechanisms behind reduced IR after long-term PA to guide further mechanistic investigations and to tailor PA interventions in the therapy of T2DM. The development of such interventions requires knowledge of normal glucose metabolism, and we briefly summarize an integrated physiological perspective on IR. We then describe the effects of long-term PA on signaling molecules involved in cellular responses to insulin, tissue-specific functions, and whole-body IR.

Serglycin Is Involved in Adipose Tissue Inflammation in Obesity.

Chronic local inflammation of adipose tissue is an important feature of obesity. Serglycin is a proteoglycan highly expressed by various immune cell types known to infiltrate adipose tissue under obese conditions. To investigate if serglycin expression has an impact on diet-induced adipose tissue inflammation, we subjected Srgn +/+ and Srgn -/- mice (C57BL/6J genetic background) to an 8-wk high-fat and high-sucrose diet. The total body weight was the same in Srgn +/+ and Srgn -/- mice after diet treatment. Expression of white adipose tissue genes linked to inflammatory pathways were lower in Srgn -/- mice. We also noted reduced total macrophage abundance, a reduced proportion of proinflammatory M1 macrophages, and reduced formation of crown-like structures in adipose tissue of Srgn -/- compared with Srgn +/+ mice. Further, Srgn -/- mice had more medium-sized adipocytes and fewer large adipocytes. Differentiation of preadipocytes into adipocytes (3T3-L1) was accompanied by reduced Srgn mRNA expression. In line with this, analysis of single-cell RNA sequencing data from mouse and human adipose tissue supports that Srgn mRNA is predominantly expressed by various immune cells, with low expression in adipocytes. Srgn mRNA expression was higher in obese compared with lean humans and mice, accompanied by an increased expression of immune cell gene markers. SRGN and inflammatory marker mRNA expression was reduced upon substantial weight loss in patients after bariatric surgery. Taken together, this study introduces a role for serglycin in the regulation of obesity-induced adipose inflammation.

The effect of toll-like receptor ligands on energy metabolism and myokine expression and secretion in cultured human skeletal muscle cells.

Skeletal muscle plays an important role in glycaemic control and metabolic homeostasis, making it a tissue of interest with respect to type 2 diabetes mellitus. The aim of the present study was to determine if ligands of Toll-like receptors (TLRs) could have an impact on energy metabolism and myokine expression and secretion in cultured human skeletal muscle cells. The myotubes expressed mRNA for TLRs 1-6. TLR3, TLR4, TLR5 and TLR6 ligands (TLRLs) increased glucose metabolism. Furthermore, TLR4L and TLR5L increased oleic acid metabolism. The metabolic effects of TLRLs were not evident until after at least 24 h pre-incubation of the cells and here the metabolic effects were more evident for the metabolism of glucose than oleic acid, with a shift towards effects on oleic acid metabolism after chronic exposure (168 h). However, the stimulatory effect of TLRLs on myokine expression and secretion was detected after only 6 h, where TLR3-6L stimulated secretion of interleukin-6 (IL-6). TLR5L also increased secretion of interleukin-8 (IL-8), while TLR6L also increased secretion of granulocyte-macrophage colony stimulating factor (GM-CSF). Pre-incubation of the myotubes with IL-6 for 24 h increased oleic acid oxidation but had no effect on glucose metabolism. Thus IL-6 did not mimic all the metabolic effects of the TLRLs, implying metabolic effects beyond the actions of this myokine.

Effect of voluntary exercise upon the metabolic syndrome and gut microbiome composition in mice.

The metabolic syndrome is a cluster of conditions that increase an individual's risk of developing diseases. Being physically active throughout life is known to reduce the prevalence and onset of some aspects of the metabolic syndrome. Furthermore, previous studies have demonstrated that an individual's gut microbiome composition has a large influence on several aspects of the metabolic syndrome. However, the mechanism(s) by which physical activity may improve metabolic health are not well understood. We sought to determine if endurance exercise is sufficient to prevent or ameliorate the development of the metabolic syndrome and its associated diseases. We also analyzed the impact of physical activity under metabolic syndrome progression upon the gut microbiome composition. Utilizing whole-body low-density lipoprotein receptor (LDLR) knockout mice on a "Western Diet," we show that long-term exercise acts favorably upon glucose tolerance, adiposity, and liver lipids. Exercise increased mitochondrial abundance in skeletal muscle but did not reduce liver fibrosis, aortic lesion area, or plasma lipids. Lastly, we observed several changes in gut bacteria and their novel associations with metabolic parameters of clinical importance. Altogether, our results indicate that exercise can ameliorate some aspects of the metabolic syndrome progression and alter the gut microbiome composition.

Effect of oral and transdermal oestrogen therapy on bone mineral density in functional hypothalamic amenorrhoea: a systematic review and meta-analysis.

BACKGROUND: Female athletes might develop reduced bone mineral density (BMD) and amenorrhoea due to low energy intake. OBJECTIVE: To systematically review the literature of randomised controlled trials (RCTs) assessing the effect of oestrogen oral contraceptives (OCP), conjugated oestrogens (CE) and transdermal estradiol (TE) on BMD in premenopausal women with functional hypothalamic amenorrhoea (FHA) due to weight loss, vigorous exercise and/or stress. METHODS: A comprehensive literature search in PubMed, MEDLINE, Cochrane Library, Ovid and CINAHL from inception to 1 October 2020. DATA EXTRACTION AND SYNTHESIS: Two authors independently extracted data. When possible, the data were pooled in a random-effects meta-analysis. MAIN OUTCOMES: Difference in BMD (g/cm2) at the lumbar spine. RESULTS: Nine RCTs comprising 770 participants met the inclusion criteria; five studies applied OCP, two CE and two TE. Four RCTs (two OCP, two TE) found an increased BMD in premenopausal women with FHA, and five (three OCP, two CE) found a decreased BMD compared with controls. A meta-analysis showed no difference in BMD between the treatment and control groups, (standardised mean difference (SMD) 0.30, 95% CI -0.12 to 0.73). A secondary analysis for change scores from baseline to first assessment point, showed a similar overall result (SMD 0.17, 95% CI -0.16 to 0.51). No serious adverse events were reported. CONCLUSION: The literature suggests that TE might increase lumbar BMD in premenopausal women with FHA, but pooled results revealed no effect of the intervention. The findings do not support oestrogen therapy to improve BMD in these patient groups.

Plin2 deletion increases cholesteryl ester lipid droplet content and disturbs cholesterol balance in adrenal cortex.

Cholesteryl esters (CEs) are the water-insoluble transport and storage form of cholesterol. Steroidogenic cells primarily store CEs in cytoplasmic lipid droplet (LD) organelles, as contrasted to the majority of mammalian cell types that predominantly store triacylglycerol (TAG) in LDs. The LD-binding Plin2 binds to both CE- and TAG-rich LDs, and although Plin2 is known to regulate degradation of TAG-rich LDs, its role for regulation of CE-rich LDs is unclear. To investigate the role of Plin2 in the regulation of CE-rich LDs, we performed histological and molecular characterization of adrenal glands from Plin2+/+ and Plin2-/- mice. Adrenal glands of Plin2-/- mice had significantly enlarged organ size, increased size and numbers of CE-rich LDs in cortical cells, elevated cellular unesterified cholesterol levels, and increased expression of macrophage markers and genes facilitating reverse cholesterol transport. Despite altered LD storage, mobilization of adrenal LDs and secretion of corticosterone induced by adrenocorticotropic hormone stimulation or starvation were similar in Plin2+/+ and Plin2-/- mice. Plin2-/- adrenals accumulated ceroid-like structures rich in multilamellar bodies in the adrenal cortex-medulla boundary, which increased with age, particularly in females. Finally, Plin2-/- mice displayed unexpectedly high levels of phosphatidylglycerols, which directly paralleled the accumulation of these ceroid-like structures. Our findings demonstrate an important role of Plin2 for regulation of CE-rich LDs and cellular cholesterol balance in the adrenal cortex.

Genetic regulation of liver lipids in a mouse model of insulin resistance and hepatic steatosis.

To elucidate the contributions of specific lipid species to metabolic traits, we integrated global hepatic lipid data with other omics measures and genetic data from a cohort of about 100 diverse inbred strains of mice fed a high-fat/high-sucrose diet for 8 weeks. Association mapping, correlation, structure analyses, and network modeling revealed pathways and genes underlying these interactions. In particular, our studies lead to the identification of Ifi203 and Map2k6 as regulators of hepatic phosphatidylcholine homeostasis and triacylglycerol accumulation, respectively. Our analyses highlight mechanisms for how genetic variation in hepatic lipidome can be linked to physiological and molecular phenotypes, such as microbiota composition.

Progress and Challenges in the Biology of FNDC5 and Irisin.

In 2002, a transmembrane protein-now known as FNDC5-was discovered and shown to be expressed in skeletal muscle, heart, and brain. It was virtually ignored for 10 years, until a study in 2012 proposed that, in response to exercise, the ectodomain of skeletal muscle FNDC5 was cleaved, traveled to white adipose tissue, and induced browning. The wasted energy of this browning raised the possibility that this myokine, named irisin, might mediate some beneficial effects of exercise. Since then, more than 1000 papers have been published exploring the roles of irisin. A major interest has been on adipose tissue and metabolism, following up the major proposal from 2012. Many studies correlating plasma irisin levels with physiological conditions have been questioned for using flawed assays for irisin concentration. However, experiments altering irisin levels by injecting recombinant irisin or by gene knockout are more promising. Recent discoveries have suggested potential roles of irisin in bone remodeling and in the brain, with effects potentially related to Alzheimer's disease. We discuss some discrepancies between research groups and the mechanisms that are yet to be determined. Some important questions raised in the initial discovery of irisin, such as the role of the mutant start codon of human FNDC5 and the mechanism of ectodomain cleavage, remain to be answered. Apart from these specific questions, a promising new tool has been developed-mice with a global or tissue-specific knockout of FNDC5. In this review, we critically examine the current knowledge and delineate potential solutions to resolve existing ambiguities.

Branched-chain amino acid metabolism, insulin sensitivity and liver fat response to exercise training in sedentary dysglycaemic and normoglycaemic men.

AIMS/HYPOTHESIS: Obesity and insulin resistance may be associated with elevated plasma concentration of branched-chain amino acids (BCAAs) and impaired BCAA metabolism. However, it is unknown whether the insulin-sensitising effect of long-term exercise can be explained by concomitant change in BCAAs and their metabolism. METHODS: We included 26 sedentary overweight and normal-weight middle-aged men from the MyoGlu clinical trial, with or without dysglycaemia, for 12 weeks of supervised intensive exercise intervention, including two endurance and two resistance sessions weekly. Insulin sensitivity was measured as the glucose infusion rate (GIR) from a hyperinsulinaemic-euglycaemic clamp. In addition, maximum oxygen uptake, upper and lower body strength and adipose tissue depots (using MRI and spectroscopy) were measured, and subcutaneous white adipose tissue (ScWAT) and skeletal muscle (SkM) biopsies were harvested both before and after the 12 week intervention. In the present study we have measured plasma BCAAs and related metabolites using CG-MS/MS and HPLC-MS/MS, and performed global mRNA-sequencing pathway analysis on ScWAT and SkM. RESULTS: In MyoGlu, men with dysglycaemia displayed lower GIR, more fat mass and higher liver fat content than normoglycaemic men at baseline, and 12 weeks of exercise increased GIR, improved body composition and reduced liver fat content similarly for both groups. In our current study we observed higher plasma concentrations of BCAAs (14.4%, p = 0.01) and related metabolites, such as 3-hydroxyisobutyrate (19.4%, p = 0.034) in dysglycaemic vs normoglycaemic men at baseline. Baseline plasma BCAA levels correlated negatively to the change in GIR (ρ = -0.41, p = 0.037) and [Formula: see text] (ρ = -0.47, p = 0.015) after 12 weeks of exercise and positively to amounts of intraperitoneal fat (ρ = 0.40, p = 0.044) and liver fat (ρ = 0.58, p = 0.01). However, circulating BCAAs and related metabolites did not respond to 12 weeks of exercise, with the exception of isoleucine, which increased in normoglycaemic men (10 µmol/l, p = 0.01). Pathway analyses of mRNA-sequencing data implied reduced BCAA catabolism in both SkM and ScWAT in men with dysglycaemia compared with men with normoglycaemia at baseline. Gene expression levels related to BCAA metabolism correlated positively with GIR and markers of mitochondrial content in both SkM and ScWAT, and negatively with fat mass generally, and particularly with intraperitoneal fat mass. mRNA-sequencing pathway analysis also implied increased BCAA metabolism after 12 weeks of exercise in both groups and in both tissues, including enhanced expression of the gene encoding branched-chain α-ketoacid dehydrogenase (BCKDH) and reduced expression of the BCKDH phosphatase in both groups and tissues. Gene expression of SLC25A44, which encodes a mitochondrial BCAA transporter, was increased in SkM in both groups, and gene expression of BCKDK, which encodes BCKDH kinase, was reduced in ScWAT in dysglycaemic men. Mediation analyses indicated a pronounced effect of enhanced SkM (~53%, p = 0.022), and a moderate effect of enhanced ScWAT (~18%, p = 0.018) BCAA metabolism on improved insulin sensitivity after 12 weeks of exercise, based on mRNA sequencing. In comparison, plasma concentration of BCAAs did not mediate any effect in this regard. CONCLUSION/INTERPRETATION: Plasma BCAA concentration was largely unresponsive to long-term exercise and unrelated to exercise-induced insulin sensitivity. On the other hand, the insulin-sensitising effect of long-term exercise in men may be explained by enhanced SkM and, to a lesser degree, also by enhanced ScWAT BCAA catabolism. Graphical abstract.

Isolated Plin5-deficient cardiomyocytes store less lipid droplets than normal, but without increased sensitivity to hypoxia.

Plin5 is abundantly expressed in the heart where it binds to lipid droplets (LDs) and facilitates physical interaction between LDs and mitochondria. We isolated cardiomyocytes from adult Plin5+/+ and Plin5-/- mice to study the role of Plin5 for fatty acid uptake, LD accumulation, fatty acid oxidation, and tolerance to hypoxia. Cardiomyocytes isolated from Plin5-/- mice cultured with oleic acid stored less LDs than Plin5+/+, but comparable levels to Plin5+/+ cardiomyocytes when adipose triglyceride lipase activity was inhibited. The ability to oxidize fatty acids into CO2 was similar between Plin5+/+ and Plin5-/- cardiomyocytes, but Plin5-/- cardiomyocytes had a transient increase in intracellular fatty acid oxidation intermediates. After pre-incubation with oleic acids, Plin5-/- cardiomyocytes retained a higher content of glycogen and showed improved tolerance to hypoxia compared to Plin5+/+. In isolated, perfused hearts, deletion of Plin5 had no important effect on ventricular pressures or infarct size after ischemia. Old Plin5-/- mice had reduced levels of cardiac triacylglycerides, increased heart weight, and apart from modest elevated expression of mRNAs for beta myosin heavy chain Myh7 and the fatty acid transporter Cd36, other genes involved in fatty acid oxidation, glycogen metabolism and glucose utilization were essentially unchanged by removal of Plin5. Plin5 seems to facilitate cardiac LD storage primarily by repressing adipose triglyceride lipase activity without altering cardiac fatty acid oxidation capacity. Expression of Plin5 and cardiac LD content of isolated cardiomyocytes has little importance for tolerance to acute hypoxia and ischemia, which contrasts the protective role for Plin5 in mouse models during myocardial ischemia.

Estrogen receptor α controls metabolism in white and brown adipocytes by regulating Polg1 and mitochondrial remodeling.

Obesity is heightened during aging, and although the estrogen receptor α (ERα) has been implicated in the prevention of obesity, its molecular actions in adipocytes remain inadequately understood. Here, we show that adipose tissue ESR1/Esr1 expression inversely associated with adiposity and positively associated with genes involved in mitochondrial metabolism and markers of metabolic health in 700 Finnish men and 100 strains of inbred mice from the UCLA Hybrid Mouse Diversity Panel. To determine the anti-obesity actions of ERα in fat, we selectively deleted Esr1 from white and brown adipocytes in mice. In white adipose tissue, Esr1 controlled oxidative metabolism by restraining the targeted elimination of mitochondria via the E3 ubiquitin ligase parkin. mtDNA content was elevated, and adipose tissue mass was reduced in adipose-selective parkin knockout mice. In brown fat centrally involved in body temperature maintenance, Esr1 was requisite for both mitochondrial remodeling by dynamin-related protein 1 (Drp1) and uncoupled respiration thermogenesis by uncoupled protein 1 (Ucp1). In both white and brown fat of female mice and adipocytes in culture, mitochondrial dysfunction in the context of Esr1 deletion was paralleled by a reduction in the expression of the mtDNA polymerase γ subunit Polg1 We identified Polg1 as an ERα target gene by showing that ERα binds the Polg1 promoter to control its expression in 3T3L1 adipocytes. These findings support strategies leveraging ERα action on mitochondrial function in adipocytes to combat obesity and metabolic dysfunction.

Effects of dietary methionine and cysteine restriction on plasma biomarkers, serum fibroblast growth factor 21, and adipose tissue gene expression in women with overweight or obesity: a double-blind randomized controlled pilot study.

BACKGROUND: Dietary restriction of methionine and cysteine is a well-described model that improves metabolic health in rodents. To investigate the translational potential in humans, we evaluated the effects of dietary methionine and cysteine restriction on cardiometabolic risk factors, plasma and urinary amino acid profile, serum fibroblast growth factor 21 (FGF21), and subcutaneous adipose tissue gene expression in women with overweight and obesity in a double-blind randomized controlled pilot study. METHODS: Twenty women with overweight or obesity were allocated to a diet low (Met/Cys-low, n = 7), medium (Met/Cys-medium, n = 7) or high (Met/Cys-high, n = 6) in methionine and cysteine for 7 days. The diets differed only by methionine and cysteine content. Blood and urine were collected at day 0, 1, 3 and 7 and subcutaneous adipose tissue biopsies were taken at day 0 and 7. RESULTS: Plasma methionine and cystathionine and urinary total cysteine decreased, whereas FGF21 increased in the Met/Cys-low vs. Met/Cys-high group. The Met/Cys-low group had increased mRNA expression of lipogenic genes in adipose tissue including DGAT1. When we excluded one participant with high fasting insulin at baseline, the Met/Cys-low group showed increased expression of ACAC, DGAT1, and tendencies for increased expression of FASN and SCD1 compared to the Met/Cys-high group. The participants reported satisfactory compliance and that the diets were moderately easy to follow. CONCLUSIONS: Our data suggest that dietary methionine and cysteine restriction may have beneficial effects on circulating biomarkers, including FGF21, and influence subcutaneous adipose tissue gene expression. These results will aid in the design and implementation of future large-scale dietary interventions with methionine and cysteine restriction. Trial registration ClinicalTrials.gov Identifier: NCT03629392, registration date: 14/08/2018 https://clinicaltrials.gov/ct2/show/NCT03629392.

Effects of long-term exercise on plasma adipokine levels and inflammation-related gene expression in subcutaneous adipose tissue in sedentary dysglycaemic, overweight men and sedentary normoglycaemic men of healthy weight.

AIMS/HYPOTHESIS: Obesity and insulin resistance may be associated with altered expression and secretion of adipokines. Physical activity can markedly improve insulin sensitivity, but the association with adipokines remains largely unknown. In this study, we examined the effects of physical activity on the subcutaneous white adipose tissue (scWAT) secretome and its relationship to insulin sensitivity. METHODS: As reported previously, we enrolled 26 sedentary, middle-aged men (13 dysglycaemic and overweight; 13 normoglycaemic and of healthy weight) into a 12 week, supervised, intensive physical exercise intervention that included two endurance and two resistance sessions each week. Insulin sensitivity was measured as the glucose infusion rate from a euglycaemic-hyperinsulinaemic clamp. In our previous study, we measured maximum oxygen uptake, upper- and lower-body strength and a range of circulating biomarkers, and quantified adipose tissue depots using MRI and magnetic resonance spectroscopy. We have now performed global mRNA sequencing, microarrays and RT-PCR of scWAT and skeletal muscle biopsies, and quantified selected plasma adipokines by ELISA. RESULTS: Insulin sensitivity increased similarly in both dysglycaemic (45%) and normoglycaemic (38%) men after 12 weeks of exercise, as reported previously. mRNA sequencing of scWAT revealed 90 transcripts that responded to exercise in dysglycaemic men, whereas only marginal changes were observed in normoglycaemic men. These results were validated using microarrays and RT-PCR. A total of 62 out of 90 transcripts encoded secreted proteins. Overall, 17 transcripts were upregulated and 73 transcripts were downregulated. Downregulated transcripts included several macrophage markers, and were associated with inflammatory and immune-related pathways. Levels of these immune-related transcripts were enhanced in dysglycaemic men vs normoglycaemic men at baseline, but were normalised after the exercise intervention. Principal component and correlation analyses revealed inverse correlations between levels of these immune-related transcripts and insulin sensitivity at baseline, after the intervention, and for the change between baseline and after the intervention. In addition, levels of these transcripts at baseline could predict exercise-induced improvements in insulin sensitivity. Adipokine levels in scWAT (but not in skeletal muscle) were significantly correlated with corresponding plasma adipokine concentrations, as exemplified by leptin, high-molecular-weight adiponectin and secreted frizzled-related protein 4 (SFRP4). SFRP4 mRNA was the most exercise-responsive transcript in scWAT from dysglycaemic men, and plasma SFRP4 concentrations were reduced in dysglycaemic men, but not in normoglycaemic men, after 12 weeks of exercise. CONCLUSIONS/INTERPRETATION: This study indicates that scWAT may be an important mediator of exercise-induced improvements in insulin sensitivity, especially in overweight dysglycaemic individuals at increased risk of developing type 2 diabetes.

Gene-by-Sex Interactions in Mitochondrial Functions and Cardio-Metabolic Traits.

We studied sex differences in over 50 cardio-metabolic traits in a panel of 100 diverse inbred strains of mice. The results clearly showed that the effects of sex on both clinical phenotypes and gene expression depend on the genetic background. In support of this, genetic loci associated with the traits frequently showed sex specificity. For example, Lyplal1, a gene implicated in human obesity, was shown to underlie a sex-specific locus for diet-induced obesity. Global gene expression analyses of tissues across the panel implicated adipose tissue "beiging" and mitochondrial functions in the sex differences. Isolated mitochondria showed gene-by-sex interactions in oxidative functions, such that some strains (C57BL/6J) showed similar function between sexes, whereas others (DBA/2J and A/J) showed increased function in females. Reduced adipose mitochondrial function in males as compared to females was associated with increased susceptibility to obesity and insulin resistance. Gonadectomy studies indicated that gonadal hormones acting in a tissue-specific manner were responsible in part for the sex differences.

The impact of exercise on mitochondrial dynamics and the role of Drp1 in exercise performance and training adaptations in skeletal muscle.

OBJECTIVE: Mitochondria are organelles primarily responsible for energy production, and recent evidence indicates that alterations in size, shape, location, and quantity occur in response to fluctuations in energy supply and demand. We tested the impact of acute and chronic exercise on mitochondrial dynamics signaling and determined the impact of the mitochondrial fission regulator Dynamin related protein (Drp)1 on exercise performance and muscle adaptations to training. METHODS: Wildtype and muscle-specific Drp1 heterozygote (mDrp1+/-) mice, as well as dysglycemic (DG) and healthy normoglycemic men (control) performed acute and chronic exercise. The Hybrid Mouse Diversity Panel, including 100 murine strains of recombinant inbred mice, was used to identify muscle Dnm1L (encodes Drp1)-gene relationships. RESULTS: Endurance exercise impacted all aspects of the mitochondrial life cycle, i.e. fission-fusion, biogenesis, and mitophagy. Dnm1L gene expression and Drp1Ser616 phosphorylation were markedly increased by acute exercise and declined to baseline during post-exercise recovery. Dnm1L expression was strongly associated with transcripts known to regulate mitochondrial metabolism and adaptations to exercise. Exercise increased the expression of DNM1L in skeletal muscle of healthy control and DG subjects, despite a 15% ↓(P = 0.01) in muscle DNM1L expression in DG at baseline. To interrogate the role of Dnm1L further, we exercise trained male mDrp1+/- mice and found that Drp1 deficiency reduced muscle endurance and running performance, and altered muscle adaptations in response to exercise training. CONCLUSION: Our findings highlight the importance of mitochondrial dynamics, specifically Drp1 signaling, in the regulation of exercise performance and adaptations to endurance exercise training.