Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human.

Brown fat generates heat via the mitochondrial uncoupling protein UCP1, defending against hypothermia and obesity. Recent data suggest that there are two distinct types of brown fat: classical brown fat derived from a myf-5 cellular lineage and UCP1-positive cells that emerge in white fat from a non-myf-5 lineage. Here, we report the isolation of "beige" cells from murine white fat depots. Beige cells resemble white fat cells in having extremely low basal expression of UCP1, but, like classical brown fat, they respond to cyclic AMP stimulation with high UCP1 expression and respiration rates. Beige cells have a gene expression pattern distinct from either white or brown fat and are preferentially sensitive to the polypeptide hormone irisin. Finally, we provide evidence that previously identified brown fat deposits in adult humans are composed of beige adipocytes. These data provide a foundation for studying this mammalian cell type with therapeutic potential. PAPERCLIP:

Advances and challenges in measuring hepatic glucose uptake with FDG PET: implications for diabetes research.

The liver plays a crucial role in the control of glucose homeostasis and is therefore of great interest in the investigation of the development of type 2 diabetes. Hepatic glucose uptake (HGU) can be measured through positron emission tomography (PET) imaging with the tracer [18F]-2-fluoro-2-deoxy-D-glucose (FDG). HGU is dependent on many variables (e.g. plasma glucose, insulin and glucagon concentrations), and the metabolic state for HGU assessment should be chosen with care and coherence with the study question. In addition, as HGU is influenced by many factors, protocols and measurement conditions need to be standardised for reproducible results. This review provides insights into the protocols that are available for the measurement of HGU by FDG PET and discusses the current state of knowledge of HGU and its impairment in type 2 diabetes. Overall, a scanning modality that allows for the measurement of detailed kinetic information and influx rates (dynamic imaging) may be preferable to static imaging. The combination of FDG PET and insulin stimulation is crucial to measure tissue-specific insulin sensitivity. While the hyperinsulinaemic-euglycaemic clamp allows for standardised measurements under controlled blood glucose levels, some research questions might require a more physiological approach, such as oral glucose loading, with both advantages and complexities relating to fluctuations in blood glucose and insulin levels. The available approaches to address HGU hold great potential but await more systematic exploitation to improve our understanding of the mechanisms underlying metabolic diseases. Current findings from the investigation of HGU by FDG PET highlight the complex interplay between insulin resistance, hepatic glucose metabolism, NEFA levels and intrahepatic lipid accumulation in type 2 diabetes and obesity. Further research is needed to fully understand the underlying mechanisms and potential therapeutic targets for improving HGU in these conditions.

Timing of physical activity in relation to liver fat content and insulin resistance.

AIMS/HYPOTHESIS: We hypothesised that the insulin-sensitising effect of physical activity depends on the timing of the activity. Here, we examined cross-sectional associations of breaks in sedentary time and timing of physical activity with liver fat content and insulin resistance in a Dutch cohort. METHODS: In 775 participants of the Netherlands Epidemiology of Obesity (NEO) study, we assessed sedentary time, breaks in sedentary time and different intensities of physical activity using activity sensors, and liver fat content by magnetic resonance spectroscopy (n=256). Participants were categorised as being most active in the morning (06:00-12:00 hours), afternoon (12:00-18:00 hours) or evening (18:00-00:00 hours) or as engaging in moderate-to-vigorous-physical activity (MVPA) evenly distributed throughout the day. Most active in a certain time block was defined as spending the majority (%) of total daily MVPA in that block. We examined associations between sedentary time, breaks and timing of MVPA with liver fat content and HOMA-IR using linear regression analyses, adjusted for demographic and lifestyle factors including total body fat. Associations of timing of MVPA were additionally adjusted for total MVPA. RESULTS: The participants (42% men) had a mean (SD) age of 56 (4) years and a mean (SD) BMI of 26.2 (4.1) kg/m2. Total sedentary time was not associated with liver fat content or insulin resistance, whereas the amount of breaks in sedentary time was associated with higher liver fat content. Total MVPA (-5%/h [95% CI -10%/h, 0%/h]) and timing of MVPA were associated with reduced insulin resistance but not with liver fat content. Compared with participants who had an even distribution of MVPA throughout the day, insulin resistance was similar (-3% [95% CI -25%, 16%]) in those most active in morning, whereas it was reduced in participants who were most active in the afternoon (-18% [95% CI -33%, -2%]) or evening (-25% [95% CI -49%, -4%]). CONCLUSIONS/INTERPRETATION: The number of daily breaks in sedentary time was not associated with lower liver fat content or reduced insulin resistance. Moderate-to-vigorous activity in the afternoon or evening was associated with a reduction of up to 25% in insulin resistance. Further studies should assess whether timing of physical activity is also important for the occurrence of type 2 diabetes.

Exercise training modifies skeletal muscle clock gene expression but not 24-hour rhythmicity in substrate metabolism of men with insulin resistance.

Twenty-four hour rhythmicity in whole-body substrate metabolism, skeletal muscle clock gene expression and mitochondrial respiration is compromised upon insulin resistance. With exercise training known to ameliorate insulin resistance, our objective was to test if exercise training can reinforce diurnal variation in whole-body and skeletal muscle metabolism in men with insulin resistance. In a single-arm longitudinal design, 10 overweight and obese men with insulin resistance performed 12 weeks of high-intensity interval training recurrently in the afternoon (between 14.00 and 18.00 h) and were tested pre- and post-exercise training, while staying in a metabolic research unit for 2 days under free-living conditions with regular meals. On the second days, indirect calorimetry was performed at 08.00, 13.00, 18.00, 23.00 and 04.00 h, muscle biopsies were taken from the vastus lateralis at 08.30, 13.30 and 23.30 h, and blood was drawn at least bi-hourly over 24 h. Participants did not lose body weight over 12 weeks, but improved body composition and exercise capacity. Exercise training resulted in reduced 24-h plasma glucose levels, but did not modify free fatty acid and triacylglycerol levels. Diurnal variation of muscle clock gene expression was modified by exercise training with period genes showing an interaction (time × exercise) effect and reduced mRNA levels at 13.00 h. Exercise training increased mitochondrial respiration without inducing diurnal variation. Twenty-four-hour substrate metabolism and energy expenditure remained unchanged. Future studies should investigate alternative exercise strategies or types of interventions (e.g. diet or drugs aiming at improving insulin sensitivity) for their capacity to reinforce diurnal variation in substrate metabolism and mitochondrial respiration. KEY POINTS: Insulin resistance is associated with blunted 24-h flexibility in whole-body substrate metabolism and skeletal muscle mitochondrial respiration, and disruptions in the skeletal muscle molecular circadian clock. We hypothesized that exercise training modifies 24-h rhythmicity in whole-body substrate metabolism and diurnal variation in skeletal muscle molecular clock and mitochondrial respiration in men with insulin resistance. We found that metabolic inflexibility over 24 h persisted after exercise training, whereas mitochondrial respiration increased independent of time of day. Gene expression of Per1-3 and Rorα in skeletal muscle changed particularly close to the time of day at which exercise training was performed. These results provide the rationale to further investigate the differential metabolic impact of differently timed exercise to treat metabolic defects of insulin resistance that manifest at a particular time of day.

Three weeks of time-restricted eating improves glucose homeostasis in adults with type 2 diabetes but does not improve insulin sensitivity: a randomised crossover trial.

AIMS/HYPOTHESIS: Time-restricted eating (TRE) is suggested to improve metabolic health by limiting food intake to a defined time window, thereby prolonging the overnight fast. This prolonged fast is expected to lead to a more pronounced depletion of hepatic glycogen stores overnight and might improve insulin sensitivity due to an increased need to replenish nutrient storage. Previous studies showed beneficial metabolic effects of 6-8 h TRE regimens in healthy, overweight adults under controlled conditions. However, the effects of TRE on glucose homeostasis in individuals with type 2 diabetes are unclear. Here, we extensively investigated the effects of TRE on hepatic glycogen levels and insulin sensitivity in individuals with type 2 diabetes. METHODS: Fourteen adults with type 2 diabetes (BMI 30.5±4.2 kg/m2, HbA1c 46.1±7.2 mmol/mol [6.4±0.7%]) participated in a 3 week TRE (daily food intake within 10 h) vs control (spreading food intake over ≥14 h) regimen in a randomised, crossover trial design. The study was performed at Maastricht University, the Netherlands. Eligibility criteria included diagnosis of type 2 diabetes, intermediate chronotype and absence of medical conditions that could interfere with the study execution and/or outcome. Randomisation was performed by a study-independent investigator, ensuring that an equal amount of participants started with TRE and CON. Due to the nature of the study, neither volunteers nor investigators were blinded to the study interventions. The quality of the data was checked without knowledge on intervention allocation. Hepatic glycogen levels were assessed with 13C-MRS and insulin sensitivity was assessed using a hyperinsulinaemic-euglycaemic two-step clamp. Furthermore, glucose homeostasis was assessed with 24 h continuous glucose monitoring devices. Secondary outcomes included 24 h energy expenditure and substrate oxidation, hepatic lipid content and skeletal muscle mitochondrial capacity. RESULTS: Results are depicted as mean ± SEM. Hepatic glycogen content was similar between TRE and control condition (0.15±0.01 vs 0.15±0.01 AU, p=0.88). M value was not significantly affected by TRE (19.6±1.8 vs 17.7±1.8 µmol kg-1 min-1 in TRE vs control, respectively, p=0.10). Hepatic and peripheral insulin sensitivity also remained unaffected by TRE (p=0.67 and p=0.25, respectively). Yet, insulin-induced non-oxidative glucose disposal was increased with TRE (non-oxidative glucose disposal 4.3±1.1 vs 1.5±1.7 µmol kg-1 min-1, p=0.04). TRE increased the time spent in the normoglycaemic range (15.1±0.8 vs 12.2±1.1 h per day, p=0.01), and decreased fasting glucose (7.6±0.4 vs 8.6±0.4 mmol/l, p=0.03) and 24 h glucose levels (6.8±0.2 vs 7.6±0.3 mmol/l, p<0.01). Energy expenditure over 24 h was unaffected; nevertheless, TRE decreased 24 h glucose oxidation (260.2±7.6 vs 277.8±10.7 g/day, p=0.04). No adverse events were reported that were related to the interventions. CONCLUSIONS/INTERPRETATION: We show that a 10 h TRE regimen is a feasible, safe and effective means to improve 24 h glucose homeostasis in free-living adults with type 2 diabetes. However, these changes were not accompanied by changes in insulin sensitivity or hepatic glycogen. TRIAL REGISTRATION: ClinicalTrials.gov NCT03992248 FUNDING: ZonMW, 459001013.

The influence of bright and dim light on substrate metabolism, energy expenditure and thermoregulation in insulin-resistant individuals depends on time of day.

AIMS/HYPOTHESIS: In our modern society, artificial light is available around the clock and most people expose themselves to electrical light and light-emissive screens during the dark period of the natural light/dark cycle. Such suboptimal lighting conditions have been associated with adverse metabolic effects, and redesigning indoor lighting conditions to mimic the natural light/dark cycle more closely holds promise to improve metabolic health. Our objective was to compare metabolic responses to lighting conditions that resemble the natural light/dark cycle in contrast to suboptimal lighting in individuals at risk of developing metabolic diseases. METHODS: Therefore, we here performed a non-blinded, randomised, controlled, crossover trial in which overweight insulin-resistant volunteers (n = 14) were exposed to two 40 h laboratory sessions with different 24 h lighting protocols while staying in a metabolic chamber under real-life conditions. In the Bright day-Dim evening condition, volunteers were exposed to electric bright light (~1250 lx) during the daytime (08:00-18:00 h) and to dim light (~5 lx) during the evening (18:00-23:00 h). Vice versa, in the Dim day-Bright evening condition, volunteers were exposed to dim light during the daytime and bright light during the evening. Randomisation and allocation to light conditions were carried out by sequential numbering. During both lighting protocols, we performed 24 h indirect calorimetry, and continuous core body and skin temperature measurements, and took frequent blood samples. The primary outcome was plasma glucose focusing on the pre- and postprandial periods of the intervention. RESULTS: Spending the day in bright light resulted in a greater increase in postprandial triacylglycerol levels following breakfast, but lower glucose levels preceding the dinner meal at 18:00 h, compared with dim light (5.0 ± 0.2 vs 5.2 ± 0.2 mmol/l, n = 13, p=0.02). Dim day-Bright evening reduced the increase in postprandial glucose after dinner compared with Bright day-Dim evening (incremental AUC: 307 ± 55 vs 394 ± 66 mmol/l × min, n = 13, p=0.009). After the Bright day-Dim evening condition the sleeping metabolic rate was identical compared with the baseline night, whereas it dropped after Dim day-Bright evening. Melatonin secretion in the evening was strongly suppressed for Dim day-Bright evening but not for Bright day-Dim evening. Distal skin temperature for Bright day-Dim evening was lower at 18:00 h (28.8 ± 0.3°C vs 29.9 ± 0.4°C, n = 13, p=0.039) and higher at 23:00 h compared with Dim day-Bright evening (30.1 ± 0.3°C vs 28.8 ± 0.3°C, n = 13, p=0.006). Fasting and postprandial plasma insulin levels and the respiratory exchange ratio were not different between the two lighting protocols at any time. CONCLUSIONS/INTERPRETATION: Together, these findings suggest that the indoor light environment modulates postprandial substrate handling, energy expenditure and thermoregulation of insulin-resistant volunteers in a time-of-day-dependent manner. TRIAL REGISTRATION: ClinicalTrials.gov NCT03829982. FUNDING: We acknowledge the financial support from the Netherlands Cardiovascular Research Initiative: an initiative with support from the Dutch Heart Foundation (CVON2014-02 ENERGISE).

Novel insights in intestinal and hepatic fructose metabolism: from mice to men.

PURPOSE OF REVIEW: The rise in fructose consumption in parallel with the current epidemic of obesity and related cardiometabolic disease requires a better understanding of the pathophysiological pathways that are involved. RECENT FINDINGS: Animal studies have shown that fructose has various effects on the intestines that subsequently affect intrahepatic lipid accumulation and inflammation. Fructose adversely affects the gut microbiome - as a producer of endotoxins and intermediates of de novo lipogenesis - and intestinal barrier function. Furthermore, intestinal fructose metabolism shields fructose away from the liver. Finally, fructose 1-phosphate (F1-P) serves as a signal molecule that promotes intestinal cell survival and, consequently, intestinal absorption capacity. Intervention and epidemiological studies have convincingly shown that fructose, particularly derived from sugar-sweetened beverages, stimulates de novo lipogenesis and intrahepatic lipid accumulation in humans. Of interest, individuals with aldolase B deficiency, who accumulate F1-P, are characterized by a greater intrahepatic lipid content. First phase II clinical trials have recently shown that reduction of F1-P, by inhibition of ketohexokinase, reduces intrahepatic lipid content. SUMMARY: Experimental evidence supports current measures to reduce fructose intake, for example by the implementation of a tax on sugar-sweetened beverages, and pharmacological inhibition of fructose metabolism to reduce the global burden of cardiometabolic disease.

Circadian misalignment disturbs the skeletal muscle lipidome in healthy young men.

Circadian misalignment, as seen in shift work, is associated with an increased risk to develop type 2 diabetes. In an experimental setting, we recently showed that a rapid day-night shift for 3 consecutive nights leads to misalignment of the core molecular clock, induction of the PPAR pathway, and insulin resistance in skeletal muscle of young, healthy men. Here, we investigated if circadian misalignment affects the skeletal muscle lipidome and intramyocellular lipid droplet characteristics, explaining the misalignment-induced insulin resistance. Fourteen healthy men underwent one aligned and one circadian misalignment period, both consisting of ~3.5 days. In the misaligned condition, day and night were rapidly shifted by 12 hours leading to opposite eating, sleep, and activity times compared with the aligned condition. For each condition, two muscle biopsies were taken from the m. vastus lateralis in the morning and evening and subjected to semi-targeted lipidomics and confocal microscopy analysis. We found that only 2% of detected lipids were different between morning and evening in the aligned condition, whereas 12% displayed a morning-evening difference upon misalignment. Triacylglycerols, in particular species of a carbon length ≥55, were the most abundant lipid species changed upon misalignment. Cardiolipins were decreased upon misalignment, whereas phosphatidylcholines consistently followed the same morning-evening pattern, suggesting regulation by the circadian clock. Cholesteryl esters adjusted to the shifted behavior. Lipid droplet characteristics remained unaltered upon misalignment. Together, these findings show that simulated shift work disturbs the skeletal muscle lipidome, which may contribute to misalignment-induced insulin resistance.

Liver fat storage pathways: methodologies and dietary effects.

PURPOSE OF REVIEW: Nonalcoholic fatty liver is the result of an imbalance between lipid storage [from meal, de novo lipogenesis (DNL) and fatty acid (FA) uptake] and disposal (oxidation and VLDL output). Knowledge on the contribution of each of these pathways to liver fat content in humans is essential to develop tailored strategies to prevent and treat nonalcoholic fatty liver. Here, we review the techniques available to study the different storage pathways and review dietary modulation of these pathways. RECENT FINDINGS: The type of carbohydrate and fat could be of importance in modulating DNL, as complex carbohydrates and omega-3 FAs have been shown to reduce DNL. No effects were found on the other pathways, however studies investigating this are scarce. SUMMARY: Techniques used to assess storage pathways are predominantly stable isotope techniques, which require specific expertise and are costly. Validated biomarkers are often lacking. These methodological limitations also translate into a limited number of studies investigating to what extent storage pathways can be modulated by diet. Further research is needed to elucidate in more detail the impact that fat and carbohydrate type can have on liver fat storage pathways and content.

Human skeletal muscle mitochondrial dynamics in relation to oxidative capacity and insulin sensitivity.

AIMS/HYPOTHESIS: Mitochondria operate in networks, adapting to external stresses and changes in cellular metabolic demand and are subject to various quality control mechanisms. On the basis of these traits, we here hypothesise that the regulation of mitochondrial networks in skeletal muscle is hampered in humans with compromised oxidative capacity and insulin sensitivity. METHODS: In a cross-sectional design, we compared four groups of participants (selected from previous studies) ranging in aerobic capacity and insulin sensitivity, i.e. participants with type 2 diabetes (n = 11), obese participants without diabetes (n = 12), lean individuals (n = 10) and endurance-trained athletes (n = 12); basal, overnight fasted muscle biopsies were newly analysed for the current study and we compared the levels of essential mitochondrial dynamics and quality control regulatory proteins in skeletal muscle tissue. RESULTS: Type 2 diabetes patients and obese participants were older than lean participants and athletes (58.6 ± 4.0 and 56.7 ± 7.2 vs 21.8 ± 2.5 and 25.1 ± 4.3 years, p < 0.001, respectively) and displayed a higher BMI (32.4 ± 3.7 and 31.0 ± 3.7 vs 22.1 ± 1.8 and 21.0 ± 1.5 kg/m2, p < 0.001, respectively) than lean individuals and endurance-trained athletes. Fission protein 1 (FIS1) and optic atrophy protein 1 (OPA1) protein content was highest in muscle from athletes and lowest in participants with type 2 diabetes and obesity, respectively (FIS1: 1.86 ± 0.79 vs 0.79 ± 0.51 AU, p = 0.002; and OPA1: 1.55 ± 0.64 vs 0.76 ± 0.52 AU, p = 0.014), which coincided with mitochondrial network fragmentation in individuals with type 2 diabetes, as assessed by confocal microscopy in a subset of type 2 diabetes patients vs endurance-trained athletes (n = 6). Furthermore, lean individuals and athletes displayed a mitonuclear protein balance that was different from obese participants and those with type 2 diabetes. Mitonuclear protein balance also associated with heat shock protein 60 (HSP60) protein levels, which were higher in athletes when compared with participants with obesity (p = 0.048) and type 2 diabetes (p = 0.002), indicative for activation of the mitochondrial unfolded protein response. Finally, OPA1, FIS1 and HSP60 correlated positively with aerobic capacity (r = 0.48, p = 0.0001; r = 0.55, p < 0.001 and r = 0.61, p < 0.0001, respectively) and insulin sensitivity (r = 0.40, p = 0.008; r = 0.44, p = 0.003 and r = 0.48, p = 0.001, respectively). CONCLUSIONS/INTERPRETATION: Collectively, our data suggest that mitochondrial dynamics and quality control in skeletal muscle are linked to oxidative capacity in humans, which may play a role in the maintenance of muscle insulin sensitivity. CLINICAL TRIAL REGISTRY: numbers NCT00943059, NCT01298375 and NL1888 Graphical abstract.

Sitting less elicits metabolic responses similar to exercise and enhances insulin sensitivity in postmenopausal women.

AIMS/HYPOTHESIS: In our current society sedentary behaviour predominates in most people and is associated with the risk of developing type 2 diabetes. It has been suggested that replacing sitting time by standing and walking could be beneficial for individuals with type 2 diabetes but the underlying mechanisms are unknown and direct comparisons with exercise are lacking. Our objective was to directly compare metabolic responses of either sitting less or exercising, relative to being sedentary. METHODS: We performed a randomised, crossover intervention study in 12 overweight women who performed three well-controlled 4 day activity regimens: (1) sitting regimen (sitting 14 h/day); (2) exercise regimen (sitting 13 h/day, exercise 1 h/day); and (3) sitting less regimen (sitting 9 h/day, standing 4 h/day and walking 3 h/day). The primary outcome was insulin sensitivity measured by a two-step hyperinsulinaemic-euglycaemic clamp. We additionally performed metabolomics on muscle biopsies taken before the clamp to identify changes at the molecular level. RESULTS: Replacing sitting time by standing and walking over 4 days resulted in improved peripheral insulin sensitivity, comparable with the improvement achieved by moderate-to-vigorous exercise. Specifically, we report a significant improvement in peripheral insulin sensitivity in the sitting less (~13%) and the exercise regimen (~20%), compared with the sitting regimen. Furthermore, sitting less shifted the underlying muscle metabolome towards that seen with moderate-to-vigorous exercise, compared with the sitting regimen. CONCLUSIONS/INTERPRETATIONS: Replacing sitting time by standing and walking is an attractive alternative to moderate-to-vigorous exercise for improving metabolic health. TRIAL REGISTRATION: ClinicalTrials.gov NCT03912922.

Postexercise changes in myocellular lipid droplet characteristics of young lean individuals are affected by circulatory nonesterified fatty acids.

Intramyocellular lipid (IMCL) content is an energy source during acute exercise. Nonesterified fatty acid (NEFA) levels can compete with IMCL utilization during exercise. IMCL content is stored as lipid droplets (LDs) that vary in size, number, subcellular distribution, and in coating with LD protein PLIN5. Little is known about how these factors are affected during exercise and recovery. Here, we aimed to investigate the effects of acute exercise with and without elevated NEFA levels on intramyocellular LD size and number, intracellular distribution and PLIN5 coating, using high-resolution confocal microscopy. In a crossover study, 9 healthy lean young men performed a 2-h moderate intensity cycling protocol in the fasted (high NEFA levels) and glucose-fed state (low NEFA levels). IMCL and LD parameters were measured at baseline, directly after exercise and 4 h postexercise. We found that total IMCL content was not changed directly after exercise (irrespectively of condition), but IMCL increased 4 h postexercise in the fasting condition, which was due to an increased number of LDs rather than changes in size. The effects were predominantly detected in type I muscle fibers and in LDs coated with PLIN5. Interestingly, subsarcolemmal, but not intermyofibrillar IMCL content, was decreased directly after exercise in the fasting condition and was replenished during the 4 h recovery period. In conclusion, acute exercise affects IMCL storage during exercise and recovery, particularly in type I muscle fibers, in the subsarcolemmal region and in the presence of PLIN5. Moreover, the effects of exercise on IMCL content are affected by plasma NEFA levels.NEW & NOTEWORTHY Skeletal muscle stores lipids in lipid droplets (LDs) that can vary in size, number, and location and are a source of energy during exercise. Specifically, subsarcolemmal LDs were used during exercise when fasted. Exercising in the fasted state leads to postrecovery elevation in IMCL levels due to an increase in LD number in type I muscle fibers, in subsarcolemmal region and decorated with PLIN5. These effects are blunted by glucose ingestion during exercise and recovery.

The importance of 24-h metabolism in obesity-related metabolic disorders: opportunities for timed interventions.

Various metabolic processes in the body oscillate throughout the natural day, driven by a biological clock. Circadian rhythms are also influenced by time cues from the environment (light exposure) and behaviour (eating and exercise). Recent evidence from diurnal- and circadian-rhythm studies indicates rhythmicity in various circulating metabolites, insulin secretion and -sensitivity and energy expenditure in metabolically healthy adults. These rhythms have been shown to be disturbed in adults with obesity-related metabolic disturbances. Moreover, eating and being (in)active at a time that the body is not prepared for it, as in night-shift work, is related to poor metabolic outcomes. These findings indicate the relevance of 24-h metabolism in obesity-related metabolic alterations and have also led to novel strategies, such as timing of food intake and exercise, to reinforce the circadian rhythm and thereby improving metabolic health. This review aims to deepen the understanding of the influence of the circadian system on metabolic processes and obesity-related metabolic disturbances and to discuss novel time-based strategies that may be helpful in combating metabolic disease.

The effect of physical activity level and exercise training on the association between plasma branched-chain amino acids and intrahepatic lipid content in participants with obesity.

AIMS: To evaluate whether the association between plasma branched-chain amino acids (BCAA) and intrahepatic lipid (IHL) was affected by physical activity level. Furthermore, to investigate if a conventional exercise training program, a subcategory of physical activity, could lower plasma BCAA along with alterations in IHL content in patients with type 2 diabetes (T2DM) and people with nonalcoholic fatty liver (NAFL). METHODS: To investigate the effect of physical activity on the association between plasma BCAA and IHL content, linear regression analyses were performed in 1983 individuals from the Netherlands Epidemiology of Obesity (NEO) stratified by physical activity frequency. Furthermore, the effect of a 12-week supervised combined aerobic resistance-exercise program on plasma BCAA, insulin sensitivity (hyperinsulinemic-euglycemic clamp), and IHL (proton-magnetic resonance spectroscopy (1H-MRS)) was investigated in seven patients with T2DM, seven individuals with NAFL and seven BMI-matched control participants (CON). RESULTS: We observed positive associations between plasma valine, isoleucine and leucine level, and IHL content (1.29 (95% CI: 1.21, 1.38), 1.52 (95% CI: 1.43, 1.61), and 1.54 (95% CI: 1.44, 1.64) times IHL, respectively, per standard deviation of plasma amino acid level). Similar associations were observed in less active versus more active individuals. Exercise training did not change plasma BCAA levels among groups, but reduced IHL content in NAFL (from 11.6 ± 3.0% pre-exercise to 8.1 ± 2.0% post exercise, p < 0.05) and CON (from 2.4 ± 0.6% pre-exercise to 1.6 ± 1.4% post exercise, p < 0.05), and improved peripheral insulin sensitivity in NAFL as well by ~23% (p < 0.05). CONCLUSIONS: The association between plasma BCAA levels and IHL is not affected by physical activity level. Exercise training reduced IHL without affecting plasma BCAA levels in individuals with NAFL and CON. We conclude that exercise training-induced reduction in IHL content is not related to changes in plasma BCAA levels. TRIAL REGISTRATION: Trial registry number: NCT01317576.

Relationship between de novo lipogenesis and serum sex hormone binding globulin in humans.

OBJECTIVE: Obesity and liver fat are associated with decreased levels of serum sex hormone binding globulin (SHBG). Laboratory studies suggest that hepatic de novo lipogenesis (DNL) is involved in the downregulation of SHBG synthesis. The aim of the present study was to address the role of DNL on serum SHBG in humans. DESIGN: A cross-sectional study examining the association between DNL, measured by stable isotopes, and serum SHBG, stratified by sex. PARTICIPANTS: Healthy men (n = 34) and women (n = 21) were combined from two cross-sectional studies. Forty-two per cent of participants had hepatic steatosis, and the majority were overweight (62%) or obese (27%). RESULTS: DNL was inversely associated with SHBG in women (ß: -0.015, 95% CI: -0.030; 0.000), but not in men (ß: 0.007, 95% CI: -0.005; 0.019) (p for interaction = .068). Adjustment for study population, age and body mass index did not materially change these results, although statistical significance was lost after adjustment for serum insulin. CONCLUSIONS: An inverse association between DNL and SHBG may explain the decreased SHBG levels that are observed in obesity, at least in women.

Circadian misalignment induces fatty acid metabolism gene profiles and compromises insulin sensitivity in human skeletal muscle.

Circadian misalignment, such as in shift work, has been associated with obesity and type 2 diabetes. However, direct effects of circadian misalignment on skeletal muscle insulin sensitivity and the muscle molecular circadian clock have never been studied in humans. Here, we investigated insulin sensitivity and muscle metabolism in 14 healthy young lean men [age 22.4 ± 2.8 years; body mass index (BMI) 22.3 ± 2.1 kg/m2 (mean ± SD)] after a 3-d control protocol and a 3.5-d misalignment protocol induced by a 12-h rapid shift of the behavioral cycle. We show that short-term circadian misalignment results in a significant decrease in muscle insulin sensitivity due to a reduced skeletal muscle nonoxidative glucose disposal (rate of disappearance: 23.7 ± 2.4 vs. 18.4 ± 1.4 mg/kg per minute; control vs. misalignment; P = 0.024). Fasting glucose and free fatty acid levels as well as sleeping metabolic rate were higher during circadian misalignment. Molecular analysis of skeletal muscle biopsies revealed that the molecular circadian clock was not aligned to the inverted behavioral cycle, and transcriptome analysis revealed the human PPAR pathway as a key player in the disturbed energy metabolism upon circadian misalignment. Our findings may provide a mechanism underlying the increased risk of type 2 diabetes among shift workers.

Exercising your fat (metabolism) into shape: a muscle-centred view.

Fatty acids are an important energy source during exercise. Training status and substrate availability are determinants of the relative and absolute contribution of fatty acids and glucose to total energy expenditure. Endurance-trained athletes have a high oxidative capacity, while, in insulin-resistant individuals, fat oxidation is compromised. Fatty acids that are oxidised during exercise originate from the circulation (white adipose tissue lipolysis), as well as from lipolysis of intramyocellular lipid droplets. Moreover, hepatic fat may contribute to fat oxidation during exercise. Nowadays, it is clear that myocellular lipid droplets are dynamic organelles and that number, size, subcellular distribution, lipid droplet coat proteins and mitochondrial tethering of lipid droplets are determinants of fat oxidation during exercise. This review summarises recent insights into exercise-mediated changes in lipid metabolism and insulin sensitivity in relation to lipid droplet characteristics in human liver and muscle. Graphical abstract.

One-leg inactivity induces a reduction in mitochondrial oxidative capacity, intramyocellular lipid accumulation and reduced insulin signalling upon lipid infusion: a human study with unilateral limb suspension.

AIMS/HYPOTHESIS: Physical inactivity, low mitochondrial function, increased intramyocellular lipid (IMCL) deposition and reduced insulin sensitivity are common denominators of chronic metabolic disorders, like obesity and type 2 diabetes. Yet, whether low mitochondrial function predisposes to insulin resistance in humans is still unknown. METHODS: Here we investigated, in an intervention study, whether muscle with low mitochondrial oxidative capacity, induced by one-legged physical inactivity, would feature stronger signs of lipid-induced insulin resistance. To this end, ten male participants (age 22.4 ± 4.2 years, BMI 21.3 ± 2.0 kg/m2) underwent a 12 day unilateral lower-limb suspension with the contralateral leg serving as an active internal control. RESULTS: In vivo, mitochondrial oxidative capacity, assessed by phosphocreatine (PCr)-recovery half-time, was lower in the inactive vs active leg. Ex vivo, palmitate oxidation to 14CO2 was lower in the suspended leg vs the active leg; however, this did not result in significantly higher [14C]palmitate incorporation into triacylglycerol. The reduced mitochondrial function in the suspended leg was, however, paralleled by augmented IMCL content in both musculus tibialis anterior and musculus vastus lateralis, and by increased membrane bound protein kinase C (PKC) θ. Finally, upon lipid infusion, insulin signalling was lower in the suspended vs active leg. CONCLUSIONS/INTERPRETATION: Together, these results demonstrate, in a unique human in vivo model, that a low mitochondrial oxidative capacity due to physical inactivity directly impacts IMCL accumulation and PKCθ translocation, resulting in impaired insulin signalling upon lipid infusion. This demonstrates the importance of mitochondrial oxidative capacity and muscle fat accumulation in the development of insulin resistance in humans. TRIAL REGISTRATION: ClinicalTrial.gov NCT01576250. FUNDING: PS was supported by a 'VICI' Research Grant for innovative research from the Netherlands Organization for Scientific Research (Grant 918.96.618).

A Whole-Diet Approach Affects Not Only Fasting but Also Postprandial Cardiometabolic Risk Markers in Overweight and Obese Adults: A Randomized Controlled Trial.

BACKGROUND: Current dietary recommendations for cardiovascular disease (CVD) prevention focus more on dietary patterns than on single nutrients. However, randomized controlled trials using whole-diet approaches to study effects on both fasting and postprandial CVD risk markers are limited. OBJECTIVE: This randomized parallel trial compared the effects of a healthy diet (HD) with those of a typical Western diet (WD) on fasting and postprandial CVD risk markers in overweight and obese adults. METHODS: After a 2-wk run-in period, 40 men and women (50-70 y; BMI: 25-35 kg/m2) consumed the HD (high in fruit and vegetables, pulses, fibers, nuts, fatty fish, polyunsaturated fatty acids; low in salt and high-glycemic carbohydrates; n = 19) or the WD (less fruit, vegetables, and fibers; no nuts and fatty fish; and more saturated fatty acids and simple carbohydrates; n = 21) for 6 wk. Fasting and postprandial cardiometabolic risk markers were assessed as secondary outcome parameters during a 5-h mixed-meal challenge, and a per protocol analysis was performed using 1-factor ANCOVA or linear mixed models. RESULTS: Differences in diet-induced changes are expressed relative to the HD group. Changes in fasting plasma total cholesterol (-0.57 ± 0.12 mmol/L, P < 0.001), LDL cholesterol (-0.41 ± 0.12 mmol/L, P < 0.01), apolipoprotein B100 (-0.09 ± 0.03 g/L, P < 0.01), and apolipoprotein A1 (-0.06 ± 0.03 g/L, P = 0.05) were significantly different between the diet groups. Changes in postprandial plasma triacylglycerol (diet × time, P < 0.001) and apolipoprotein B48 (P < 0.01) differed significantly between the groups with clear improvements on the HD, although fasting triacylglycerols (-0.24 ± 0.13 mmol/L, P = 0.06) and apolipoprotein B48 (1.04 ± 0.67 mg/L, P = 0.40) did not. Significant differences between the diets were also detected in fasting systolic (-6.9 ± 3.1 mmHg, P < 0.05) and 24-h systolic (-5.0 ± 1.7 mmHg, P < 0.01) and diastolic (-3.3 ± 1.1 mmHg, P < 0.01) blood pressure. CONCLUSION: A whole-diet approach targeted multiple fasting and postprandial CVD risk markers in overweight and obese adults. In fact, the postprandial measurements provided important additional information to estimate CVD risk. This trial is registered at clinicaltrials.gov as NCT02519127.

In vitro effects of sitosterol and sitostanol on mitochondrial respiration in human brown adipocytes, myotubes and hepatocytes.

PURPOSE: Lowering of LDL cholesterol levels by plant sterols and stanols is associated with decreased risk of cardiovascular disease in humans. Plant sterols and stanols also lower triacylglycerol (TG). However, it is not fully understood how reduction in TG is achieved and what the full potential of plant sterols and stanols is on whole-body metabolism. We here hypothesize that high levels of plant sterols and stanols stimulate whole-body energy expenditure, which can be attributed to changes in mitochondrial function of brown adipose tissue (BAT), skeletal muscle and liver. METHODS: Phytosterolemic mice were fed chow diets for 32 weeks to examine whole-body weight gain. In vitro, 24-h incubation were performed in adipocytes derived from human BAT, human myotubes or HepG2 human hepatocytes using sitosterol or sitostanol. Following mitochondrial function was assessed using seahorse bioanalyzer. RESULTS: Chow feeding in phytosterolemic mice resulted in diminished increase in body weight compared to control mice. In vitro, sitosterol or sitostanol did not change mitochondrial function in adipocytes derived from human BAT or in cultured human myotubes. Interestingly, maximal mitochondrial function in HepG2 human hepatocytes was decreased following sitosterol or sitostanol incubation, however, only when mitochondrial function was assessed in low glucose-containing medium. CONCLUSIONS: Beneficial in vivo effects of plant sterols and stanols on lipid and lipoprotein metabolism are well recognized. Our results indicate that alterations in human mitochondrial function are apparently not involved to explain these beneficial effects.