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Aims: To test if a New Zealand food-based Mediterranean diet (NZMedDiet) with behavioural intervention improves cardiometabolic health and wellbeing., Methods: A randomised controlled trial comparing 12 weeks of the NZMedDiet to usual diet in participants with increased cardiometabolic risk (metabolic syndrome severity score [MetSSS] > 0.35). The intervention group was provided with food and recipes to meet 75% of their energy requirements, supported by a behavioural intervention to improve adherence. The primary outcome measure was (MetSSS) after 12 weeks., Results: Two hundred individuals with mean (SD) age 49.9 (10.9) years of which 62% women were enrolled with their household/whānau. After 12 weeks, the mean (SD) MetSSS was 1.0 (0.7) in the control (n = 98) and 0.8 (0.5) in the intervention (n = 102) group; estimated difference (95% confidence interval [CI]) of -0.05 (-0.16 to 0.06), p = 0.35. The Mediterranean diet score (PyrMDS) was greater in the intervention group 1.6 (1.1-2.1), p < 0.001, consistent with a change to a more Mediterranean dietary pattern. Weight reduced in the NZMedDiet group compared with control (-1.9 kg [-2.0 to -0.34]), p = 0.006 and wellbeing, assessed by the SF-36 quality of life questionnaire, and improved across all domains. For example, the physical component summary score difference (95% CI) was 4.0 (2.4-5.7), p < 0.001, and the mental component summary score difference was 3.0 (0.7-5.2), p = 0.01., Conclusion: In participants with increased cardiometabolic risk, food provision with a Mediterranean dietary pattern and a behavioural intervention did not improve metabolic risk scores but was associated with reduced weight and improved quality of life., (© 2024 John Wiley & Sons Ltd.)

Aims/hypothesis: Metformin is an important first-line treatment for type 2 diabetes and acts by increasing the body's ability to dispose of glucose. Metformin's efficacy can be affected by genetic variants in the transporters that regulate its uptake into cells. The SLC22A3 gene (also known as EMT; EMTH; OCT3) codes for organic cation transporter 3 (OCT3), which is a broad-specificity cation transporter that also transports metformin. Most SLC22A3 variants reduce the rate of metformin transport but the rs8187715 variant (p.Thr44Met) is reported to increase uptake of metformin in vitro. However, the impact of this on in vivo metformin transport and efficacy is unknown. Very few carriers of this variant have been reported globally, but, notably, all were of Pacific Island descent. Therefore, this study aims to understand the prevalence of this variant in Polynesian peoples (Māori and Pacific peoples) and to understand its impact on metformin transport and efficacy in vivo., Methods: rs8187715 was genotyped in 310 individuals with Māori and Pacific ancestry recruited in Aotearoa New Zealand. To study this variant in a physiological context, an orthologous knockin mouse model with C57BL/6J background was used. Pharmacokinetic analysis compared uptake rate of metformin into tissues. Plasma growth/differentiation factor 15 (GDF-15) was also measured as a marker of metformin efficacy. Glucose and insulin tolerance was assessed after acute or sustained metformin treatment in knockin and wild-type control mice to examine the impact of the variant on metformin's glycaemic control., Results: The minor allele frequency of this variant in the Māori and Pacific participants was 15.4%. There was no association of the variant with common metabolic parameters including diabetes status, BMI, blood pressure, lipids, or blood glucose and HbA 1c . However, in the orthologous knockin mouse model, the rate of metformin uptake into the blood and tissues was increased. Acute metformin dosing increased insulin sensitivity in variant knockin mice but this effect was lost after longer-term metformin treatment. Metformin's effects on GDF-15 levels were also lost in variant knockin mice with longer-term metformin treatment., Conclusions/interpretation: These data provide evidence that the SLC22A3 rs8187715 variant accelerates metformin uptake rate in vivo. While this acutely improves insulin sensitivity, there was no increased effect of metformin with longer-term dosing. Thus, our finding of a high prevalence of this variant specifically in Māori and Pacific peoples identifies it as a potential population-specific pharmacogenetic marker with potential to guide metformin therapy in these peoples., (© 2024. The Author(s).)

Cardiac glycogen-autophagy ('glycophagy') is disturbed in cardiometabolic pathologies. The physiological role of cardiac glycophagy is unclear. Exercise induces transient cardiac glycogen accumulation. Thus, this study experimentally examined glycophagy involvement during recovery from an exhaustive exercise protocol. Peak myocardial glycogen accumulation in mice was evident at 2 h post-exercise, preceded by transient activation of glycogen synthase. At 4 and 16 h post-exercise, glycogen degradation was associated with decreased STBD1 (glycophagy tagging protein) and increased GABARAPL1 (Atg8 protein), suggesting that glycophagy activity was increased. These findings provide the first evidence that glycophagy is involved in cardiac glycogen physiologic homeostasis post-exercise., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Lipoprotein(a) (Lp(a)) is a low-density lipoprotein (LDL)-like particle in which the apolipoprotein B component is covalently linked to apolipoprotein(a) (apo(a)). Lp(a) is a well-established independent risk factor for cardiovascular diseases. Plasma Lp(a) concentrations vary enormously between individuals and ethnic groups. Several nucleotide polymorphisms in the SLC22A3 gene associate with Lp(a) concentration in people of different ethnicities. We investigated the association of a Polynesian-specific (Māori and Pacific peoples) SLC22A3 gene coding variant p.Thr44Met) with the plasma concentration of Lp(a) in a cohort of 302 healthy Polynesian males. An apo(a)-size independent assay assessed plasma Lp(a) concentrations; all other lipid and apolipoprotein concentrations were measured using standard laboratory techniques. Quantitative real-time polymerase chain reaction was used to determine apo(a) isoforms. The range of metabolic (HbA1c, blood pressure, and blood lipids) and blood lipid variables were similar between the non-carriers and carriers in age, ethnicity and BMI adjusted models. However, rs8187715 SLC22A3 variant was significantly associated with lower Lp(a) concentrations. Median Lp(a) concentration was 10.60 nmol/L (IQR: 5.40-41.00) in non-carrier group, and was 7.60 nmol/L (IQR: 5.50-12.10) in variant carrier group (P<0.05). Lp(a) concentration inversely correlated with apo(a) isoform size. After correction for apo(a) isoform size, metabolic parameters and ethnicity, the association between the SLC22A3 variant and plasma Lp(a) concentration remained. The present study is the first to identify the association of this gene variant and low plasma Lp(a) concentrations. This provides evidence for better guidance on ethnic specific cut-offs when defining 'elevated' and 'normal' plasma Lp(a) concentrations in clinical applications., (© 2024 The Author(s).)

There is some evidence for temperature-dependent stimulation of mitochondrial biogenesis; however, the role of elevated muscle temperature during exercise in mitochondrial adaptation to training has not been studied in humans in vivo. The purpose of this study was to determine the role of elevating muscle temperature during exercise in temperate conditions through the application of mild, local heat stress on mitochondrial adaptations to endurance training. Eight endurance-trained males undertook 3 weeks of supervised cycling training, during which mild (~ 40 °C) heat stress was applied locally to the upper-leg musculature of one leg during all training sessions (HEAT), with the contralateral leg serving as the non-heated, exercising control (CON). Vastus lateralis microbiopsies were obtained from both legs before and after the training period. Training-induced increases in complex I (fold-change, 1.24 ± 0.33 vs. 1.01 ± 0.49, P = 0.029) and II (fold-change, 1.24 ± 0.33 vs. 1.01 ± 0.49, P = 0.029) activities were significantly larger in HEAT than CON. No significant effects of training, or interactions between local heat stress application and training, were observed for complex I-V or HSP70 protein expressions. Our data provides partial evidence to support the hypothesis that elevating local muscle temperature during exercise augments training-induced adaptations to mitochondrial enzyme activity., (© 2024. The Author(s).)

Background: Cardiometabolic diseases are highly prevalent in Aotearoa New Zealand. Dietary intake is a modifiable risk factor for such diseases and certain dietary patterns, specifically the Mediterranean diet (MedDiet), are associated with improved metabolic health. This study aims to test whether an intervention including a Mediterranean dietary pattern incorporating high quality New Zealand foods (NZMedDiet pattern) and behavior change science can improve the metabolic health of participants and their household/whānau., Methods and Analysis: This is a multi-center, three-stage trial with two parallel group superiority randomized controlled trials (RCTs), and a longitudinal cohort study embedded within the trial design. The first RCT (RCT 1) is a comparison of the NZMedDiet pattern compared to usual diet for 12 weeks. The Behavior Change Wheel was used to select and implement strategies to support participant adherence to the NZMedDiet, such as web-based nutrition education on healthy shopping and cooking. The second (RCT 2) compares online social support to no online social support for 12 weeks, administered to participants immediately following RCT 1. The third stage is a longitudinal cohort study where all participants are followed from the beginning of their start of the active intervention for 12 months in total. The primary outcome measure for each stage is the metabolic syndrome severity score (MetSSS). The duration of enrolment is 12-15 months. The total recruitment target is 200 index participants and their household/whānau members who participate with them, and the primary analyses will be intention to treat on index participants., Discussion: The trial will test whether the NZMedDiet pattern and behavior change support improves the cardiometabolic health of people in Aotearoa New Zealand., Clinical Trial Registration: https://www.anzctr.org.au/Default.aspx, identifier ACTRN12622000906752 and https://www.isrctn.com/, identifier ISRCTN89011056 (Spirit 2)., Competing Interests: MF was employed by Edible Research Ltd. DC was employed by the New Zealand Institute for Plant & Food Research Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Lithander, Parry Strong, Braakhuis, Worthington, Foster, Rolleston, Davies, Mullaney, Ross, Conroy, Merry, Gearry, Weatherall and Krebs.)

Aim: To assess the feasibility of a family-based dietary intervention study using a meal kit home delivery service, in people at risk of cardio-metabolic disease., Methods: A 12-week dietary intervention feasibility study of adults (termed the index participants) at increased risk of metabolic and cardiovascular disease, enriched for Māori who are indigenous New Zealanders. The study sample also included the household/whānau members living with the index participant. All participants received a 12 week intervention using weekly home delivery of meal kits and groceries consistent with a Mediterranean dietary pattern. Outcomes were the metabolic syndrome severity score (MetSSS); feasibility and acceptability of the intervention; dietary intake; and other clinical and anthropometric measures., Results: There were 29 index participants recruited and in addition, 50 household/whānau members took part in the feasibility study. The mean (SD) household/whānau size was 3.45 (1.4) people, and the mean (SD) number of people in each household/whānau who participated in the study was 2.84 (1.2). The feasibility of intervention to households/whānau was proven in this context. The mean (SD) change in MetSSS was 0.03 (0.33), N = 27, P = 0.69 and there was a statistically significant decrease in body weight of 1.37 kg (95% CI 0.11 to 2.62), p = 0.034. The food deliveries were well received, the dinner kits more so than the grocery items., Conclusion: It is feasible to recruit individuals and households/whānau to a family-based dietary intervention. Use of a meal kit home delivery service to provide food which is consistent with the intervention dietary pattern was well received. This feasibility study identified improvements to be made such as nutrition behaviour change support, more variety in food provided, more recipes, and better matching of food quantity to family size., Trial Registration: ANZCTR-ACTRN12621000856819p registered 2.JUN.2021 https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=382021&isReview=true., (© 2023. The Author(s).)

3,6,7-trimethyllumazine (Lepteridine™) is a newly discovered natural pteridine derivative unique to Mānuka ( Leptospermum scoparium ) nectar and honey, with no previously reported biological activity. Pteridine derivative-based medicines, such as methotrexate, are used to treat auto-immune and inflammatory diseases, and Mānuka honey reportedly possesses anti-inflammatory properties and is used topically as a wound dressing. MMP-9 is a potential candidate protein target as it is upregulated in recalcitrant wounds and intestinal inflammation. Using gelatin zymography, 40 μg/mL Lepteridine TM inhibited the gelatinase activities of both pro- (22%, p < 0.0001) and activated (59%, p < 0.01) MMP-9 forms. By comparison, Lepteridine TM exerted modest (~10%) inhibition against a chromogenic peptide substrate and no effect against a fluorogenic peptide substrate. These findings suggest that Lepteridine TM may not interact within the catalytic domain of MMP-9 and exerts a negligible effect on the active site hydrolysis of small soluble peptide substrates. Instead, the findings implicate fibronectin II domain interactions by Lepteridine TM which impair gelatinase activity, possibly through perturbed tethering of MMP-9 to the gelatin matrix. Molecular modelling analyses were equivocal over interactions at the S1' pocket versus the fibronectin II domain, while molecular dynamic calculations indicated rapid exchange kinetics. No significant degradation of synthetic or natural Lepteridine TM in Mānuka honey occurred during simulated gastrointestinal digestion. MMP-9 regulates skin and gastrointestinal inflammatory responses and extracellular matrix remodelling. These results potentially implicate Lepteridine TM bioactivity in Mānuka honey's reported beneficial effects on wound healing via topical application and anti-inflammatory actions in gastrointestinal disorder models via oral consumption.

There is increasing evidence that adherence to a Mediterranean dietary pattern reduces the incidence of diet-related diseases. To date, the habitual dietary intake of New Zealand (NZ) adults has not been examined in relation to its alignment with a Mediterranean-style dietary pattern. This study aimed to define the habitual dietary patterns, nutrient intakes, and adherence to the Mediterranean Diet in a sample of 1012 NZ adults (86% female, mean age 48 ± 16 years) who had their diabetes risk defined by the Australian Type 2 Diabetes Risk Assessment Tool (AUSDRISK). Dietary intakes were collected using a validated semi-quantitative NZ food frequency questionnaire, and dietary patterns were identified using principal component analysis. Reported intakes from the FFQ were used in conjunction with the Mediterranean-Style Dietary Pattern Score (MSDPS) to determine adherence to a Mediterranean dietary pattern. Mixed linear models were used to analyze the association between dietary patterns and MSDPS with demographics, health factors, and nutrient intakes. Two distinct dietary patterns were identified: Discretionary (positive loadings on processed meat, meat/poultry, fast food, sweet drinks, and sugar, sweets, and baked good) and Guideline (positive loadings on vegetables, eggs/beans, and fruits). Adherence to dietary patterns and diet quality was associated with age and ethnicity. Dietary patterns were also associated with sex. Adherence to a Mediterranean dietary pattern defined by the MSDPS was low, indicating that a significant shift in food choices will be required if the Mediterranean Diet is to be adopted in the NZ population.

A central characteristic of insulin resistance is the impaired ability for insulin to stimulate glucose uptake into skeletal muscle. While insulin resistance can occur distal to the canonical insulin receptor-PI3k-Akt signaling pathway, the signaling intermediates involved in the dysfunction are yet to be fully elucidated. β-catenin is an emerging distal regulator of skeletal muscle and adipocyte insulin-stimulated GLUT4 trafficking. Here, we investigate its role in skeletal muscle insulin resistance. Short-term (5-week) high-fat diet (HFD) decreased skeletal muscle β-catenin protein expression 27% (p = 0.03), and perturbed insulin-stimulated β-catenin S552 phosphorylation 21% (p = 0.009) without affecting insulin-stimulated Akt phosphorylation relative to chow-fed controls. Under chow conditions, mice with muscle-specific β-catenin deletion had impaired insulin responsiveness, whereas under HFD, both mice exhibited similar levels of insulin resistance (interaction effect of genotype × diet p < 0.05). Treatment of L6-GLUT4-myc myocytes with palmitate lower β-catenin protein expression by 75% (p = 0.02), and attenuated insulin-stimulated β-catenin phosphorylation S552 and actin remodeling (interaction effect of insulin × palmitate p < 0.05). Finally, β-catenin S552 phosphorylation was 45% lower in muscle biopsies from men with type 2 diabetes while total β-catenin expression was unchanged. These findings suggest that β-catenin dysfunction is associated with the development of insulin resistance., (© 2023 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Diminished insulin and insulin-like growth factor-1 signaling extends the lifespan of invertebrates 1-4 ; however, whether it is a feasible longevity target in mammals is less clear 5-12 . Clinically utilized therapeutics that target this pathway, such as small-molecule inhibitors of phosphoinositide 3-kinase p110α (PI3Ki), provide a translatable approach to studying the impact of these pathways on aging. Here, we provide evidence that dietary supplementation with the PI3Ki alpelisib from middle age extends the median and maximal lifespan of mice, an effect that was more pronounced in females. While long-term PI3Ki treatment was well tolerated and led to greater strength and balance, negative impacts on common human aging markers, including reductions in bone mass and mild hyperglycemia, were also evident. These results suggest that while pharmacological suppression of insulin receptor (IR)/insulin-like growth factor receptor (IGFR) targets could represent a promising approach to delaying some aspects of aging, caution should be taken in translation to humans., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The number of sugar-sweetened beverages consumed per day has been associated with an increased risk of acute rheumatic fever, an autoimmune disease triggered by superficial Streptococcus pyogenes infection. To explore if there could be a biological basis for this association, we used a mouse model of S. pyogenes nasopharyngeal colonisation combined with a dietary intervention. We observed an increased bacterial load in the nasopharynx of mice receiving sucrose drinking water post-infection, suggesting that high sucrose intake promotes S. pyogenes growth and/or survival. This provides new insight into the potential biological basis behind the association seen in humans., (© 2022. The Author(s).)

Aim: To investigate the relationship of metabolic syndrome severity score (MetSSS) with glucose regulatory and cardiovascular disease (CVD) status in Aotearoa New Zealand., Methods: MetSSS and MetSSS component coefficients were calculated for participants from the cross-sectional Workforce Diabetes Study (WDS) (n = 5,806) and Diabetes, Heart and Health Survey (DHAH) (n = 4,010) and compared by ethnicity (European, Māori, Pacific and Asian), glucose regulatory status [impaired fasting glucose, impaired glucose tolerance and type 2 diabetes) and history of cardiovascular disease., Results: MetSSS positively associated with impaired glucose regulatory status and history of cardiovascular disease for all ethnic groups. Ethnicity significantly affected different coefficients of the MetSSS components, however all ethnicities had an approximately normal MetSSS distribution, with Māori and Pacific curves being right-shifted compared to European. While the MetSSS thresholds that capture 80% of participant with type 2 diabetes (T2D) were higher for Māori and Pacific, the difference in MetSSS between those participants with and without type 2 diabetes within an ethnicity group was similar across ethnicities., Conclusion: MetSSS may have utility as a tool to quantify an individual's cardiometabolic disease risk within the multi-ethnic population of Aotearoa New Zealand, however ethnic-specific categories for disease risk are likely to be required., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Objective: Model-based metabolic tests require accurate identification of subject-specific parameters from measured assays. Insulin assays are used to identify insulin kinetics parameters, such as general and first-pass hepatic clearances. This study assesses the impact of intravenous insulin boluses on parameter identification precision., Method: Insulin and C-peptide data from two intravenous glucose tolerance test (IVGTT) trials of healthy adults ( N = 10 × 2; denoted A and B), with (A) and without (B) insulin modification, were used to identify insulin kinetics parameters using a grid search. Monte Carlo analysis ( N = 1000) quantifies variation in simulation error for insulin assay errors of 5%. A region of parameter values around the optimum was identified whose errors are within variation due to assay error. A smaller optimal region indicates more precise practical identifiability. Trial results were compared to assess identifiability and precision., Results: Trial B, without insulin modification, has optimal parameter regions 4.7 times larger on average than Trial A, with 1-U insulin bolus modification. Ranges of optimal parameter values between trials A and B increase from 0.04 to 0.12 min -1 for hepatic clearance and from 0.07 to 0.14 for first-pass clearance on average. Trial B's optimal values frequently lie outside physiological ranges, further indicating lack of distinct identifiability., Conclusions: A small 1-U insulin bolus improves identification of hepatic clearance parameters by providing a smaller region of optimal parameter values. Adding an insulin bolus in metabolic tests can significantly improve identifiability and outcome test precision. Assay errors necessitate insulin modification in clinical tests to ensure identifiability and precision.

Unaccustomed exercise causes muscle damage resulting in loss of muscle function, which may be attributable to exercise-induced increases in skeletal muscle reactive oxygen species. This study examined the effect of mitochondria-targeted antioxidant supplementation on recovery of muscle function following exercise. Thirty-two untrained men received MitoQ (20 mg/day) or a placebo for 14 days before performing  300 maximal eccentric contractions of the knee extensor muscles of 1 leg. Muscle function was assessed using isokinetic dynamometry before, immediately after, and 24, 48, 72, and 168 hours after exercise. Muscle soreness was assessed using a visual analogue scale 24, 48, 72, and 168 hours after exercise. Blood samples were collected before, immediately after, and 2, 24, 48, 72, and 168 hours after exercise and urine samples were collected before and during the 48 hours after exercise. The reduction in maximal voluntary isometric contraction force and peak concentric torque following exercise was unaffected by MitoQ while recovery of peak eccentric torque was delayed in the MitoQ group. Exercise-induced increases in urine F 2 -isoprostanes were unaffected by MitoQ. MitoQ augmented exercise-induced increases in plasma creatine kinase levels, while plasma IL-6 was similar between groups. Muscle soreness was not affected by MitoQ. These results indicate that MitoQ does not attenuate post-exercise muscle soreness and may delay recovery of muscle function following eccentric exercise. Trial registration number: ACTRN12620001089921. Novelty: Post-exercise recovery of maximal voluntary isometric contraction force and peak concentric torque were unaffected by MitoQ. MitoQ delayed post-exercise recovery of peak eccentric torque. Post-exercise muscle soreness was unaffected by MitoQ.

The role of mitochondrial ROS in signalling muscle adaptations to exercise training has not been explored in detail. We investigated the effect of supplementation with the mitochondria-targeted antioxidant MitoQ on a) the skeletal muscle mitochondrial and antioxidant gene transcriptional response to acute high-intensity exercise and b) skeletal muscle mitochondrial content and function following exercise training. In a randomised, double-blind, placebo-controlled, parallel design study, 23 untrained men (age: 44 ± 7 years, VO 2peak : 39.6 ± 7.9 ml/kg/min) were randomised to receive either MitoQ (20 mg/d) or a placebo for 10 days before completing a bout of high-intensity interval exercise (cycle ergometer, 10 × 60 s at VO 2peak workload with 75 s rest). Blood samples and vastus lateralis muscle biopsies were collected before exercise and immediately and 3 h after exercise. Participants then completed high-intensity interval training (HIIT; 3 sessions per week for 3 weeks) and another blood sample and muscle biopsy were collected. There was no effect of acute exercise or MitoQ on systemic (plasma protein carbonyls and reduced glutathione) or skeletal muscle (mtDNA damage and 4-HNE) oxidative stress biomarkers. Acute exercise-induced increases in skeletal muscle peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1-α) mRNA expression were augmented in the MitoQ group. Despite this, training-induced increases in skeletal muscle mitochondrial content were similar between groups. HIIT-induced increases in VO 2peak and 20 km time trial performance were also similar between groups while training-induced increases in peak power achieved during the VO 2peak test were augmented in the MitoQ group. These data suggest that training-induced increases in peak power are enhanced following MitoQ supplementation, which may be related to the augmentation of skeletal muscle PGC1α expression following acute exercise. However, these effects do not appear to be related to an effect of MitoQ supplementation on exercise-induced oxidative stress or training-induced mitochondrial biogenesis in skeletal muscle., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Objective: The minor allele (A) of the rs373863828 variant (p.Arg457Gln) in CREBRF is restricted to indigenous peoples of the Pacific islands (including New Zealand Māori and peoples of Polynesia), with a frequency of up to 25% in these populations. This allele associates with a large increase in body mass index (BMI) but with significantly lower risk of type-2 diabetes (T2D). It remains unclear whether the increased BMI is driven by increased adiposity or by increased lean mass., Methods: We undertook body composition analysis using DXA in 189 young men of Māori and Pacific descent living in Aotearoa New Zealand. Further investigation was carried out in two orthologous Arg458Gln knockin mouse models on FVB/NJ and C57BL/6j backgrounds., Results: The rs373863828 A allele was associated with lower fat mass when adjusted for BMI (p < 0.05) and was associated with significantly lower circulating levels of the muscle inhibitory hormone myostatin (p < 0.05). Supporting the human data, significant reductions in adipose tissue mass were observed in the knockin mice. This was more significant in older mice in both backgrounds and appeared to be the result of reduced age-associated increases in fat mass. The older male knockin mice on C57BL/6j background also had increased grip strength (p < 0.01) and lower levels of myostatin (p < 0.05)., Conclusion: Overall, these results prove that the rs373863828 A-allele is associated with a reduction of myostatin levels which likely contribute to an age-dependent lowering of fat mass, at least in males., (Copyright © 2022 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Reactive oxygen species (ROS) generated during exercise are considered integral for the health-promoting effects of exercise. However, the precise mechanisms by which exercise and ROS promote metabolic health remain unclear. Here, we demonstrate that skeletal muscle NADPH oxidase 4 (NOX4), which is induced after exercise, facilitates ROS-mediated adaptive responses that promote muscle function, maintain redox balance, and prevent the development of insulin resistance. Conversely, reductions in skeletal muscle NOX4 in aging and obesity contribute to the development of insulin resistance. NOX4 deletion in skeletal muscle compromised exercise capacity and antioxidant defense and promoted oxidative stress and insulin resistance in aging and obesity. The abrogated adaptive mechanisms, oxidative stress, and insulin resistance could be corrected by deleting the H 2 O 2 -detoxifying enzyme GPX-1 or by treating mice with an agonist of NFE2L2, the master regulator of antioxidant defense. These findings causally link NOX4-derived ROS in skeletal muscle with adaptive responses that promote muscle function and insulin sensitivity.

Acute exercise, and in particular aerobic exercise, increases skeletal muscle energy demand causing mitochondrial stress, and mitochondrial-related adaptations which are a hallmark of exercise training. Given that mitochondria are central players in the exercise response, it is imperative that they have networks that can communicate their status both intra- and inter-cellularly. Peptides encoded by short open-reading frames within mitochondrial DNA, mitochondrial-derived peptides (MDPs), have been suggested to form a newly recognised branch of this retrograde signalling cascade that contribute to coordinating the adaptive response to regular exercise. Here we summarise the recent evidence that acute high intensity exercise in humans can increase concentrations of the MDPs humanin and MOTS-c in skeletal muscle and plasma, and speculate on the mechanisms controlling MDP responses to exercise stress. Evidence that exercise training results in chronic changes in MDP expression within tissues and the circulation is conflicting and may depend on the mode, duration, intensity of training plan and participant characteristics. Further research is required to define the effect of these variables on MDPs and to determine whether MDPs other than MOTS-c have exercise mimetic properties. MOTS-c treatment of young and aged mice improves exercise capacity/performance and leads to adaptions that are similar to that of being physically active (weight loss, increased antioxidant capacity and improved insulin sensitivity), however, studies utilising a MOTS-c inactivating genetic variant or combination of exercise + MOTS-c treatment in mice suggest that there are distinct and overlapping pathways through which exercise and MOTS-c evoke metabolic benefits. Overall, MOTS-c, and potentially other MDPs, may be exercise-sensitive myokines and further work is required to define inter- and intra-tissue targets in an exercise context., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Aims/hypothesis: The minor A allele of rs373863828 (CREBRF p.Arg457Gln) is associated with increased BMI, but reduced risk of type 2 and gestational diabetes in Polynesian (Pacific peoples and Aotearoa New Zealand Māori) populations. This study investigates the effect of the A allele on insulin release and sensitivity in overweight/obese men without diabetes., Methods: A mixed meal tolerance test was completed by 172 men (56 with the A allele) of Māori or Pacific ancestry, and 44 (24 with the A allele) had a frequently sampled IVGTT and hyperinsulinaemic-euglycaemic clamp. Mixed linear models with covariates age, ancestry and BMI were used to analyse the association between the A allele of rs373863828 and markers of insulin release and blood glucose regulation., Results: The A allele of rs373863828 is associated with a greater increase in plasma insulin 30 min following a meal challenge without affecting the elevation in plasma glucose or incretins glucagon-like polypeptide-1 or gastric inhibitory polypeptide. Consistent with this point, following an i.v. infusion of a glucose bolus, participants with an A allele had higher early (p < 0.05 at 2 and 4 min) plasma insulin and C-peptide concentrations for a similar elevation in blood glucose as those homozygous for the major (G) allele. Despite increased plasma insulin, rs373863828 genotype was not associated with a significant difference (p > 0.05) in insulin sensitivity index or glucose disposal during hyperinsulinaemic-euglycaemic clamp., Conclusions/interpretation: rs373863828-A allele associates with increased glucose-stimulated insulin release without affecting insulin sensitivity, suggesting that CREBRF p.Arg457Gln may increase insulin release to reduce the risk of type 2 diabetes., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Mitochondrial-derived peptides (MDPs) are encoded by the mitochondrial genome and hypothesised to form part of a retrograde signalling network that modulates adaptive responses to metabolic stress. To understand how metabolic stress regulates MDPs in humans we assessed the association between circulating MOTS-c and SHLP2 and components of metabolic syndrome (MS), as well as depot-specific fat mass in participants without overt type 2 diabetes or cardiovascular disease. One-hundred and twenty-five Chinese participants (91 male, 34 female) had anthropometry, whole body dual-energy X-ray absorptiometry scans and fasted blood samples analysed. Chinese female participants and an additional 34 European Caucasian female participants also underwent magnetic resonance imaging and spectroscopy (MRI/S) for visceral, pancreatic and liver fat quantification. In Chinese participants (age = 41 ± 1 years, BMI = 27.8 ± 3.9 kg/m 2 ), plasma MOTS-c (315 ± 27 pg/ml) and SHLP2 (1393 ± 82 pg/ml) were elevated in those with MS (n = 26). While multiple components of the MS sequelae positively associated with both MOTS-c and SHLP2, including blood pressure, fasting plasma glucose and triglycerides, the most significant of these was waist circumference (p < 0.0001). Android fat had a greater effect on increasing plasma MOTS-c (p < 0.004) and SHLP2 (p < 0.009) relative to whole body fat. Associations with MRI/S parameters corrected for total body fat mass revealed that liver fat positively associated with plasma MOTS-c and SHLP2 and visceral fat with SHLP2. Consistent with hepatic stress being a driver of circulating MDP concentrations, plasma MOTS-c and SHLP2 were higher in participants with elevated liver damage markers and in male C57Bl/6j mice fed a diet that induces hepatic lipid accumulation and damage. Our findings provide evidence that in the absence of overt type 2 diabetes, components of the MS positively associated with levels of MOTS-c and SHLP2 and that android fat, in particular liver fat, is a primary driver of these associations. MOTS-c and SHLP2 have previously been shown to have cyto- and metabolo-protective properties, therefore we suggest that liver stress may be a mitochondrial peptide signal, and that mitochondrial peptides are part of a hepatic centric-hormetic response intended to restore metabolic balance., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Cancer causes mitochondrial alterations in skeletal muscle, which may progress to muscle wasting and, ultimately, to cancer cachexia. Understanding how exercise adaptations are altered by cancer and cancer treatment is important for the effective design of exercise interventions aimed at improving cancer outcomes. We conducted an exploratory study to investigate how tumor burden and cancer immunotherapy treatment (anti-PD-1) modify the skeletal muscle mitochondrial response to exercise training in mice with transplantable tumors (B16-F10 melanoma and EO771 breast cancer). Mice remained sedentary or were provided with running wheels for ~19 days immediately following tumor implant while receiving no treatment (Untreated), isotype control antibody (IgG2a) or anti-PD-1. Exercise and anti-PD-1 did not alter the growth rate of either tumor type, either alone or in combination therapy. Untreated mice with B16-F10 tumors showed increases in most measured markers of skeletal muscle mitochondrial content following exercise training, as did anti-PD-1-treated mice, suggesting increased mitochondrial content following exercise training in these groups. However, mice with B16-F10 tumors receiving the isotype control antibody did not exhibit increased skeletal muscle mitochondrial content following exercise. In untreated mice with EO771 tumors, only citrate synthase activity and complex IV activity were increased following exercise. In contrast, IgG2a and anti-PD-1-treated groups both showed robust increases in most measured markers following exercise. These results indicate that in mice with B16-F10 tumors, IgG2a administration prevents exercise adaptation of skeletal muscle mitochondria, but adaptation remains intact in mice receiving anti-PD-1. In mice with EO771 tumors, both IgG2a and anti-PD-1-treated mice show robust skeletal muscle mitochondrial exercise responses, while untreated mice do not. Taken together, we postulate that immune modulation may enhance skeletal muscle mitochondrial response to exercise in tumor-bearing mice, and suggest this as an exciting new avenue for future research in exercise oncology., Competing Interests: The authors have declared that no competing interests exist.

Neutrophils accumulate in insulin-sensitive tissues during obesity and may play a role in impairing insulin sensitivity. The major serine protease expressed by neutrophils is neutrophil elastase (NE), which is inhibited endogenously by α1-antitrypsin A (A1AT). We investigated the effect of exogenous (A1AT) treatment on diet-induced metabolic dysfunction. Male C57Bl/6j mice fed a chow or a high-fat diet (HFD) were randomized to receive intraperitoneal injections three times weekly of either Prolastin (human A1AT; 2 mg) or vehicle (PBS) for 10 wk. Prolastin treatment did not affect plasma NE concentration, body weight, glucose tolerance, or insulin sensitivity in chow-fed mice. In contrast, Prolastin treatment attenuated HFD-induced increases in plasma and white adipose tissue (WAT) NE without affecting circulatory neutrophil levels or increases in body weight. Prolastin-treated mice fed a HFD had improved insulin sensitivity, as assessed by insulin tolerance test, and this was associated with higher insulin-dependent IRS-1 (insulin receptor substrate) and Akt Ser473 phosphorylation, and reduced inflammation markers in WAT but not liver or muscle. In 3T3-L1 adipocytes, Prolastin reversed recombinant NE-induced impairment of insulin-stimulated glucose uptake and IRS-1 phosphorylation. Furthermore, PDGF mediated p-Akt Ser473 activation and glucose uptake (which is independent of IRS-1) was not affected by recombinant NE treatment. Collectively, our findings suggest that NE infiltration of WAT during metabolic overload contributes to insulin resistance by impairing insulin-induced IRS-1 signaling. NEW & NOTEWORTHY Neutrophils accumulate in peripheral tissues during obesity and are critical coordinators of tissue inflammatory responses. Here, we provide evidence that inhibition of the primary neutrophil protease, neutrophil elastase, with α1-antitrypsin A (A1AT) can improve insulin sensitivity and glucose homeostasis of mice fed a high-fat diet. This was attributed to improved insulin-induced IRS-1 phosphorylation in white adipose tissue and provides further support for a role of neutrophils in mediating diet-induced peripheral tissue insulin resistance.

Background: Exercise increases skeletal muscle reactive oxygen species (ROS) production, which may contribute to the onset of muscular fatigue and impair athletic performance. Mitochondria-targeted antioxidants such as MitoQ, which contains a ubiquinone moiety and is targeted to mitochondria through the addition of a lipophilic triphenylphosphonium cation, are becoming popular amongst active individuals as they are designed to accumulate within mitochondria and may provide targeted protection against exercise-induced oxidative stress. However, the effect of MitoQ supplementation on cycling performance is currently unknown. Here, we investigate whether MitoQ supplementation can improve cycling performance measured as time to complete an 8 km time trial., Method: In a randomized, double-blind, placebo-controlled crossover study, 19 middle-aged (age: 44 ± 4 years) recreationally trained (VO 2peak : 58.5 ± 6.2 ml·kg - 1 ·min - 1 , distance cycled per week during 6 months prior to study enrollment: 158.3 ± 58.4 km) male cyclists completed 45 min cycling at 70% VO 2peak followed by an 8 km time trial after 28 days of supplementation with MitoQ (20 mg·day - 1 ) and a placebo. Free F 2 -isoprostanes were measured in plasma samples collected at rest, after 45 min cycling at 70% VO 2peak and after completion of the time trial. Respiratory gases and measures of rating of perceived exertion (RPE) were also collected., Results: Mean completion time for the time trial was 1.3% faster with MitoQ (12.91 ± 0.94 min) compared to placebo (13.09 ± 0.95 min, p = 0.04, 95% CI [0.05, 2.64], d = 0.2). There was no difference in RPE during the time trial between conditions (p = 0.82) despite there being a 4.4% increase in average power output during the time trial following MitoQ supplementation compared to placebo (placebo; 270 ± 51 W, MitoQ; 280 ± 53 W, p = 0.04, 95% CI [0.49, 8.22], d = 0.2). Plasma F 2 -isoprostanes were lower on completion of the time trial following MitoQ supplementation (35.89 ± 13.6 pg·ml - 1 ) compared to placebo (44.7 ± 16.9 pg·ml - 1 p = 0.03)., Conclusion: These data suggest that MitoQ supplementation may be an effective nutritional strategy to attenuate exercise-induced increases in oxidative damage to lipids and improve cycling performance., (© 2021. The Author(s).)

Key Points: Loss of β-catenin impairs in vivo and isolated muscle exercise/contraction-stimulated glucose uptake. β-Catenin is required for exercise-induced skeletal muscle actin cytoskeleton remodelling. β-Catenin 675 phosphorylation during exercise may be intensity dependent., Abstract: The conserved structural protein β-catenin is an emerging regulator of vesicle trafficking in multiple tissues and supports insulin-stimulated glucose transporter 4 (GLUT4) translocation in skeletal muscle by facilitating cortical actin remodelling. Actin remodelling may be a convergence point between insulin and exercise/contraction-stimulated glucose uptake. Here we investigated whether β-catenin is involved in regulating exercise/contraction-stimulated glucose uptake. We report that the muscle-specific deletion of β-catenin induced in adult mice (BCAT-mKO) impairs both exercise- and contraction (isolated muscle)-induced glucose uptake without affecting running performance or canonical exercise signalling pathways. Furthermore, high intensity exercise in mice and contraction of myotubes and isolated muscles led to the phosphorylation of β-catenin S675 , and this was impaired by Rac1 inhibition. Moderate intensity exercise in control and Rac1 muscle-specific knockout mice did not induce muscle β-catenin S675 phosphorylation, suggesting exercise intensity-dependent regulation of β-catenin S675 . Introduction of a non-phosphorylatable S675A mutant of β-catenin into myoblasts impaired GLUT4 translocation and actin remodelling stimulated by carbachol, a Rac1 and RhoA activator. Exercise-induced increases in cross-sectional phalloidin staining (F-actin marker) of gastrocnemius muscle was impaired in muscle from BCAT-mKO mice. Collectively our findings suggest that β-catenin is required for optimal glucose transport in muscle during exercise/contraction, potentially via facilitating actin cytoskeleton remodelling., (© 2021 The Authors. The Journal of Physiology © 2021 The Physiological Society.)

While it has long been known that contraction robustly stimulates skeletal muscle glucose uptake, the molecular steps regulating this increase remain incompletely defined. The mammalian ortholog of Sir2, sirtuin 1 (SIRT1), is an NAD + -dependent protein deacetylase that is thought to link perturbations in energy flux associated with exercise to subsequent cellular adaptations. Nevertheless, its role in contraction-stimulated glucose uptake has not been described. The objective of this study was to determine the importance of SIRT1 to contraction-stimulated glucose uptake in mouse skeletal muscle. Using a radioactive 2-deoxyglucose uptake (2DOGU) approach, we measured ex vivo glucose uptake in unstimulated (rested) and electrically stimulated (100 Hz contraction every 15 s for 10 min; contracted) extensor digitorum longus (EDL) and soleus from ∼15-wk-old male and female mice with muscle-specific knockout of SIRT1 deacetylase activity and their wild-type littermates. Skeletal muscle force decreased over the contraction protocol, although there were no differences in the rate of fatigue between genotypes. In EDL and soleus, loss of SIRT1 deacetylase activity did not affect contraction-induced increase in glucose uptake in either sex. Interestingly, the absolute rate of contraction-stimulated 2DOGU was ∼1.4-fold higher in female compared with male mice, regardless of muscle type. Taken together, our findings demonstrate that SIRT1 is not required for contraction-stimulated glucose uptake in mouse skeletal muscle. Moreover, to our knowledge, this is the first demonstration of sex-based differences in contraction-stimulated glucose uptake in mouse skeletal muscle. NEW & NOTEWORTHY Here, we demonstrate that glucose uptake in response to ex vivo contractions is not affected by the loss of sirtuin 1 (SIRT1) deacetylase function in muscle, regardless of sex or muscle type. Interestingly, however, similar to studies on insulin-stimulated glucose uptake, we demonstrate that contraction-stimulated glucose uptake is robustly higher in female compared with the male skeletal muscle. To our knowledge, this is the first demonstration of sex-based differences in contraction-stimulated glucose uptake in skeletal muscle.

Disuse-induced muscle atrophy is accompanied by a blunted postprandial response of the mammalian target of rapamycin complex 1 (mTORC1) pathway. Conflicting observations exist as to whether postabsorptive mTORC1 pathway activation is also blunted by disuse and plays a role in atrophy. It is unknown whether changes in habitual protein intake alter mTORC1 regulatory proteins and how they may contribute to the development of anabolic resistance. The primary objective of this study was to characterize the downstream responsiveness of skeletal muscle mTORC1 activation and its upstream regulatory factors, following 14 days of lower limb disuse in middle-aged men (45-60 yr). The participants were further randomized to receive daily supplementation of 20 g/d of protein ( n = 12; milk protein concentrate) or isocaloric carbohydrate placebo ( n = 13). Immobilization reduced postabsorptive skeletal muscle phosphorylation of the mTORC1 downstream targets, 4E-BP1, P70S6K, and ribosomal protein S6 (RPS6), with phosphorylation of the latter two decreasing to a greater extent in the placebo, compared with the protein supplementation groups (37% ± 13% vs. 14% ± 11% and 38% ± 20% vs. 25% ± 8%, respectively). Sestrin2 protein was also downregulated following immobilization irrespective of supplement group, despite a corresponding increase in its mRNA content. This decrease in Sestrin2 protein was negatively correlated with the immobilization-induced change in the in silico-predicted regulator miR-23b-3p. No other measured upstream proteins were altered by immobilization or supplementation. Immobilization downregulated postabsorptive mTORC1 pathway activation, and 20 g/day of protein supplementation attenuated the decrease in phosphorylation of targets regulating muscle protein synthesis.

BYL719 (alpelisib) is a small molecule inhibitor of PI3K p110α developed for cancer therapy. Targeted suppression of PI3K has led to lifespan extension in rodents and model organisms. If PI3K inhibitors are to be considered as an aging therapeutic, it is important to understand the potential consequences of long-term exposure, and the most practical way to achieve this is through diet administration. Here, we investigated the pharmacokinetics of BYL719 delivered in diet and the efficacy of BYL719 to suppress insulin signaling when administered in the diet of 8-month-old male and female mice. Compared to oral gavage, diet incorporation resulted in a lower peak plasma BYL719 (3.6 vs. 9.2 μM) concentration but similar half-life (~1.5 h). Consuming BYL719 resulted in decreased insulin signaling in liver and muscle within 72 h, and mice still showed impaired glucose tolerance and insulin sensitivity following 6 weeks of access to a diet containing 0.3 g/kg BYL719. However, consuming BYL719 did not affect food intake, body mass, muscle function (rotarod and hang time performance) or cognitive behaviors. This provides evidence that BYL719 has long-term efficacy without major toxicity or side effects, and suggests that administering BYL719 in diet is suitable for studying the effect of pharmacological suppression of PI3K p110α on aging and metabolic function.

Healthy aging can be promoted by enhanced metabolic fitness and physical capacity. Mitochondria are chief metabolic organelles with strong implications in aging that also coordinate broad physiological functions, in part, using peptides that are encoded within their independent genome. However, mitochondrial-encoded factors that actively regulate aging are unknown. Here, we report that mitochondrial-encoded MOTS-c can significantly enhance physical performance in young (2 mo.), middle-age (12 mo.), and old (22 mo.) mice. MOTS-c can regulate (i) nuclear genes, including those related to metabolism and proteostasis, (ii) skeletal muscle metabolism, and (iii) myoblast adaptation to metabolic stress. We provide evidence that late-life (23.5 mo.) initiated intermittent MOTS-c treatment (3x/week) can increase physical capacity and healthspan in mice. In humans, exercise induces endogenous MOTS-c expression in skeletal muscle and in circulation. Our data indicate that aging is regulated by genes encoded in both of our co-evolved mitochondrial and nuclear genomes.

Objective: Skeletal muscle glucose disposal following a meal is mediated through insulin-stimulated movement of the GLUT4-containing vesicles to the cell surface. The highly conserved scaffold-protein β-catenin is an emerging regulator of vesicle trafficking in other tissues. Here, we investigated the involvement of β-catenin in skeletal muscle insulin-stimulated glucose transport., Methods: Glucose homeostasis and transport was investigated in inducible muscle specific β-catenin knockout (BCAT-mKO) mice. The effect of β-catenin deletion and mutation of β-catenin serine 552 on signal transduction, glucose uptake and protein-protein interactions were determined in L6-G4-myc cells, and β-catenin insulin-responsive binding partners were identified via immunoprecipitation coupled to label-free proteomics., Results: Skeletal muscle specific deletion of β-catenin impaired whole-body insulin sensitivity and insulin-stimulated glucose uptake into muscle independent of canonical Wnt signalling. In response to insulin, β-catenin was phosphorylated at serine 552 in an Akt-dependent manner, and in L6-G4-myc cells, mutation of β-catenin S552 impaired insulin-induced actin-polymerisation, resulting in attenuated insulin-induced glucose transport and GLUT4 translocation. β-catenin was found to interact with M-cadherin in an insulin-dependent β-catenin S552 -phosphorylation dependent manner, and loss of M-cadherin in L6-G4-myc cells attenuated insulin-induced actin-polymerisation and glucose transport., Conclusions: Our data suggest that β-catenin is a novel mediator of glucose transport in skeletal muscle and may contribute to insulin-induced actin-cytoskeleton remodelling to support GLUT4 translocation., (Copyright © 2020 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Genetic inhibition of the p110α isoform of phosphatidylinositol-3-kinase (PI3K) can increase murine lifespan, enhance mitochondrial function and alter tissue-specific oxidative balance. Here, we investigated whether pharmacological inhibition of the p110α isoform of PI3K induces similar enhancement of mitochondrial function in middle-aged mice. Eight-month-old male and female mice were fed a diet containing 0.3 g/kg of the p110α-selective inhibitor BYL-719 (BYL) or a vehicle diet (VEH) for 6 weeks. Mice consuming BYL-719 had higher blood glucose and insulin, and tended towards decreased body weight. After 72 h, gene expression of the mitochondrial biogenesis mediators Pgc1α, Tfam and Nrf1 was greater in liver of BYL-719 males only, but unchanged in skeletal muscle of either sex. Six weeks of BYL-719 treatment did not affect mitochondrial content or function in the liver or skeletal muscle of either sex. In livers of males only, the expression of the antioxidant genes Nfe2l2, Cat, Sod1 and Sod2 increased within 72 h of BYL-719 treatment, and remained higher after 6 weeks. This was associated with an increase in hepatic GSH content and catalase protein expression, and lower H2O2 levels. Our results suggest that pharmacological inhibition of p110α in adult mice does not affect liver or skeletal muscle mitochondrial function, but does show sex- and tissue-specific effects on up-regulation of antioxidant response., (© 2020 The Author(s).)

Mitochondrial-derived peptides (MDPs) are small bioactive peptides encoded by short open-reading frames (sORF) in mitochondrial DNA that do not necessarily have traditional hallmarks of protein-coding genes. To date, eight MDPs have been identified, all of which have been shown to have various cyto- or metaboloprotective properties. The 12S ribosomal RNA ( MT-RNR1 ) gene harbors the sequence for MOTS-c, whereas the other seven MDPs [humanin and small humanin-like peptides (SHLP) 1-6] are encoded by the 16S ribosomal RNA gene. Here, we review the evidence that endogenous MDPs are sensitive to changes in metabolism, showing that metabolic conditions like obesity, diabetes, and aging are associated with lower circulating MDPs, whereas in humans muscle MDP expression is upregulated in response to stress that perturbs the mitochondria like exercise, some mtDNA mutation-associated diseases, and healthy aging, which potentially suggests a tissue-specific response aimed at restoring cellular or mitochondrial homeostasis. Consistent with this, treatment of rodents with humanin, MOTS-c, and SHLP2 can enhance insulin sensitivity and offer protection against a range of age-associated metabolic disorders. Furthermore, assessing how mtDNA variants alter the functions of MDPs is beginning to provide evidence that MDPs are metabolic signal transducers in humans. Taken together, MDPs appear to form an important aspect of a retrograde signaling network that communicates mitochondrial status with the wider cell and to distal tissues to modulate adaptative responses to metabolic stress. It remains to be fully determined whether the metaboloprotective properties of MDPs can be harnessed into therapies for metabolic disease.

New Findings: What is the central question of this study? Does short-term high-intensity interval training alter the composition of the microbiome and is this associated with exercise-induced improvements in cardiorespiratory fitness and insulin sensitivity? What is the main finding and its importance? Although high-intensity interval training increased insulin sensitivity and cardiovascular fitness, it did not alter the composition of the microbiome. This suggests that changes in the composition of the microbiome that occur with prolonged exercise training might be in response to changes in metabolic health rather than driving exercise training-induced adaptations., Abstract: Regular exercise reduces the risk of metabolic diseases, and the composition of the gut microbiome has been associated with metabolic function. We investigated whether short-term high-intensity interval training (HIIT) altered the diversity and composition of the bacterial community and whether there were associations with markers of insulin sensitivity or aerobic fitness. Cardiorespiratory fitness ( V ̇ O 2 peak ) and body composition (dual energy X-ray absorptiometry scan) were assessed and faecal and fasted blood samples collected from 14 lean (fat mass 21 ± 2%, aged 29 ± 2 years) and 15 overweight (fat mass 33 ± 2%, aged 31 ± 2 years) men before and after 3 weeks of HIIT training (8-12 × 60 s cycle ergometer bouts at V ̇ O 2 peak power output interspersed by 75 s rest, three times per week). Gut microbiome composition was analysed by 16S rRNA gene amplicon sequencing. The HIIT significantly increased the aerobic fitness of both groups (P < 0.001) and improved markers of insulin sensitivity (lowered fasted insulin and HOMA-IR; P < 0.001) in the overweight group. Despite differences in the abundance of several bacterial taxa being evident between the lean and overweight group, HIIT did not affect the overall bacterial diversity or community structure (α-diversity or β-diversity). No associations were found between the top 50 most abundant bacterial genera and cardiorespiratory fitness markers; however, significant associations (P < 0.05) were observed between the abundance of the bacterial species Coprococcus&#95;3, Blautia, Lachnospiraceae&#95;ge and Dorea and insulin sensitivity markers in the overweight group. Our results suggest that short-term HIIT does not greatly impact the overall composition of the gut microbiome, but that certain microbiome genera are associated with insulin sensitivity markers that were improved by HIIT in overweight participants., (© 2020 The Authors. Experimental Physiology © 2020 The Physiological Society.)

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Purpose: Excess production of reactive oxygen species (ROS) from the mitochondria can promote mitochondrial dysfunction and has been implicated in the development of a range of chronic diseases. As such there is interest in whether mitochondrial-targeted antioxidant supplementation can attenuate mitochondrial-associated oxidative stress. We investigated the effect of MitoQ and CoQ10 supplementation on oxidative stress and skeletal muscle mitochondrial ROS levels and function in healthy middle-aged men., Methods: Skeletal muscle and blood samples were collected from twenty men (50 ± 1 y) before and following six weeks of daily supplementation with MitoQ (20 mg) or CoQ10 (200 mg). High-resolution respirometry was used to determine mitochondrial respiration and H 2 O 2 levels, markers of mitochondrial mass and antioxidant defences were measured in muscle samples and oxidative stress markers in urine and blood samples., Results: Both MitoQ and CoQ10 supplementation suppressed mitochondrial net H 2 O 2 levels during leak respiration, while MitoQ also elevated muscle catalase expression. However, neither supplement altered urine F 2 -isoprostanes nor plasma TBARS levels. Neither MitoQ nor CoQ10 supplementation had a significant impact on mitochondrial respiration or mitochondrial density markers (citrate synthase, mtDNA/nDNA, PPARGC1A, OXPHOS expression)., Conclusion: Our results suggest that neither MitoQ and CoQ10 supplements impact mitochondrial function, but both can mildly suppress mitochondrial ROS levels in healthy middle-aged men, with some indication that MitoQ may be more effective than CoQ10.

Humanin is a small regulatory peptide encoded within the 16S ribosomal RNA gene ( MT-RNR2 ) of the mitochondrial genome that has cellular cyto- and metabolo-protective properties similar to that of aerobic exercise training. Here we investigated whether acute high-intensity interval exercise or short-term high-intensity interval training (HIIT) impacted skeletal muscle and plasma humanin levels. Vastus lateralis muscle biopsies and plasma samples were collected from young healthy untrained men ( n = 10, 24.5 ± 3.7 yr) before, immediately following, and 4 h following the completion of 10 × 60 s cycle ergometer bouts at V̇o 2peak power output (untrained). Resting and postexercise sampling was also performed after six HIIT sessions (trained) completed over 2 wk. Humanin protein abundance in muscle and plasma were increased following an acute high-intensity exercise bout. HIIT trended ( P = 0.063) to lower absolute humanin plasma levels, without effecting the response in muscle or plasma to acute exercise. A similar response in the plasma was observed for the small humanin-like peptide 6 (SHLP6), but not SHLP2, indicating selective regulation of peptides encoded by MT-RNR2 gene. There was a weak positive correlation between muscle and plasma humanin levels, and contraction of isolated mouse EDL muscle increased humanin levels ~4-fold. The increase in muscle humanin levels with acute exercise was not associated with MT-RNR2 mRNA or humanin mRNA levels (which decreased following acute exercise). Overall, these results suggest that humanin is an exercise-sensitive mitochondrial peptide and acute exercise-induced humanin responses in muscle are nontranscriptionally regulated and may partially contribute to the observed increase in plasma concentrations. NEW & NOTEWORTHY Small regulatory peptides encoded within the mitochondrial genome (mitochondrial derived peptides) have been shown to have cellular cyto- and metabolo-protective roles that parallel those of exercise. Here we provide evidence that humanin and SHLP6 are exercise-sensitive mitochondrial derived peptides. Studies to determine whether mitochondrial derived peptides play a role in regulating exercise-induced adaptations are warranted.

Excessive insulin signaling through the insulin receptor (IR) may play a role in the pathogenesis of diet-induced metabolic disease, including obesity and type 2 diabetes. Here we investigate whether heterozygous impairment of insulin receptor (IR) expression limited to peripheral, i.e. non-CNS, tissues of adult mice impacts the development of high-fat diet-induced metabolic deterioration. While exhibiting some features of insulin resistance, PerIRKO +/- mice display a hepatic energy deficit accompanied by induction of energy-sensing AMPK, mitochondrial biogenesis, PPARα, unexpectedly leading to protection from, and reversal of hepatic lipid accumulation (steatosis hepatis, NAFLD). Consistently, and unlike in control mice, the PPARα activator fenofibrate fails to further affect hepatic lipid accumulation in PerIRKO +/- mice. Taken together, and opposing previously established diabetogenic features of insulin resistance, incomplete impairment of insulin signaling may mimic central aspects of calorie restriction to limit hepatic lipid accumulation during conditions of metabolic stress.

Mitochondria putatively regulate the aging process, in part, through the small regulatory peptide, mitochondrial open reading frame of the 12S rRNA-c (MOTS-c) that is encoded by the mitochondrial genome. Here we investigated the regulation of MOTS-c in the plasma and skeletal muscle of healthy aging men. Circulating MOTS-c reduced with age, but older (70-81 y) and middle-aged (45-55 y) men had ~1.5-fold higher skeletal muscle MOTS-c expression than young (18-30 y). Plasma MOTS-c levels only correlated with plasma in young men, was associated with markers of slow-type muscle, and associated with improved muscle quality in the older group (maximal leg-press load relative to thigh cross-sectional area). Using small mRNA assays we provide evidence that MOTS-c transcription may be regulated independently of the full length 12S rRNA gene in which it is encoded, and expression is not associated with antioxidant response element (ARE)-related genes as previously seen in culture. Our results suggest that plasma and muscle MOTS-c are differentially regulated with aging, and the increase in muscle MOTS-c expression with age is consistent with fast-to-slow type muscle fiber transition. Further research is required to determine the molecular targets of endogenous MOTS-c in human muscle but they may relate to factors that maintain muscle quality.

Background: The CREBRF missense variant (p.Arg457Gln) is paradoxically associated with lower risk of type 2 diabetes, yet higher body mass index (BMI). Here we sought to determine whether this CREBRF variant might be associated with adult height., Methods: Linear regression was used to analyse the association of the CREBRF minor (A) allele with height in 2286 Māori and Pacific adults living in Aotearoa/New Zealand. A potential type 2 diabetes index event was corrected to account for a bias that may be the cause of paradoxical association between the CREBRF diabetes-protective allele and higher BMI and height., Results: The CREBRF protective allele was associated with increased adult height (ß = 1.25 cm, P = 3.9 × 10 -6 ), with the effect being more pronounced in males. The lower odds of diabetes remained similar when analyses were adjusted for height (OR = 0.67-0.65). We found no evidence of a diabetes index event bias to explain the paradoxical effect of CREBRF with either BMI or height and diabetes. The orthologous CREBRF p.Arg457Gln variant was created in knock-in mice to independently assess the effect of the variant, and length was found to be greater in male mice at 8 weeks of age., Conclusion: These data taken together indicate that CREBRF p.Arg457Gln is associated with taller stature in Māori and Pacific adults.

Oxidative stress induced by acute exercise may regulate exercise training-induced adaptations that improve metabolic health. One of the central transcription regulatory targets of reactive oxygen species (ROS) is Nrf2 (nuclear factor erythroid-derived 2-like 2, or NFE2L2). Here, we investigated whether global deficiency of Nrf2 in mice would impact exercise training-induced changes in glucose and lipid homeostasis. We report that following 6 wk of treadmill exercise training, Nrf2-deficient mice have elevated fasting plasma triglycerides and free fatty acids and higher blood glucose levels following a meal despite having a similar fat mass to wild-type controls. This impaired glucose homeostasis appears to be related to reduced insulin sensitivity primarily in adipose and liver tissue, and although a clear mechanism was not evident, Nrf2-deficient mice had increased markers of hepatic oxidative stress and stress-related kinase activation in white adipose tissue (WAT) without overt inflammation alteration in WAT or modulation of hepatic and WAT fibroblast growth factor 21 gene expression. Our results suggest that Nrf2 facilitates exercise training-induced improvements in glucose homeostasis; however, further research is required to determine whether this occurs through direct regulation of exercise adaptations or via the maintenance of redox balance during training.

Background and Objectives: Excessive adipose tissue macrophage accumulation in obesity has been implicated in mediating inflammatory responses that impair glucose homeostasis and promote insulin resistance. Colony-stimulating factor 1 (CSF1) controls macrophage differentiation, and here we sought to determine the effect of a CSF1 receptor inhibitor, PLX3397, on adipose tissue macrophage levels and understand the impact on glucose homeostasis in mice., Methods: A Ten-week-old mice were fed a chow or high-fat diet for 10 weeks and then treated with PLX3397 via oral gavage (50 mg/kg) every second day for 3 weeks, with subsequent monitoring of glucose tolerance, insulin sensitivity and assessment of adipose tissue immune cells., Results: PLX3397 treatment substantially reduced macrophage numbers in adipose tissue of both chow and high-fat diet fed mice without affecting total myeloid cell levels. Despite this, PLX3397 did not greatly alter glucose homeostasis, did not affect high-fat diet-induced increases in visceral fat cytokine expression (Il-6 and Tnfa) and had limited effect on the phosphorylation of the stress kinases JNK and ERK and macrophage polarization., Conclusions: Our results indicate that macrophage infiltration of adipose tissue induced by a high-fat diet may not be the trigger for impairments in whole body glucose homeostasis, and that anti-CSF1 therapies are not likely to be useful as treatments for insulin resistance.

Measurement of skeletal muscle mitochondrial respiration requires invasive biopsy to obtain a muscle sample. Peripheral blood mononuclear cell (PBMC) mitochondrial protein content appears to reflect training status in young men; however, no studies have investigated whether there are training-induced changes in PBMC mitochondrial respiration. Therefore, we determined whether PBMC mitochondrial respiration could be used as a marker of skeletal muscle mitochondrial respiration in young healthy men and whether PBMC mitochondrial respiration responds to short-term training. Skeletal muscle and PBMC samples from 10 healthy young (18-35 yr) male participants were taken before and after a 2-wk high-intensity interval training protocol. High-resolution respirometry was used to determine mitochondrial respiration from muscle and PBMCs, and Western blotting and quantitative PCR were used to assess mitochondrial biogenesis in PBMCs. PBMC mitochondrial respiration was not correlated with muscle mitochondrial respiration at baseline ( R 2 = 0.012-0.364, P > 0.05). While muscle mitochondrial respiration increased in response to training (32.1-61.5%, P < 0.05), PBMC respiration was not affected by training. Consequently, PBMCs did not predict training effect on muscle mitochondrial respiration ( R 2 = 0.024-0.283, P > 0.05). Similarly, gene and protein markers of mitochondrial biogenesis did not increase in PBMCs following training. This suggests PBMC mitochondrial function does not reflect that of skeletal muscle and does not increase following short-term high-intensity training. PBMCs are therefore not a suitable biomarker for muscle mitochondrial function in young healthy men. It may be useful to study PBMC mitochondrial function as a biomarker of muscle mitochondrial function in pathological populations with different respiration capacities. NEW & NOTEWORTHY Research in primates has suggested that peripheral blood mononuclear cells (PBMCs) may provide a less-invasive alternative to a muscle biopsy for measuring muscle mitochondrial function. Furthermore, trained individuals appear to have greater mitochondrial content in PBMCs. Here we show that in healthy young men, PBMCs do not reflect skeletal muscle mitochondrial function and do not adapt in response to a training intervention that increases muscle mitochondrial function, suggesting PBMCs are a poor marker of muscle mitochondrial function in humans.

Pancreatitis is increasingly recognized as not merely a local inflammation of the pancreas but also a disease with high frequency of systemic sequelae. Current understanding of the cellular mechanisms that trigger it and affect the development of sequelae are limited. Genetically engineered mouse models can be a useful tool to study the pathophysiology of pancreatitis. This article gives an overview of the genetically engineered mouse models that spontaneously develop pancreatitis and discusses those that most closely replicate different pancreatitis hallmarks observed in humans.

MicroRNAs (miRNAs) regulate gene expression via transcript degradation and translational inhibition, and they may also function as long distance signaling molecules. Circulatory miRNAs are either protein-bound or packaged within vesicles (exosomes). Ten young men (24.6 ± 4.0 yr) underwent a single bout of high-intensity interval cycling exercise. Vastus lateralis biopsies and plasma were collected immediately before and after exercise, as well as 4 h following the exercise bout. Twenty-nine miRNAs previously reported to be regulated by acute exercise were assessed within muscle, venous plasma, and enriched circulatory exosomes via qRT-PCR. Of the 29 targeted miRNAs, 11 were altered in muscle, 8 in plasma, and 9 in the exosome fraction. Although changes in muscle and plasma expression were bidirectional, all regulated exosomal miRNAs increased following exercise. Three miRNAs were altered in all three sample pools (miR-1-3p, -16-5p, and -222-3p), three in both muscle and plasma (miR-21-5p, -134-3p, and -107), three in both muscle and exosomes (miR-23a-3p, -208a-3p, and -150-5p), and three in both plasma and exosomes (miR-486-5p, -126-3p, and -378a-5p). There was a marked discrepancy between the observed alterations between sample pools. A subset of exosomal miRNAs increased in abundance following exercise, suggesting an exercise-induced release of exosomes enriched in specific miRNAs. The uniqueness of the exosomal miRNA response suggests its relevance as a sample pool that needs to be further explored in better understanding biological functions.

One of the main sources of reactive oxygen species (ROS) in skeletal muscle is the mitochondria. Prolonged or very high ROS exposure causes oxidative damage, which can be deleterious to muscle function, and as such, there is growing interest in targeting antioxidants to the mitochondria in an effort to prevent or treat muscle dysfunction and damage associated with disease and injury. Paradoxically, however, ROS also act as important signalling molecules in controlling cellular homeostasis, and therefore caution must be taken when supplementing with antioxidants. It is possible that mitochondria-targeted antioxidants may limit oxidative stress without suppressing ROS from non-mitochondrial sources that might be important for cell signalling. Therefore, in this review, we summarise literature relating to the effect of mitochondria-targeted antioxidants on skeletal muscle function. Overall, mitochondria-targeted antioxidants appear to exert beneficial effects on mitochondrial capacity and function, insulin sensitivity and age-related declines in muscle function. However, it seems that this is dependent on the type of mitochondrial-trageted antioxidant employed, and its specific mechanism of action, rather than simply targeting to the mitochondria.

Purpose: Dietary protein and resistance exercise (RE) are both potent stimuli of the mammalian target of rapamycin complex 1 (mTORC1). Sestrins1, 2, 3 are multifunctional proteins that regulate mTORC1, stimulate autophagy and alleviate oxidative stress. Of this family, Sestrin2 is a putative leucine sensor implicated in mTORC1 and AMP-dependent protein kinase (AMPK) regulation. There is currently no data examining the responsiveness of Sestrin2 to dietary protein ingestion, with or without RE., Methods: In Study 1, 16 males ingested either 10 or 20 g of milk protein concentrate (MPC) with muscle biopsies collected pre, 90 and 210 min post-beverage consumption. In Study 2, 20 males performed a bout of RE immediately followed by the consumption of 9 g of MPC or carbohydrate placebo. Analysis of Sestrins, AMPK and antioxidant responses was examined., Results: Dietary protein ingestion did not result in Sestrin2 mobility shift. After RE, Sestrin2 phosphorylation state was significantly altered and was not further modified by post-exercise protein or carbohydrate ingestion. With RE, AMPK phosphorylation remained stable, while the mRNA expressions of several antioxidants were upregulated., Conclusions: Dietary protein ingestion did not affect the signalling by the family of Sestrins. With RE, Sestrin2 was hyperphosphorylated, with no further evidence of a relationship to AMPK signalling.

Impaired insulin/IGF1 signalling has been shown to extend lifespan in model organisms ranging from yeast to mammals. Here we sought to determine the effect of targeted disruption of the insulin receptor (IR) in non-neuronal tissues of adult mice on the lifespan. We induced hemizygous (PerIRKO +/- ) or homozygous (PerIRKO -/- ) disruption of the IR in peripheral tissue of 15-weeks-old mice using a tamoxifen-inducible Cre transgenic mouse with only peripheral tissue expression, and subsequently monitored glucose metabolism, insulin signalling and spontaneous death rates over 4 years. Complete peripheral IR disruption resulted in a diabetic phenotype with increased blood glucose and plasma insulin levels in young mice. Although blood glucose levels returned to normal, and fat mass was reduced in aged PerIRKO -/- mice, their lifespan was reduced. By contrast, heterozygous disruption had no effect on lifespan. This was despite young male PerIRKO +/- mice showing reduced fat mass and mild increase in hepatic insulin sensitivity. In conflict with findings in metazoans like Caenorhabditis elegans and Drosophila melanogaster, our results suggest that heterozygous impairment of the insulin signalling limited to peripheral tissues of adult mice fails to extend lifespan despite increased systemic insulin sensitivity, while homozygous impairment shortens lifespan., (© 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.)

Aims/hypothesis: In obesity oxidative stress is thought to contribute to the development of insulin resistance, non-alcoholic fatty liver disease and the progression to non-alcoholic steatohepatitis. Our aim was to examine the precise contributions of hepatocyte-derived H 2 O 2 to liver pathophysiology., Methods: Glutathione peroxidase (GPX) 1 is an antioxidant enzyme that is abundant in the liver and converts H 2 O 2 to water. We generated Gpx1 lox/lox mice to conditionally delete Gpx1 in hepatocytes (Alb-Cre;Gpx1 lox/lox ) and characterised mice fed chow, high-fat or choline-deficient amino-acid-defined (CDAA) diets., Results: Chow-fed Alb-Cre;Gpx1 lox/lox mice did not exhibit any alterations in body composition or energy expenditure, but had improved insulin sensitivity and reduced fasting blood glucose. This was accompanied by decreased gluconeogenic and increased glycolytic gene expression as well as increased hepatic glycogen. Hepatic insulin receptor Y1163/Y1163 phosphorylation and Akt Ser-473 phosphorylation were increased in fasted chow-fed Alb-Cre;Gpx1 lox/lox mice, associated with increased H 2 O 2 production and insulin signalling in isolated hepatocytes. The enhanced insulin signalling was accompanied by the increased oxidation of hepatic protein tyrosine phosphatases previously implicated in the attenuation of insulin signalling. High-fat-fed Alb-Cre;Gpx1 lox/lox mice did not exhibit alterations in weight gain or hepatosteatosis, but exhibited decreased hepatic inflammation, decreased gluconeogenic gene expression and increased insulin signalling in the liver. Alb-Cre;Gpx1 lox/lox mice fed a CDAA diet that promotes non-alcoholic steatohepatitis exhibited decreased hepatic lymphocytic infiltrates, inflammation and liver fibrosis., Conclusions/interpretation: Increased hepatocyte-derived H 2 O 2 enhances hepatic insulin signalling, improves glucose control and protects mice from the development of non-alcoholic steatohepatitis.