Physiologic Adaptation to Macronutrient Change Distorts Findings from Short Dietary Trials: Reanalysis of a Metabolic Ward Study.

An influential 2-wk cross-over feeding trial without a washout period purported to show advantages of a low-fat diet (LFD) compared with a low-carbohydrate diet (LCD) for weight control. In contrast to several other macronutrient trials, the diet order effect was originally reported as not significant. In light of a new analysis by the original investigative group identifying an order effect, we aimed to examine, in a reanalysis of publicly available data (16 of 20 original participants; 7 female; mean BMI, 27.8 kg/m2), the validity of the original results and the claims that trial data oppose the carbohydrate-insulin model of obesity (CIM). We found that energy intake on the LCD was much lower when this diet was consumed first compared with second (a difference of -1164 kcal/d, P = 3.6 × 10-13); the opposite pattern was observed for the LFD (924 kcal/d, P = 2.0 × 10-16). This carry-over effect was significant (P interaction = 0.0004) whereas the net dietary effect was not (P = 0.4). Likewise, the between-arm difference (LCD - LFD) was -320 kcal/d in the first period and +1771 kcal/d in the second. Body fat decreased with consumption of the LCD first and increased with consumption of this diet second (-0.69 ± 0.33 compared with 0.57 ± 0.32 kg, P = 0.007). LCD-first participants had higher ß-hydroxybutyrate levels while consuming the LCD and lower respiratory quotients while consuming LFD when compared with LFD-first participants on their respective diets. Change in insulin secretion as assessed by C-peptide in the first diet period predicted higher energy intake and less fat loss in the second period. These findings, which tend to support rather than oppose the CIM, suggest that differential (unequal) carry-over effects and short duration, with no washout period, preclude causal inferences regarding chronic macronutrient effects from this trial.

Total water intake guidelines are sufficient for optimal hydration in United States adults.

PURPOSE: Recent studies suggest that 24-h urine osmolality (UOsm) for optimal water intake should be maintained < 500 mmol·kg-1. The purpose of this study was to determine the total water intake (TWI) requirement for healthy adults to maintain optimal hydration as indicated by 24-h urine osmolality < 500 mmol·kg-1. METHODS: Twenty-four-hour UOsm was assessed in 49 men and 50 women residing in the United States (age: 41 ± 14 y, body mass index: 26.3 ± 5.2 kg·m-2). TWI was assessed from 7-day water turnover, using a dilution of deuterium oxide, corrected for metabolic water production. The diagnostic accuracy of TWI to identify UOsm < 500 mmol·kg-1 was evaluated using receiver operating characteristic (ROC) analysis in men and women separately. RESULTS: Twenty-four-hour UOsm was 482 ± 229 and 346 ± 182 mmol·kg-1 and TWI was 3.57 ± 1.10 L·d-1 and 3.20 ± 1.27 L·d-1 in men and women, respectively. ROC analysis for TWI detecting 24-h UOsm < 500 mmol·kg-1 in men yielded an area under the curve (AUC) of 77.4% with sensitivity, specificity, and threshold values of 83.3%, 64.5%, and 3.39 L·d-1, respectively. The AUC was 82.4% in women with sensitivity, specificity, and threshold values of 85.7%, 72.1%, and 2.61 L·d-1. CONCLUSION: Considering threshold values in men and women of 3.4 L·d-1 and 2.6 L·d-1, respectively, maintaining TWI in line with National Academy of Medicine guidelines of 3.7 L·d-1 in men and 2.7 L·d-1 in women should be sufficient for most individuals in the United States to maintain 24-h UOsm < 500 mmol·kg-1.

Cellular dehydration acutely degrades mood mainly in women: a counterbalanced, crossover trial.

It is unclear if mild-to-moderate dehydration independently affects mood without confounders like heat exposure or exercise. This study examined the acute effect of cellular dehydration on mood. Forty-nine adults (55 % female, age 39 (sd 8) years) were assigned to counterbalanced, crossover trials. Intracellular dehydration was induced with 2-h (0·1 ml/kg per min) 3 % hypertonic saline (HYPER) infusion or 0·9 % isotonic saline (ISO) as a control. Plasma osmolality increased in HYPER (pre 285 (sd 3), post 305 (sd 4) mmol/kg; P < 0·05) but remained unchanged in ISO (pre 285 (sd 3), post 288 (sd 3) mmol/kg; P > 0·05). Mood was assessed with the short version of the Profile of Mood States Questionnaire (POMS). The POMS sub-scale (confusion-bewilderment, depression-dejection, fatigue-inertia) increased in HYPER compared with ISO (P < 0·05). Total mood disturbance score (TMD) assessed by POMS increased from 10·3 (sd 0·9) to 16·6 (sd 1·7) in HYPER (P < 0·01), but not in ISO (P > 0·05). When TMD was stratified by sex, the increase in the HYPER trial was significant in females (P < 0·01) but not in males (P > 0·05). Following infusion, thirst and copeptin (surrogate for vasopressin) were also higher in females than in males (21·3 (sd 2·0), 14·1 (sd 1·4) pmol/l; P < 0·01) during HYPER. In conclusion, cellular dehydration acutely degraded specific aspects of mood mainly in women. The mechanisms underlying sex differences may be related to elevated thirst and vasopressin.

Smartphone-Based Analysis of Urine Reagent Strips Is Inaccurate for Assessing Underhydration.

Background: Proper hydration is vital for both exercise and general health. Although various methods for hydration assessment exist, many are not valid for either use or never tested. Introduction: The purpose of this study was to determine whether the uChek© smart phone application can be used to diagnose underhydration based on elevated urine specific gravity (USG) assessed by refractometry. Methods: One hundred forty-seven (n = 147) fresh human urine samples from young and middle-age adults were analyzed for USG with a refractometer and the uChek© application by reading the Siemens Multistix 10G urine reagent strip. Results: Bland-Altman analysis showed agreement of the two methods of assessment. Overall diagnostic ability of the uChek© to identify underhydration was fair (area under the curve 79%). However, the sensitivity to correctly identify underhydration was poor (60%) as well as the specificity of correctly identifying euhydration (53%). Conclusion: The uChek© application does not accurately detect underhydration.

Validity of Predictive Equations for Resting Energy Expenditure in Greek Adults.

AIM: To examine the validity of published resting energy expenditure (REE) equations in Greek adults, and if indicated, develop new cohort-specific predictive REE equations. METHODS: Indirect calorimetry and anthropometric data were obtained from 226 adult volunteers of diverse age groups and body mass index ranges (18-60 years, 16.6-67.7 kg·m-2). Measured REE was compared to preexisting prediction equations via correlation, regression, and Bland-Altman analysis. Then, cohort-specific REE equations were developed using curve estimation and nonlinear regression. To reduce type I error, presently derived equations were validated by splitting the sample into a training and validation group. RESULTS: Preexisting equations over-predicted in-cohort REE. Equations by Livigston and Kohlstadt were most accurate at the individual level (63% accuracy), while formulas by Owen and collaborators elicited highest accuracy at the group level (-1.8% bias). Bland-Altman analysis showed proportional bias for most equations. Currently developed equations showed highest overall accuracy with 70% at the individual and group level (1.0% bias), with small differences between measured and predicted REE values (mean, 95% CI 36 [-15 to 88] kcal·day-1). CONCLUSION: Data indicate currently developed equations to be the most accurate and valid for estimating REE in Greek adults. Further studies should examine the developed equations in an independent sample.

Validation Testing Demonstrates Efficacy of a 7-Day Fluid Record to Estimate Daily Water Intake in Adult Men and Women When Compared with Total Body Water Turnover Measurement.

Background: Mean daily water intake from fluids (WATER-FL) has proven to be difficult to measure because of a range of nonvalidated data collection techniques. Few questionnaires have been validated to estimate WATER-FL against self-reported diaries or urinary hydration markers, which may limit their objectivity.Objectives: The goals of this investigation were 1) to assess the validity of a 7-d fluid record (7dFLR) to measure WATER-FL (WATER-FL-7dFLR) through comparison with WATER-FL as calculated by measuring deuterium oxide (D2O) disappearance (WATER-FL-D2O), and 2) to evaluate the reliability of the 7dFLR in measuring WATER-FL.Methods: Participants [n = 96; 51% female; mean ± SD age: 41 ± 14 y; mean ± SD body mass index (in kg/m2): 26.2 ± 5.1] completed body water turnover analysis over 3 consecutive weeks. They completed the 7dFLR and food diaries during weeks 2 and 4 of the observation. The records were entered into nutritional software to determine the water content of all foods and fluids consumed. WATER-FL-D2O was calculated from water turnover (via the D2O dilution method), minus water from food and metabolic water. The agreement between the 2 methods of determining WATER-FL were compared according to a Bland-Altman plot at week 2. The test-retest reliability of 7dFLR between weeks 2 and 4 was assessed via intraclass correlation (ICC).Results: The mean ± SD difference between WATER-FL-7dFLR and WATER-FL-D2O was -131 ± 845 mL/d. In addition, no bias was observed (F[1,94] = 0.484; R2 = 0.006; P = 0.488). When comparing WATER-FL-7dFLR from weeks 2 and 4, no significant difference (mean ± SD difference: 71 ± 75 mL/d; t[79] = 0.954; P = 0.343) and an ICC of 0.85 (95% CI: 0.77, 0.90) was observed.Conclusions: The main findings of this study were that the use of the 7dFLR is an effective and reliable method to estimate WATER-FL in adults. This style of questionnaire may be extremely helpful for collecting water intake data for large-scale epidemiologic studies.

The Effect of Storing Temperature and Duration on Urinary Hydration Markers.

The purpose of this investigation was to quantify the effects of storage temperature, duration, and the urinary sediment on urinary hydration markers. Thirty-six human urine samples were analyzed fresh and then the remaining sample was separated into 24 separate vials, six in each of the following four temperatures: 22 °C, 7 °C, -20 °C, and -80 °C. Two of each sample stored in any given temperature, were analyzed after 1, 2, and 7 days either following vortexing or centrifugation. Each urine sample was analyzed for osmolality (UOsm), urine specific gravity (USG), and urine color (UC). UOsm was stable at 22 °C, for 1 day (+5-9 mmol∙kg-1, p > .05) and at 7 °C, UOsm up to 7 days (+8-8 mmol∙kg-1, p > .05). At -20 and -80 °C, UOsm decreased after 1, 2, and 7 days (9-61 mmol∙kg-1, p < .05). Vortexing the sample before analysis further decreased only UOsm in the -20 °C and -80 °C storage. USG remained stable up to 7 days when samples were stored in 22 °C or 7 °C (p > .05) but declined significantly when stored in -20 °C, and -80 °C (p < .001). UC was not stable in any of the storing conditions for 1, 2, and 7 days. In conclusion, these data indicate that urine specimens analyzed for UOsm or USG remained stable in refrigerated (7 °C) environment for up to 7 days, and in room temperature for 1 day. However, freezing (-20 and -80 °C) samples significantly decreased the values of hydration markers.

A Mediterranean diet plan in lactating women with obesity reduces maternal energy intake and modulates human milk composition - a feasibility study.

Introduction: Maternal obesity is associated with increased concentrations of human milk (HM) obesogenic hormones, pro-inflammatory cytokines, and oligosaccharides (HMOs) that have been associated with infant growth and adiposity. The objective of this pilot study was to determine if adherence to a Mediterranean meal plan during lactation modulates macronutrients and bioactive molecules in human milk from mothers with obesity. Methods: Sixteen healthy, exclusively breastfeeding women with obesity (body mass index ≥30 kg/m2) enrolled between 4 and 5 months postpartum. The women followed a 4-week Mediterranean meal plan which was provided at no cost. Maternal and infant anthropometrics, HM composition, and infant intakes were measured at enrollment and at weeks 2 and 4 of the intervention. Thirteen mother-infant dyads completed the study. Additionally, participants from an adjacent, observational cohort who had obesity and who collected milk at 5 and 6 months postpartum were compared to this cohort. Results: Participants' healthy eating index scores improved (+27 units, p < 0.001), fat mass index decreased (-4.7%, p < 0.001), and daily energy and fat intake were lower (-423.5 kcal/day, p < 0.001 and-32.7 g/day, p < 0.001, respectively) following the intervention. While HM macronutrient concentrations did not change, HM leptin, total human milk oligosaccharides (HMOs), HMO-bound fucose, Lacto-N-fucopentaose (LNFP)-II, LNFP-III, and difucosyllacto-N-tetrose (DFLNT) concentrations were lower following the intervention. Infant intakes of leptin, tumor necrosis factor (TNF)-α, total HMOs, HMO-bound fucose, LNFP-III and DFLNT were lower following the intervention. Specific components of the maternal diet (protein and fat) and specific measures of maternal diet quality (protein, dairy, greens and beans, fruit and vegetables) were associated with infant intakes and growth. Discussion: Adherence to a Mediterranean meal plan increases dietary quality while reducing total fat and caloric intake. In effect, body composition in women with obesity improved, HM composition and infants' intakes were modulated. These findings provide, for the first time, evidence-based data that enhancing maternal dietary quality during lactation may promote both maternal and child health. Longer intervention studies examining the impact of maternal diet quality on HM composition, infant growth, and infant development are warranted.

Design and conduct of a randomized controlled feeding trial in a residential setting with mitigation for COVID-19.

BACKGROUND: Evaluating effects of different macronutrient diets in randomized trials requires well defined infrastructure and rigorous methods to ensure intervention fidelity and adherence. METHODS: This controlled feeding study comprised two phases. During a Run-in phase (14-15 weeks), study participants (18-50 years, BMI, ≥27 kg/m2) consumed a very-low-carbohydrate (VLC) diet, with home delivery of prepared meals, at an energy level to promote 15 ± 3% weight loss. During a Residential phase (13 weeks), participants resided at a conference center. They received a eucaloric VLC diet for three weeks and then were randomized to isocaloric test diets for 10 weeks: VLC (5% energy from carbohydrate, 77% from fat), high-carbohydrate (HC)-Starch (57%, 25%; including 20% energy from refined grains), or HC-Sugar (57%, 25%; including 20% sugar). Outcomes included measures of body composition and energy expenditure, chronic disease risk factors, and variables pertaining to physiological mechanisms. Six cores provided infrastructure for implementing standardized protocols: Recruitment, Diet and Meal Production, Participant Support, Assessments, Regulatory Affairs and Data Management, and Statistics. The first participants were enrolled in May 2018. Participants residing at the conference center at the start of the COVID-19 pandemic completed the study, with each core implementing mitigation plans. RESULTS: Before early shutdown, 77 participants were randomized, and 70 completed the trial (65% of planned completion). Process measures indicated integrity to protocols for weighing menu items, within narrow tolerance limits, and participant adherence, assessed by direct observation and continuous glucose monitoring. CONCLUSION: Available data will inform future research, albeit with less statistical power than originally planned.

Prolonged Glycemic Adaptation Following Transition From a Low- to High-Carbohydrate Diet: A Randomized Controlled Feeding Trial.

OBJECTIVE: Consuming ≥150 g/day carbohydrate is recommended for 3 days before an oral glucose tolerance test (OGTT) for diabetes diagnosis. For evaluation of this recommendation, time courses of glycemic changes following transition from a very-low-carbohydrate (VLC) to high-carbohydrate diet were assessed with continuous glucose monitoring (CGM). RESEARCH DESIGN AND METHODS: After achieving a weight loss target of 15% (±3%) on the run-in VLC diet, participants (18-50 years old, BMI ≥27 kg/m2) were randomly assigned for 10 weeks to one of three isoenergetic diets: VLC (5% carbohydrate and 77% fat); high carbohydrate, high starch (HC-Starch) (57% carbohydrate and 25% fat, including 20% refined grains); and high carbohydrate, high sugar (HC-Sugar) (57% carbohydrate and 25% fat, including 20% sugar). CGM was done throughout the trial (n = 64) and OGTT at start and end (n = 41). All food was prepared in a metabolic kitchen and consumed under observation. RESULTS: Glucose metrics continued to decline after week 1 in the HC-Starch and HC-Sugar groups (P < 0.05) but not VLC. During weeks 2-5, fasting and 2-h glucose (millimoles per liter per week) decreased in HC-Starch (fasting -0.10, P = 0.001; 2 h -0.10, P = 0.04). During weeks 6-9, 2-h glucose decreased in HC-Starch (-0.07, P = 0.01) and fasting and 2-h glucose decreased in HC-Sugar (fasting -0.09, P = 0.001; 2 h -0.09, P = 0.003). The number of participants with abnormal glucose tolerance by OGTT remained 10 (of 16) in VLC at start and end but decreased from 17 to 9 (of 25) in both high-carbohydrate groups. CONCLUSIONS: Physiological adaptation from a low- to high-carbohydrate diet may require many weeks, with implications for the accuracy of diabetes tests, interpretation of macronutrient trials, and risks of periodic planned deviations from a VLC diet.

Development of metabolic and contractile alterations in development of cancer cachexia in female tumor-bearing mice.

Cancer cachexia (CC) results in impaired muscle function and quality of life and is the primary cause of death for ∼20%-30% of patients with cancer. We demonstrated mitochondrial degeneration as a precursor to CC in male mice; however, whether such alterations occur in females is currently unknown. The purpose of this study was to elucidate muscle alterations in CC development in female tumor-bearing mice. Sixty female C57BL/6J mice were injected with PBS or Lewis lung carcinoma at 8 wk of age, and tumors developed for 1, 2, 3, or 4 wk to assess the time course of cachectic development. In vivo muscle contractile function, protein fractional synthetic rate (FSR), protein turnover, and mitochondrial health were assessed. Three- and four-week tumor-bearing mice displayed a dichotomy in tumor growth and were reassigned to high tumor (HT) and low tumor (LT) groups. HT mice exhibited lower soleus, tibialis anterior, and fat weights than PBS mice. HT mice showed lower peak isometric torque and slower one-half relaxation time than PBS mice. HT mice had lower FSR than PBS mice, whereas E3 ubiquitin ligases were greater in HT than in other groups. Bnip3 (mitophagy) and pMitoTimer red puncta (mitochondrial degeneration) were greater in HT mice, whereas Pgc1α1 and Tfam (mitochondrial biogenesis) were lower in HT mice than in PBS mice. We demonstrate alterations in female tumor-bearing mice where HT exhibited greater protein degradation, impaired muscle contractility, and mitochondrial degeneration compared with other groups. Our data provide novel evidence for a distinct cachectic development in tumor-bearing female mice compared with previous male studies.NEW & NOTEWORTHY Our study demonstrates divergent tumor development and tissue wasting within 3- and 4-wk mice, where approximately half the mice developed large tumors and subsequent cachexia. Unlike previous male studies, where metabolic perturbations precede the onset of cachexia, females appear to exhibit protections from the metabolic perturbations and cachexia development. Our data provide novel evidence for divergent cachectic development in tumor-bearing female mice compared with previous male CC studies, suggesting different mechanisms of CC between sexes.

Combining urine color and void number to assess hydration in adults and children.

BACKGROUND/OBJECTIVES: To test the diagnostic ability of two combined practical markers for elevated urine osmolality (underhydration) in free-living adults and children. SUBJECTS/METHODS: One hundred and one healthy adults (females n = 52, 40 ± 14 y, 1.70 ± 0.95 m, 76.7 ± 17.4 kg, 26.5 ± 5.5 kg/m2) and 210 children (females = 105, 1.49 ± 0.13 m, 43.4 ± 12.6 kg, 19.2 ± 3.2 kg m-2) collected urine for 24-h. Urine was analyzed for urine osmolality (UOsm), color (UC), while the number of voids (void) was also recorded. Receiver Operating Characteristic (ROC) analysis was performed for UC, void, and combination of UC and void, to determine markers' diagnostic ability for detecting underhydration based on elevated UOsm (UOsm ≥ 800 mmol kg-1). RESULTS: Linear regression analysis revealed that UC was significantly associated with UOsm in both adults (R2 = 0.38; P < 0.001) and children (R2 = 0.45; P < 0.001). Void was significantly associated with UOsm in both adults (R2 = 0.13; P < 0.001) and children (R2 = 0.15; P < 0.001). In adults, when UC > 3 and void <7 were combined, the overall diagnostic ability for underhydration was 97% with sensitivity and specificity of 100% and 88%, respectively. In children, UC > 3 and void <5 had an overall diagnostic ability for underhydration of 89% with sensitivity and specificity of 100% and 62%, respectively. CONCLUSIONS: Urine color alone and the combination of urine color with void number can a valid and simple field-measure to detect underhydration based on elevated urine osmolality.

Mitochondrial aberrations during the progression of disuse atrophy differentially affect male and female mice.

BACKGROUND: Disuse decreases muscle size and is predictive of mortality across multiple pathologies. Detriments to mitochondrial function are hypothesized to underlie disuse-induced muscle atrophy. Little data exist on early mechanisms contributing to onset of these pathologies, nor is it known how they differ between sexes. The purpose of this study was to examine differential and conserved responses to mitochondrial quality control in male and female mice during the development and progression of disuse-induced atrophy. METHODS: One hundred C57BL/6J mice (50 male and 50 female) were hindlimb unloaded to induce disuse atrophy for 0 (con), 24, 48, 72, or 168 h. At designated time-points, extensor digitorum longus, gastrocnemius, and soleus muscles were collected for analysis of mitochondrial quality control markers. RESULTS: One hundred sixty-eight hours of disuse resulted in ~25% lower oxidative muscle fibre CSA in both male (P = 0.003) and female (P = 0.02) mice without any differences due to disuse in glycolytic fibres. In male mice, 48 h of unloading was sufficient to result in ~67% greater mitochondrial oxidative stress as assessed by the reporter gene pMitoTimer compared with 0 h (P = 0.002), this mitochondrial stress preceded detectable muscle loss. However in female mice, mitochondrial oxidative stress did not occur until 168 h of disuse (~40% greater mitochondrial oxidative stress in 168 h compared with 0 h of disuse, P < 0.0001). Blunted oxidative stress in female mice appeared to coincide with greater inductions of autophagy and mitophagy in female mice (~3-fold greater BNIP3 and ~6-fold greater LC3II/I ratio P < 0.0001 and P = 0.038 respectively). Male mice overall had greater reactive oxygen species (ROS) production compared with female mice. Female mice had a greater induction of ROS within 24 h of disuse (~4-fold greater compared with 0 h, P < 0.0001); whereas male mice did not have greater ROS production until 168 h of disuse (~2-fold greater, P < 0.0001). Although all muscle types exhibited some alterations to mitochondrial quality control, such as increased markers of mitophagy and fission, the soleus muscle in both male and female mice exhibited consistent alterations to various markers of mitochondrial quality. Markers of mitochondrial translation were approximately 30-50% lower within 24 h of unloading in both male and female soleus muscle (P value ranges: <0.0001-0.03). CONCLUSIONS: Disuse negatively affects mitochondria differentially between sexes during development of muscle wasting. Acutely, female mice may forgo muscle mass to maintain mitochondrial quality compared with male mice. These differences may contribute to divergent clinical manifestations of atrophy.

Validity and Reliability of a Water Frequency Questionnaire to Estimate Daily Total Water Intake in Adults.

The purpose of this investigation was to assess the validity and reliability of a seven-day water frequency questionnaire (TWI-FQ) to estimate daily total water intake (TWI) in comparison to a water turnover objective reference value via deuterium oxide (D2O). Data collection occurred over 3 weeks, with a wash-out period during week two. Healthy adults (n = 98; 52% female; 41 ± 14 y; BMI, 26.4 ± 5.5 kg·m-2) retrospectively self-reported consumption frequencies of 17 liquids and 35 foods with specified volumes/amounts for weeks one and three via TWI-FQ. Standard water content values were utilized to determine the volume of water consumed from each liquid and food for calculation of mean daily TWI for each week. Diet records were completed daily during week two to estimate metabolic water production. To assess validity of the TWI-FQ, participants consumed D2O at the start of each week and provided urine samples immediately before ingestion, the following day, and at the end of the week to calculate water turnover. Metabolic water was subtracted from water turnover to estimate TWI. TWI-FQ validity was assessed via Bland-Altman plot for multiple observations. Reliability was assessed via intraclass correlation and Pearson's correlation between weeks. TWI-FQ significantly underestimated D2O TWI by -350 ± 1,431 mL·d-1 (95% confidence interval (CI): -551, -149 mL·d-1). TWI-FQ TWI was significantly correlated (r = 0.707, P <0.01) and not different (198 ± 1,180 mL·d-1, 95% CI: -38, 435 mL·d-1) between weeks. TWI-FQ intraclass correlation = 0.706 was significant [95% CI: 0.591, 0.793; F (97, 98) = 5.799], indicating moderate test-retest reliability. While this tool would not be suitable for individual TWI assessment, the magnitude of bias may be acceptable for assessment at the sample-level.

Female mice may have exacerbated catabolic signalling response compared to male mice during development and progression of disuse atrophy.

BACKGROUND: Muscle atrophy is a common pathology associated with disuse, such as prolonged bed rest or spaceflight, and is associated with detrimental health outcomes. There is emerging evidence that disuse atrophy may differentially affect males and females. Cellular mechanisms contributing to the development and progression of disuse remain elusive, particularly protein turnover cascades. The purpose of this study was to investigate the initial development and progression of disuse muscle atrophy in male and female mice using the well-established model of hindlimb unloading (HU). METHODS: One hundred C57BL/6J mice (50 male and 50 female) were hindlimb suspended for 0 (control), 24, 48, 72, or 168 h to induce disuse atrophy (10 animals per group). At designated time points, animals were euthanized, and tissues (extensor digitorum longus, gastrocnemius, and soleus for mRNA analysis, gastrocnemius and extensor digitorum longus for protein synthesis rates, and tibialis anterior for histology) were collected for analysis of protein turnover mechanisms (protein anabolism and catabolism). RESULTS: Both males and females lost ~30% of tibialis anterior cross-sectional area after 168 h of disuse. Males had no statistical difference in MHCIIB fibre area, whereas unloaded females had ~33% lower MHCIIB cross-sectional area by 168 h of unloading. Both males and females had lower fractional protein synthesis rates (FSRs) within 24-48 h of HU, and females appeared to have a greater reduction compared with males within 24 h of HU (~23% lower FSRs in males vs. 40% lower FSRs in females). Males and females exhibited differential patterns and responses in multiple markers of protein anabolism, catabolism, and myogenic capacity during the development and progression of disuse atrophy. Specifically, females had greater mRNA inductions of catabolic factors Ubc and Gadd45a (~4-fold greater content in females compared with ~2-fold greater content in males) and greater inductions of anabolic inhibitors Redd1 and Deptor with disuse across multiple muscle tissues exhibiting different fibre phenotypes. CONCLUSIONS: These results suggest that the aetiology of disuse muscle atrophy is more complicated and nuanced than previously thought, with different responses based on muscle phenotypes and between males and females, with females having greater inductions of atrophic markers early in the development of disuse atrophy.

Neither autophagy nor exercise training mode affect exercise-induced beneficial adaptations in high fat-fed mice.

Exercise mitigates obesity-associated pathologies; however, there is controversy regarding optimal exercise interventions. Autophagy, is known to decrease during obesity and is an important moderator for exercise adaptations. Purpose: To investigate individual and combined effects of different exercise interventions and autophagy inhibition on exercise adaptations during obesity. Methods: C57BL/6J mice initiated 45% high fat diet at 8 weeks of age. After 6 weeks of diet, animals were divided into moderate (MOD) or high intensity interval training interventions (HIIT), animals were further divided into autophagy inhibition or vehicle conditions (n = 10/group). Animals exercised and autophagy was inhibited 3X/week by NSC185058 injections, thereby blocking autophagosome formation. Interventions continued for 4 weeks. Results: High fat diet impaired glucose handling ∼17%; exercise interventions normalized glucoregulation to pre-high fat diet levels, without differences between any interventions. High fat diet induced ∼25% decrease in aerobic capacity, which returned to baseline after exercise interventions, with no differences between any interventions. No effects of autophagy inhibition were noted. Conclusions: HIIT and MOD training confer similar health-related adaptations.