Myosin-binding protein H-like regulates myosin-binding protein distribution and function in atrial cardiomyocytes.

Mutations in atrial-enriched genes can cause a primary atrial myopathy that can contribute to overall cardiovascular dysfunction. MYBPHL encodes myosin-binding protein H-like (MyBP-HL), an atrial sarcomere protein that shares domain homology with the carboxy-terminus of cardiac myosin-binding protein-C (cMyBP-C). The function of MyBP-HL and the relationship between MyBP-HL and cMyBP-C is unknown. To decipher the roles of MyBP-HL, we used structured illumination microscopy, immuno-electron microscopy, and mass spectrometry to establish the localization and stoichiometry of MyBP-HL. We found levels of cMyBP-C, a major regulator of myosin function, were half as abundant compared to levels in the ventricle. In genetic mouse models, loss of MyBP-HL doubled cMyBP-C abundance in the atria, and loss of cMyBP-C doubled MyBP-HL abundance in the atria. Structured illumination microscopy showed that both proteins colocalize in the C-zone of the A-band, with MyBP-HL enriched closer to the M-line. Immuno-electron microscopy of mouse atria showed MyBP-HL strongly localized 161 nm from the M-line, consistent with localization to the third 43 nm repeat of myosin heads. Both cMyBP-C and MyBP-HL had less-defined sarcomere localization in the atria compared to ventricle, yet areas with the expected 43 nm repeat distance were observed for both proteins. Isometric force measurements taken from control and Mybphl null single atrial myofibrils revealed that loss of Mybphl accelerated the linear phase of relaxation. These findings support a mechanism where MyBP-HL regulates cMyBP-C abundance to alter the kinetics of sarcomere relaxation in atrial sarcomeres.

Heat tolerance, oxidative stress response tuning and robust gene activation in early-stage Drosophila melanogaster embryos.

In organisms with complex life cycles, life stages that are most susceptible to environmental stress may determine species persistence in the face of climate change. Early embryos of Drosophila melanogaster are particularly sensitive to acute heat stress, yet tropical embryos have higher heat tolerance than temperate embryos, suggesting adaptive variation in embryonic heat tolerance. We compared transcriptomic responses to heat stress among tropical and temperate embryos to elucidate the gene regulatory basis of divergence in embryonic heat tolerance. The transcriptomes of tropical and temperate embryos differed in both constitutive and heat-stress-induced responses of the expression of relatively few genes, including genes involved in oxidative stress. Most of the transcriptomic response to heat stress was shared among all embryos. Embryos shifted the expression of thousands of genes, including increases in the expression of heat shock genes, suggesting robust zygotic gene activation and demonstrating that, contrary to previous reports, early embryos are not transcriptionally silent. The involvement of oxidative stress genes corroborates recent reports on the critical role of redox homeostasis in coordinating developmental transitions. By characterizing adaptive variation in the transcriptomic basis of embryonic heat tolerance, this study is a novel contribution to the literature on developmental physiology and developmental genetics.

Sex differences in iron status during military training: a prospective cohort study of longitudinal changes and associations with endurance performance and musculoskeletal outcomes.

This study investigated sex differences in Fe status, and associations between Fe status and endurance and musculoskeletal outcomes, in military training. In total, 2277 British Army trainees (581 women) participated. Fe markers and endurance performance (2·4 km run) were measured at the start (week 1) and end (week 13) of training. Whole-body areal body mineral density (aBMD) and markers of bone metabolism were measured at week 1. Injuries during training were recorded. Training decreased Hb in men and women (mean change (-0·1 (95 % CI -0·2, -0·0) and -0·7 (95 % CI -0·9, -0·6) g/dl, both P < 0·001) but more so in women (P < 0·001). Ferritin decreased in men and women (-27 (95 % CI -28, -23) and -5 (95 % CI -8, -1) µg/l, both P ≤ 0·001) but more so in men (P < 0·001). Soluble transferrin receptor increased in men and women (2·9 (95 % CI 2·3, 3·6) and 3·8 (95 % CI 2·7, 4·9) nmol/l, both P < 0·001), with no difference between sexes (P = 0·872). Erythrocyte distribution width increased in men (0·3 (95 % CI 0·2, 0·4)%, P < 0·001) but not in women (0·1 (95 % CI -0·1, 0·2)%, P = 0·956). Mean corpuscular volume decreased in men (-1·5 (95 % CI -1·8, -1·1) fL, P < 0·001) but not in women (0·4 (95 % CI -0·4, 1·3) fL, P = 0·087). Lower ferritin was associated with slower 2·4 km run time (P = 0·018), sustaining a lower limb overuse injury (P = 0·048), lower aBMD (P = 0·021) and higher beta C-telopeptide cross-links of type 1 collagen and procollagen type 1 N-terminal propeptide (both P < 0·001) controlling for sex. Improving Fe stores before training may protect Hb in women and improve endurance and protect against injury.

Cardiac Myosin Filaments are Maintained by Stochastic Protein Replacement.

Myosin and myosin-binding protein C are exquisitely organized into giant filamentous macromolecular complexes within cardiac muscle sarcomeres, yet these proteins must be continually replaced to maintain contractile fidelity. The overall hypothesis that myosin filament structure is dynamic and allows for the stochastic replacement of individual components was tested in vivo, using a combination of mass spectrometry- and fluorescence-based proteomic techniques. Adult mice were fed a diet that marked all newly synthesized proteins with a stable isotope-labeled amino acid. The abundance of unlabeled and labeled proteins was quantified by high-resolution mass spectrometry over an 8-week period. The rates of change in the abundance of these proteins were well described by analytical models in which protein synthesis defined stoichiometry and protein degradation was governed by the stochastic selection of individual molecules. To test whether the whole myosin filaments or the individual components were selected for replacement, cardiac muscle was chemically skinned to remove the cellular membrane and myosin filaments were solubilized with ionic solutions. The composition of the filamentous and soluble fractions was quantified by mass spectrometry, and filament depolymerization was visualized by real-time fluorescence microscopy. Myosin molecules were preferentially extracted from ends of the filaments in the presence of the ionic solutions, and there was only a slight bias in the abundance of unlabeled molecules toward the innermost region on the myosin filaments. These data demonstrate for the first time that the newly synthesized myosin and myosin-binding protein C molecules are randomly mixed into preexisting thick filaments in vivo and the rate of mixing may not be equivalent along the length of the thick filament. These data collectively support a new model of cardiac myosin filament structure, with the filaments being dynamic macromolecular assemblies that allow for replacement of their components, rather than rigid bodies.

Distal Tibial Bone Properties and Bone Stress Injury Risk in Young Men Undergoing Arduous Physical Training.

Trabecular microarchitecture contributes to bone strength, but its role in bone stress injury (BSI) risk in young healthy adults is unclear. Tibial volumetric BMD (vBMD), geometry, and microarchitecture, whole-body areal BMD, lean and fat mass, biochemical markers of bone metabolism, aerobic fitness, and muscle strength and power were measured in 201 British Army male infantry recruits (age 20.7 [4.3] years, BMI 24.0 ± 2.7 kg·m2) in week one of basic training. Tibial scans were performed at the ultra-distal site, 22.5 mm from the distal endplate of the non-dominant leg using High Resolution Peripheral Quantitative Computed Tomography (XtremeCT, Scanco Medical AG, Switzerland). Binary logistic regression analysis was performed to identify associations with lower body BSI confirmed by MRI. 20 recruits (10.0%) were diagnosed with a lower body BSI. Pre-injured participants had lower cortical area, stiffness and estimated failure load (p = 0.029, 0.012 and 0.011 respectively) but tibial vBMD, geometry, and microarchitecture were not associated with BSI incidence when controlling for age, total body mass, lean body mass, height, total 25(OH)D, 2.4-km run time, peak power output and maximum dynamic lift strength. Infantry Regiment (OR 9.3 [95%CI, 2.6, 33.4]) Parachute versus Line Infantry, (p ≤ 0.001) and 2.4-km best effort run time (1.06 [95%CI, 1.02, 1.10], p < 0.033) were significant predictors. Intrinsic risk factors, including ultradistal tibial density, geometry, and microarchitecture, were not associated with lower body BSI during arduous infantry training. The ninefold increased risk of BSI in the Parachute Regiment compared with Line Infantry suggests that injury propensity is primarily a function of training load and risk factors are population-specific.

The effect of calcium supplementation on calcium and bone metabolism during load carriage in women: protocol for a randomised controlled crossover trial.

BACKGROUND: Military field exercises are characterised by high volumes of exercise and prolonged periods of load carriage. Exercise can decrease circulating serum calcium and increase parathyroid hormone and bone resorption. These disturbances to calcium and bone metabolism can be attenuated with calcium supplementation immediately before exercise. This randomised crossover trial will investigate the effect of calcium supplementation on calcium and bone metabolism, and bone mineral balance, during load carriage exercise in women. METHODS: Thirty women (eumenorrheic or using the combined oral contraceptive pill, intrauterine system, or intrauterine device) will complete two experimental testing sessions either with, or without, a calcium supplement (1000 mg). Each experimental testing session will involve one 120 min session of load carriage exercise carrying 20 kg. Venous blood samples will be taken and analysed for biochemical markers of bone resorption and formation, calcium metabolism, and endocrine function. Urine will be collected pre- and post-load carriage to measure calcium isotopes for the calculation of bone calcium balance. DISCUSSION: The results from this study will help identify whether supplementing women with calcium during load carriage is protective of bone and calcium homeostasis. TRIAL REGISTRATION: NCT04823156 (clinicaltrials.gov).

Resistance exercise differentially alters extracellular vesicle size and subpopulation characteristics in healthy men and women: an observational cohort study.

Extracellular vesicles (EVs) are established mediators of adaptation to exercise. Currently, there are no published data comparing changes in EVs between men and women after resistance exercise. We tested the hypothesis that EV profiles would demonstrate a sex-specific signature following resistance exercise. Ten men and 10 women completed an acute heavy resistance exercise test for back squats using 75% of their one-repetition maximum. Blood was drawn before and immediately after exercise. EVs were isolated from plasma using size exclusion chromatography and stained with antibodies associated with exosomes (CD63), microvesicles (VAMP3), apoptotic bodies (THSD1), and a marker for skeletal muscle EVs (SGCA). CD63+ EV concentration and proportion of total EVs increased 23% (P = 0.006) and 113% (P = 0.005) in both sexes. EV mean size declined in men (P = 0.020), but not in women, suggesting a relative increase in small EVs in men. VAMP3+ EV concentration and proportion of total EVs increased by 93% (P = 0.025) and 61% (P = 0.030) in men and women, respectively. SGCA+ EV concentration was 69% higher in women compared with men independent of time (P = 0.007). Differences were also observed for CD63, VAMP3, and SGCA median fluorescence intensity, suggesting altered surface protein density according to sex and time. There were no significant effects of time or sex on THSD1+ EVs or fluorescence intensity. EV profiles, particularly among exosome-associated and muscle-derived EVs, exhibit sex-specific differences in response to resistance exercise which should be further studied to understand their relationship to training adaptations.

Skeletal MyBP-C isoforms tune the molecular contractility of divergent skeletal muscle systems.

Skeletal muscle myosin-binding protein C (MyBP-C) is a myosin thick filament-associated protein, localized through its C terminus to distinct regions (C-zones) of the sarcomere. MyBP-C modulates muscle contractility, presumably through its N terminus extending from the thick filament and interacting with either the myosin head region and/or the actin thin filament. Two isoforms of MyBP-C (fast- and slow-type) are expressed in a muscle type-specific manner. Are the expression, localization, and Ca2+-dependent modulatory capacities of these isoforms different in fast-twitch extensor digitorum longus (EDL) and slow-twitch soleus (SOL) muscles derived from Sprague-Dawley rats? By mass spectrometry, 4 MyBP-C isoforms (1 fast-type MyBP-C and 3 N-terminally spliced slow-type MyBP-C) were expressed in EDL, but only the 3 slow-type MyBP-C isoforms in SOL. Using EDL and SOL native thick filaments in which the MyBP-C stoichiometry and localization are preserved, native thin filament sliding over these thick filaments showed that, only in the C-zone, MyBP-C Ca2+ sensitizes the thin filament and slows thin filament velocity. These modulatory properties depended on MyBP-C's N terminus as N-terminal proteolysis attenuated MyBP-C's functional capacities. To determine each MyBP-C isoform's contribution to thin filament Ca2+ sensitization and slowing in the C-zone, we used a combination of in vitro motility assays using expressed recombinant N-terminal fragments and in silico mechanistic modeling. Our results suggest that each skeletal MyBP-C isoform's N terminus is functionally distinct and has modulatory capacities that depend on the muscle type in which they are expressed, providing the potential for molecular tuning of skeletal muscle performance through differential MyBP-C expression.

Reproductive and metabolic adaptation to multistressor training in women.

Hypothalamic-pituitary-gonadal (HPG) axis suppression in exercising women can be caused by low energy availability (EA), but the impact of a real-world, multistressor training environment on reproductive and metabolic function is unknown. This study aimed to characterize reproductive and metabolic adaptation in women undertaking basic military training. A prospective cohort study in women undertaking 11-month initial military training (n = 47) was carried out. Dynamic low-dose 1-h gonadotrophin-releasing hormone (GnRH) tests were completed after 0 and 7 mo of training. Urine progesterone was sampled weekly throughout. Body composition (dual X-ray absorptiometry), fasting insulin resistance (homeostatic modeling assessment 2, HOMA2), leptin, sex steroids, anti-Müllerian hormone (AMH), and inhibin B were measured after 0, 7, and 11 mo with an additional assessment of body composition at 3 mo. Luteinizing hormone (LH) and follicle-stimulating hormone (FSH) responses were suppressed after 7 mo (both P < 0.001). Among noncontraceptive users (n = 20), 65% had regular (23-35 days) cycles preenrollment, falling to 24% by 7 mo of training. Of women in whom urine progesterone was measured (n = 24), 87% of cycles showed no evidence of ovulation. There was little change in AMH, LH, and estradiol, although inhibin B and FSH increased (P < 0.05). Fat mass fluctuated during training but at month 11 was unchanged from baseline. Fat-free mass did not change. Visceral adiposity, HOMA2, and leptin increased (all P < 0.001). HPG axis suppression with anovulation occurred in response to training without evidence of low EA. Increased insulin resistance may have contributed to the observed pituitary and ovarian dysfunction. Our findings are likely to represent an adaptive response of reproductive function to the multistressor nature of military training.NEW & NOTEWORTHY We characterized reproductive endocrine adaptation to prolonged arduous multistressor training in women. We identified marked suppression of hypothalamic-pituitary-gonadal (HPG) axis function during training but found no evidence of low energy availability despite high energy requirements. Our findings suggest a complex interplay of psychological and environmental stressors with suppression of the HPG axis via activation of the hypothalamic-pituitary adrenal (HPA) axis. The neuroendocrine impact of nonexercise stressors on the HPG axis during arduous training should be considered.

Biomechanical Basis of Predicting and Preventing Lower Limb Stress Fractures During Arduous Training.

PURPOSE OF REVIEW: Stress fractures at weight-bearing sites, particularly the tibia, are common in military recruits and athletes. This review presents recent findings from human imaging and biomechanics studies aimed at predicting and preventing stress fractures. RECENT FINDINGS: Peripheral quantitative computed tomography (pQCT) provides evidence that cortical bone geometry (tibial width and area) is associated with tibial stress fracture risk during weight-bearing exercise. The contribution of bone trabecular microarchitecture, cortical porosity, and bone material properties in the pathophysiology of stress fractures is less clear, but high-resolution pQCT and new techniques such as impact microindentation may improve our understanding of the role of microarchitecture and material properties in stress fracture prediction. Military studies demonstrate osteogenic outcomes from high impact, repetitive tibial loading during training. Kinetic and kinematic characteristics may influence stress fracture risk, but there is no evidence that interventions to modify biomechanics can reduce the incidence of stress fracture. Strategies to promote adaptive bone formation, in combination with improved techniques to assess bone strength, present exciting opportunities for future research to prevent stress fractures.

Conditions to optimise the developmental competence of immature equine oocytes.

Optimising the developmental potential of immature equine oocytes and invitro-produced (IVP) embryos was explored through modifications of established media and holding temperature. In Experiment 1, delaying spontaneous resumption of meiosis through the process of simulated physiological oocyte maturation with the addition of the adenylate cyclase activator forskolin (50µM) and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (100µM) to overnight holding medium before maturation improved blastocyst production (P<0.05). In Experiment 2, the blastocyst production rate was increased significantly when cumulin (100ng mL-1) was added to the overnight holding or culture media (P<0.05). In Experiment 3, immature oocytes held overnight at 16°C before maturation had improved developmental competence than those held at 20°C and 5°C (P<0.05). There was no difference between maturation rates, but blastocyst formation per cleaved oocyte was significantly greater in oocytes held overnight at 16°C than at 20°C or 5°C. Furthermore, blastocyst formation per recovered oocyte and per fertilised oocyte was greater when oocytes were held before maturation at 16°C than at 5°C (P<0.05). In Experiment 4, the addition of sodium ascorbate (AC; 50µg mL-1) to the maturation and/or culture media of oocytes and IVP embryos did not improve blastocyst production, but did appear to lower cleavage rates compared with oocytes and embryos cultured without AC.

MYBPC3 truncation mutations enhance actomyosin contractile mechanics in human hypertrophic cardiomyopathy.

RATIONALE: Truncation mutations in the MYBPC3 gene, encoding for cardiac myosin-binding protein C (MyBP-C), are the leading cause of hypertrophic cardiomyopathy (HCM). Whole heart, fiber and molecular studies demonstrate that MyBP-C is a potent modulator of cardiac contractility, but how these mutations contribute to HCM is unresolved. OBJECTIVES: To readdress whether MYBPC3 truncation mutations result in loss of MyBP-C content and/or the expression of truncated MyBP-C from the mutant allele and determine how these mutations effect myofilament sliding in human myocardium. METHODS AND RESULTS: Septal wall tissue samples were obtained from HCM patients undergoing myectomy (n = 18) and donor controls (n = 8). The HCM samples contained 40% less MyBP-C and reduced levels of MyBP-C phosphorylation, when compared to the donor control samples using quantitative mass spectrometry. These differences occurred in the absence of changes in the stoichiometry of other myofilament proteins or production of truncated MyBP-C from the mutant MYBPC3 allele. The functional impact of MYBPC3 truncation mutations on myofilament sliding was determined using a total internal reflection microscopy (TIRFM) single particle assay. Myosin-thick filaments containing their native complement of MyBP-C, and actin-thin filaments decorated with the troponin/tropomyosin calcium regulatory proteins, were isolated from a subgroup of the HCM (n = 4) and donor (n = 5) heart samples. The maximal sliding velocity of native thin filaments was enhanced within the C-zones of the native thick filaments isolated from the HCM samples, when compared to velocity within the C-zones of thick filaments isolated from the donor samples. Analytical modeling demonstrated that the 40% reduction in MyBP-C content was sufficient to enhance the myofilament sliding velocity, as observed in the TIRFM assay. CONCLUSIONS: HCM-causing MYBPC3 truncation mutations result in a loss of MyBP-C content that enhances maximal myofilament sliding velocities, only where MyBP-C is localized within the C-zone. These findings support therapeutic rationale for restoring normal levels of MyBP-C and/or dampening maximal contractile velocities for the treatment of human HCM.

High-intensity exhaustive exercise reduces long-interval intracortical inhibition.

The development of fatigue during single-joint isolated muscle contractions is accompanied by an increase in long-interval intracortical inhibition (LICI). However, the effect of whole-body locomotor endurance exercise on LICI is unknown. Eighteen healthy men completed three exercise trials on a cycle ergometer. The first trial was completed to determine the lactate threshold (LT) and maximal oxygen uptake ([Formula: see text]). The remaining two trials (familiarisation and experimental) involved cycling to volitional exhaustion at an intensity equivalent to halfway between the LT and [Formula: see text] (50%Δ). Responses to stimulation of the femoral nerve [motor nerve stimulation (MNS)] and motor cortex [transcranial magnetic stimulation (TMS)] were determined pre- and post-exercise to determine the level of peripheral fatigue [potentiated quadriceps twitch (Qtw,pot)] and central fatigue [voluntary activation measured by MNS and TMS (VAMNS and VATMS, respectively)]. Corticospinal excitability (motor evoked potentials) and intracortical inhibition [LICI and corticospinal silent period (SP)] were also measured from electromyography recordings on the vastus lateralis. There were exercise-induced reductions in maximal voluntary contraction torque (- 21 ± 10%), Qtw,pot (- 37 ± 18%), VAMNS (- 7 ± 7%) and VATMS (- 8 ± 10) (all P < 0.01). There were increases in the LICI ratio and reductions in SP duration from pre- to post-exercise (mean absolute change of 16 ± 14% and - 31 ± 28 s, respectively) (both P < 0.01). The pre- and post-exercise MEP amplitudes were not different (P = 0.86). The neural inhibitory circuits that mediate the LICI and SP became less excitable with fatigue following high-intensity exhaustive cycling, which could be important in the aetiology of central fatigue during whole-body locomotor endurance exercise.

High but not moderate-intensity endurance training increases pain tolerance: a randomised trial.

PURPOSE: To examine the effect of high-intensity interval training (HIIT) compared to volume-matched moderate-intensity continuous training (CONT) on muscle pain tolerance and high-intensity exercise tolerance. METHODS: Twenty healthy adults were randomly assigned (1:1) to either 6 weeks of HIIT [6-8 × 5 min at halfway between lactate threshold and maximal oxygen uptake (50%Δ)] or volume-matched CONT (~60-80 min at 90% lactate threshold) on a cycle ergometer. A tourniquet test to examine muscle pain tolerance and two time to exhaustion (TTE) trials at 50%Δ to examine exercise tolerance were completed pre- and post-training; the post-training TTE trials were completed at the pre-training 50%Δ (same absolute-intensity) and the post-training 50%Δ (same relative-intensity). RESULTS: HIIT and CONT resulted in similar improvements in markers of aerobic fitness (all P ≥ 0.081). HIIT increased TTE at the same absolute- and relative-intensity as pre-training (148 and 43%, respectively) to a greater extent than CONT (38 and -4%, respectively) (both P ≤ 0.019). HIIT increased pain tolerance (41%, P < 0.001), whereas CONT had no effect (-3%, P = 0.720). Changes in pain tolerance demonstrated positive relationships with changes in TTE at the same absolute- (r = 0.44, P = 0.027) and relative-intensity (r = 0.51, P = 0.011) as pre-training. CONCLUSION: The repeated exposure to a high-intensity training stimulus increases muscle pain tolerance, which is independent of the improvements in aerobic fitness induced by endurance training, and may contribute to the increase in high-intensity exercise tolerance following HIIT.

Reliability of single and paired-pulse transcranial magnetic stimulation in the vastus lateralis muscle.

INTRODUCTION: Transcranial magnetic stimulation (TMS) is an important tool to examine neurological pathologies, movement disorders, and central nervous system responses to exercise, fatigue, and training. The reliability has not been examined in a functional locomotor knee extensor muscle. METHODS: Within- (n = 10) and between-day (n = 16) reliability of single and paired-paired pulse TMS was examined from the active vastus lateralis. RESULTS: Motor evoked potential amplitude and cortical silent period duration showed good within- and between-day reliability (intraclass correlation coefficient [ICC] ≥ 0.82). Short- and long-interval intracortical inhibition (SICI and LICI, respectively) demonstrated good within-day reliability (ICC ≥ 0.84). SICI had moderate to good between-day reliability (ICC ≥ 0.67), but LICI was not repeatable (ICC = 0.47). Intracortical facilitation showed moderate to good within-day reliability (ICC ≥ 0.73) but poor to moderate reliability between days (ICC ≥ 0.51). CONCLUSIONS: TMS can reliably assess cortical function in a knee extensor muscle. This may be useful to examine neurological disorders that affect locomotion.

Treatment of human embryos with the TGFß inhibitor SB431542 increases epiblast proliferation and permits successful human embryonic stem cell derivation.

STUDY QUESTION: Is there an effect of the TGFß inhibitor SB431542 (SB) on the epiblast compartment of human blastocysts, and does it affect subsequent human embryonic stem cell (hESC) derivation? SUMMARY ANSWER: SB increases the mean number of NANOG-positive cells in the inner cell mass (ICM), and allows for subsequent hESC derivation. WHAT IS KNOWN ALREADY: It is known that inhibition of TGFß by SB has a positive effect on mouse ESC self-renewal, while active TGFß signalling is needed for self-renewal of primed ESC. STUDY DESIGN, SIZE, DURATION: From December 2011 until March 2012, 263 donated spare embryos were used from patients who had undergone IVF/ICSI in our centre. PARTICIPANTS/MATERIALS, SETTING, METHODS: Donated human embryos were cultured in the presence of SB or Activin A, and immunocytochemistry was performed on Day 6 blastocysts for NANOG and GATA6. Moreover, blastocysts were used for the derivation of hESC, with or without exposure to SB. MAIN RESULTS AND THE ROLE OF CHANCE: Immunocytochemistry revealed a significantly higher number of NANOG-positive ICM cells in the SB group compared with the control (12.0 ± 5.9 versus 6.1 ± 4.7), while no difference was observed in the Activin A group compared with other groups (6.7 ± 3.7). The number of GATA6-positive ICM cells did not differ between the SB, Activin A and control group (8.8 ± 4.3, 8.0 ± 4.6 and 7.2 ± 4.0, respectively). Blocking TGFß signalling did not prevent subsequent hESC line derivation. LIMITATIONS, REASONS FOR CAUTION: The number of human blastocysts available for this study was too low to reveal if the observed increase in NANOG-positive epiblast cells after exposure to SB affected the efficiency of hESC derivation (12.5% compared with 16.7%). WIDER IMPLICATIONS OF THE FINDINGS: This work can contribute to the derivation of naive hESC lines in the future. STUDY FUNDING/COMPETING INTEREST(S): M.V.d.J. is holder of a Ph.D. grant of the Agency for Innovation by Science and Technology (IWT, grant number SB093128), Belgium. G.D. and this research are supported by the Research Foundation Flanders (FWO), grant number FWO-3G062910) and a Concerted Research Actions funding from BOF (Bijzonder Onderzoeksfonds University Ghent, grant number BOF GOA 01G01112). S.M.C.d. S.L. is supported by the Netherlands Organization of Scientific Research (NWO) (ASPASIA 015.007.037) and the Interuniversity Attraction Poles (PAI) (no. P7/07). P.D.S. is holder of a fundamental clinical research mandate by the FWO. We would like to thank Ferring Company (Aalst, Belgium) for financial support of this study. The authors do not have any competing interests to declare. TRIAL REGISTRATION NUMBER: Not applicable.

Hormonal contraceptive use is associated with altered bone structural and metabolic responses to military training in women: An observational cohort study.

Military training increases tibial density and size. Female sex hormones may influence the adaption of bone to loading, but it is unknown if women using different hormonal contraceptives adapt similarly to military training. One hundred and sixteen women (57 women not using hormonal contraceptives [non-users], 38 combined oral contraceptive pill [COCP] users, 21 depot medroxyprogesterone acetate [DMPA] users) completed this study. Tibial volumetric bone mineral density (vBMD) and geometry were measured by peripheral quantitative computed tomography (4 %, 14 %, 38 %, and 66 % sites) at the start (week 1) and end (week 14) of British Army basic training. Circulating markers of bone and calcium metabolism were measured at weeks 1, 2, 4, 6, 10, and 14. Training increased trabecular vBMD at the 4 % site, periosteal perimeter at the 14 % and 66 % sites, and total area, cortical area, cortical thickness, and bone strength at all sites (0.1 to 1.6 %, p ≤ 0.009), with no differences between hormonal contraceptive groups (p ≥ 0.127). Trabecular vBMD increased at the 14 % site in non-users (0.8 %, p = 0.005), but not in COCP or DMPA users (p ≥ 0.205). Periosteal perimeter increased at the 38 % site in COCP (0.4 %, p < 0.001) and DMPA (0.5 %, p < 0.001) users, but not in non-users (p = 0.058). Training had no effect on periosteal perimeter at the 4 % site or cortical vBMD or endosteal perimeter at any site (p ≥ 0.168). ßCTX decreased and PINP increased during training with no difference between hormonal contraceptive groups. Training increased iPTH in non-users, but not COCP or DMPA users. Hormonal contraceptives may exert site-specific effects on the mechanobiology of bone, with higher endogenous oestradiol promoting trabecularisation and inhibiting periosteal expansion in non-users compared with hormonal contraceptive users.

Eating Disorder Risk and Common Mental Disorders in British Servicewomen: A Cross-Sectional Observational Study.

PURPOSE: Servicewomen are at increased risk of common mental disorders compared with servicemen and their female civilian counterparts. The prevalence of eating disorder risk and common mental disorders, and associated risk factors in British servicewomen are poorly understood. METHODS: All women younger than 45 yr in the UK Armed Forces were invited to complete a survey about demographics, exercise behaviors, eating behaviors, and common mental disorders. RESULTS: A total of 3022 women participated; 13% of participants were at high risk of an eating disorder based on Brief Eating Disorder in Athletes Questionnaire and Female Athlete Screening Tool scores. Twenty-five percent of participants had symptoms of anxiety (seven-item Generalized Anxiety Disorder Assessment score ≥10), and 26% had symptoms of depression (nine-item Patient Health Questionnaire score ≥10). Older age was associated with a lower risk, and heavier body mass was associated with a higher risk, of eating disorders ( P ≤ 0.043). Older age and higher rank were associated with a lower risk of symptoms of anxiety and depression ( P ≤ 0.031), and a heavier body mass was associated with a higher risk of symptoms of depression ( P ≤ 0.012). Longer habitual sleep duration was associated with a lower risk of eating disorders and symptoms of anxiety and depression ( P ≤ 0.028). A higher volume of field exercise was associated with a lower risk, and a higher volume of military physical training and personal physical training was associated with a higher risk, of eating disorders ( P ≤ 0.024). Job role and deployment history were not associated with any outcome. CONCLUSIONS: Sleeping and training habits provide potential novel targets for exploring how common mental disorders can be managed in British servicewomen.

Unhealthy air quality secondary to wildfires is associated with lower blastocyst yield.

OBJECTIVE: To study the impact of unhealthy air quality from the 2020 Oregon wildfires on outcomes for patients undergoing in vitro fertilization (IVF) treatment. DESIGN: A retrospective cohort study. SETTING: A university-based fertility clinic. PATIENTS: Subjects were undergoing IVF treatment from the 6 weeks preceding the wildfires through a 10-day exposure period. Cohorts were classified on the basis of whether subjects experienced patient and/or laboratory exposure to unhealthy air quality. Patient exposure was defined as at least 4 days of ovarian stimulation overlapping with the exposure, and laboratory exposure was defined as at least 2 days of IVF treatment and embryogenesis overlapping with the exposure. The unexposed cohort consisted of remaining subjects without defined exposure, with cycles in the 6 weeks preceding the wildfires. As some subjects had dual exposure and appeared in both patient and laboratory exposure cohorts, each cohort was separately compared with the unexposed control cohort. INTERVENTION: A 10-day period of unhealthy air quality caused by smoke plumes from a wildfire event. MAIN OUTCOME MEASURES: The primary outcome was the blastulation rate. Secondary outcomes included fertilization rate, number of blastocysts obtained, and cycles with no blastocysts frozen or transferred. RESULTS: Sixty-nine subjects underwent ovarian stimulation and IVF treatment during the 6 weeks preceding the wildfires through the 10-day period of unhealthy air quality. Of these, 15 patients were in the laboratory exposure cohort, 16 were in the patient exposure cohort, and 44 were unexposed. Six subjects appeared in both laboratory and patient exposure cohorts. Although neither exposure cohort had significantly decreased blastulation rate compared with the unexposed, the median number of blastocysts obtained was significantly lower in the laboratory exposure cohort than the unexposed group (2 [range 0-14] vs. 4.5 [range 0-21], respectively). The laboratory exposure cohort had significantly more cycles with no blastocysts obtained (3/15 [20%] vs. 1/44 [2%]). There were no significant differences in IVF treatment outcomes between patient exposure and unexposed cohorts. These findings persisted after controlling for age. There were no significant differences in pregnancy outcomes observed after embryo transfer between the exposure group and the unexposed group. CONCLUSION: For a cohort of patients undergoing IVF treatment, an acute episode of outside wildfire smoke exposure during fertilization and embryogenesis was associated with decreased blastocyst yield.

Circulating extracellular vesicle characteristics differ between men and women following 12 weeks of concurrent exercise training.

Concurrent resistance and endurance exercise training (CET) has well-studied benefits; however, inherent hormonal and genetic differences alter adaptive responses to exercise between sexes. Extracellular vesicles (EVs) are factors that contribute to adaptive signaling. Our purpose was to test if EV characteristics differ between men and women following CET. 18 young healthy participants underwent 12-weeks of CET. Prior to and following CET, subjects performed an acute bout of heavy resistance exercise (AHRET) consisting of 6 × 10 back squats at 75% 1RM. At rest and following AHRET, EVs were isolated from plasma and characteristics and miRNA contents were analyzed. AHRET elevated EV abundance in trained men only (+51%) and AHRET-induced changes were observed for muscle-derived EVs and microvesicles. There were considerable sex-specific effects of CET on EV miRNAs, highlighted by larger variation following the 12-week program in men compared to women at rest. Pathway analysis based on differentially expressed EV miRNAs predicted that AHRET and 12 weeks of CET in men positively regulates hypertrophy and growth pathways more so than in women. This report highlights sex-based differences in the EV response to resistance and concurrent exercise training and suggests that EVs may be important adaptive signaling factors altered by exercise training.