Confidence in eating disorder knowledge does not predict actual knowledge in collegiate female athletes

Background Eating disorders are serious psychological disorders with long term health impacts. Athletic populations, tend to have higher incidences of eating disorders compared to the general population. Yet there is little known about athletes’ eating disorder knowledge and how it relates to their confidence in their knowledge. Therefore, the purpose of our study was to evaluate collegiate female athletes’ eating disorder (ED) knowledge and confidence in their knowledge. 51 participants were recruited from a National Association of Intercollegiate Athletics (NAIA) university in the mid-west and asked to complete a 30-question exam assessing one’s knowledge of five different categories related to eating disorders. Confidence in the correctness of answers was assessed with a 5-point Likert-scale (1 = very unconfident, 5 = very confident). A one-way ANOVA was used to determine differences between scores on different categories and overall scores. A simple regression analysis was used to determine if confidence or age was predictive in knowledge scores. Results The average score of participants was 69.1%, SD = 10.8% with an average confidence of 3.69/5, SD = 0.33. Athletes scored lowest with regards to Identifying Signs and Symptoms of EDs compared to other sub-scores (p < 0.05). There was no relationship between knowledge and confidence scores. Discussion There is limited ED knowledge among collegiate female athletes. This may be problematic as many athletes appear confident in the correctness of their answers despite these low scores. Coaches should be aware of this lack of knowledge and work with clinical practitioners, such as dieticians, team physicians and athletic trainers to educate and monitor their athletes on eating disorders, specifically signs and symptoms

Hindlimb suspension in Wistar rats: Sex-based differences in muscle response

Ground-based animal models have been used extensively to understand the effects of microgravity on various physiological systems. Among them, hindlimb suspension (HLS), developed in 1979 in rats, remains the gold-standard and allows researchers to study the consequences of total unloading of the hind limbs while inducing a cephalic fluid shift. While this model has already brought valuable insights to space biology, few studies have directly compared functional decrements in the muscles of males and females during HLS. We exposed 28 adult Wistar rats (14 males and 14 females) to 14 days of HLS or normal loading (NL) to better assess how sex impacts disuse-induced muscle deconditioning. Females better maintained muscle function during HLS than males, as shown by a more moderate reduction in grip strength at 7 days (males: −37.5 ± 3.1%, females: −22.4 ± 6.5%, compared to baseline), that remains stable during the second week of unloading (males: −53.3 ± 5.7%, females: −22.4 ± 5.5%, compared to day 0) while the males exhibit a steady decrease over time (effect of sex × loading p = 0.0002, effect of sex × time × loading p = 0.0099). This was further supported by analyzing the force production in response to a tetanic stimulus. Further functional analyses using force production were also shown to correspond to sex differences in relative loss of muscle mass and CSA. Moreover, our functional data were supported by histomorphometric analyzes, and we highlighted differences in relative muscle loss and CSA. Specifically, female rats seem to experience a lesser muscle deconditioning during disuse than males thus emphasizing the need for more studies that will assess male and female animals concomitantly to develop tailored, effective countermeasures for all astronauts

Mitochondrial Dysfunction in Diaphragm Muscle Precedes the Cachectic Phenotype in LLC Tumor-Bearing Mice.

The defining feature of cancer cachexia is extensive weight loss and skeletal muscle atrophy. It is clinically important because cachexia reduces patient survival, results in functional impairment, and is estimated to be directly responsible for 20-40% of cancer deaths. Unfortunately, no clinical therapy exists and therefore, it is important to understand the molecular mechanisms responsible for rapid muscle wasting. Compared to limb muscles, the diaphragm is relatively understudied in cancer cachexia, but is likely to be adversely affected because cachexia is a systemic disease. Wasting of the primary inspiratory muscle may result in difficulty breathing and inability to adjust minute ventilation in response to a respiratory challenge. Based on emerging evidence, it is clear that oxidative stress is present in cachexia-induced wasting of the diaphragm; PURPOSE: we developed the hypothesis that mitochondrial dysfunction in the diaphragm precedes cachexia. METHODS: 1X106 Lewis Lung Carcinoma cells (LLC) or Phosphate-Buffered Saline (PBS, control) were implanted to the hind-flank of C57BL6/J mice at 8 wks of age. Tumors were allowed to develop for 1, 2, 3, or 4 wks. At designated time points diaphragms were collected and mitochondrial function was assessed by respiration and ROS production. RESULTS: Cancer cachexia was evident only at the 4 wk time point demonstrated by decrease in body mass and muscle atrophy in several limb muscles. Mitochondrial respiration, assessed by respiratory control ratio (state3/state 4 respiration), was significantly lower at 1 wk (pCONCLUSIONS:The molecular events that lead to muscle atrophy in cancer cachexia are unknown. We demonstrate that two hallmarks of mitochondrial dysfunction, altered respiration and ROS production, occur in the diaphragm well before the cancer cachexia phenotype is evident in the LLC model. These data suggest that the mitochondria are likely a suitable target to treat or prevent cancer cachexia-induced muscle wasting in the diaphragm

Fractional Synthetic Rate and Markers of Protein Turnover are Altered in the Diaphragms of Cachectic Mice

Cancer cachexia, a wasting syndrome characterized by rapid skeletal muscle wasting and fat loss, directly accounts for up to 20-40% of cancer-related deaths. All muscles, including respiratory muscles, are susceptible to atrophy because cancer cachexia is a systemic disease. Atrophy of the primary breathing muscle, the diaphragm, can lead to respiratory distress, which is commonly associated with a cachectic phenotype. Indeed, the diaphragm is more susceptible to atrophy in certain conditions, but little is known about the effects of cancer-cachexia on protein turnover in the diaphragm. Therefore, investigations into the alterations in protein turnover could provide insight to the molecular events and provide valuable information in the search for therapeutic targets. PURPOSE: The purpose of this study was to describe changes in diaphragmatic protein synthesis and molecular markers of synthesis and degradation during the progression of cancer cachexia. METHODS: C57BL6/J mice (8 wks old) were implanted with 1X106 Lewis Lung Carcinoma cells (LLC) or Phosphate-Buffered Saline (PBS, control). Tumors developed over a 1-4 wk time course and diaphragms were harvested at each time point (1, 2, 3, or 4 wks). Fractional synthetic rates (FSR) were determined using deuterium incorporation into muscle. Selected markers of protein synthesis and degradation pathways were analyzed by immunoblot analysis. One-Way ANOVA was used for statistical analyses, with significance set at pRESULTS: FSR trended downward over time, but did not reach significance. Similar to FSR, anabolic signaling markers (4EBP-1, ERK1/2, Deptor) did not demonstrate significant differences. p62, an autophagic degradation marker, was significantly less than PBS in 3 wk diaphragms (

Leucine Supplementation Exacerbates Morbidity in Male but Not Female Mice with Colorectal Cancer-Induced Cachexia

Cancer cachexia (CC) is a multifactorial wasting syndrome characterized by a significant loss in lean and/or fat mass and represents a leading cause of mortality in cancer patients. Nutraceutical treatments have been proposed as a potential treatment strategy to mitigate cachexia-induced muscle wasting. However, contradictory findings warrant further investigation. The purpose of this study was to determine the effects of leucine supplementation on skeletal muscle in male and female ApcMin/+ mice (APC). APC mice and their wild-type (WT) littermates were given normal drinking water or 1.5% leucine-supplemented water (n = 4–10/group/sex). We measured the gene expression of regulators of inflammation, protein balance, and myogenesis. Leucine treatment lowered survival rates, body mass, and muscle mass in males, while in females, it had no effect on body or muscle mass. Leucine treatment altered inflammatory gene expression by lowering Il1b 87% in the APC group and decreasing Tnfa 92% in both WT and APC males, while it had no effect in females (p \u3c 0.05). Leucine had no effect on regulators of protein balance and myogenesis in either sex. We demonstrated that leucine exacerbates moribundity in males and is not sufficient for mitigating muscle or fat loss during CC in either sex in the ApcMin/+ mouse

Disuse Atrophy Occurs Without a Change in Mitochondrial Respiratory Control Ratio During Hindlimb Unloading in Mice

Skeletal muscle atrophy commonly occurs during prolonged periods of inactivity, however, the precise mechanisms that cause muscle atrophy have yet to be determined. Specifically, a controversy exists on whether mitochondrial dysfunction is a cause or consequence of disuse muscle atrophy. PURPOSE: The purpose of this study was to determine if a change in the respiratory control ratio, which is a ratio of maximal O2 respiration to leak respiration, could be detected prior to muscle atrophy in a time-course study in mice. METHODS: Disuse atrophy was induced using hindlimb unloading (HU) in adult, C57BL/6J male mice for 0 (control), 1, 2, 3, or 7 days (n=~6-8/group). Following completion, gastrocnemius and soleus muscles were weighed and assessed for mitochondrial function in permeabilized muscle fibers. Here, we define mitochondrial function as the respiratory control ratio (RCR) determined by maximal ADP stimulated respiration (State 3) divided by leak or ATP synthase inhibited (state 4) respiration. A one-way ANOVA was used to determine differences between means. When significant F ratios were found, a Tukey post-hoc was used to compare differences between means. Values presented are mean ± standard error RESULTS: In both the gastrocnemius and soleus, muscle mass was not significantly different from control at day 1, but was significantly lower at 2, 3, and 7-day timepoints. In contrast, there was no significant difference in RCR in gastrocnemius (control 3.11±0.20, 1 day 2.96±0.60, 2 day 3.07±0.31, 3 day 3.08±0.25, 7 day 3.41± .29) or soleus (control 2.33±0.33, 1 day 2.77±0.33, 2 day 3.03±0.51, 3 day 2.93±0.30, 7 day 2.78±0.48). CONCLUSION: It is well established that HU causes rapid muscle atrophy. These data support mitochondrial RCR does decrease before muscle atrophy in either gastrocnemius or soleus muscle, and therefore may not be a primary cause of HU-induced muscle atrophy in mice

Mitochondrial function and protein turnover in the diaphragm are altered in llc tumor model of cancer cachexia

It is established that cancer cachexia causes limb muscle atrophy and is strongly associated with morbidity and mortality; less is known about how the development of cachexia impacts the diaphragm. The purpose of this study was to investigate cellular signaling mechanisms related to mitochondrial function, reactive oxygen species (ROS) production, and protein synthesis during the development of cancer cachexia. C57BL/J6 mice developed Lewis Lung Carcinoma for either 0 weeks (Control), 1 week, 2 weeks, 3 weeks, or 4 weeks. At designated time points, diaphragms were harvested and analyzed. Mitochondrial respiratory control ratio was ~50% lower in experimental groups, which was significant by 2 weeks of cancer development, with no difference in mitochondrial content markers COXIV or VDAC. Compared to the controls, ROS was 4-fold elevated in 2-week animals but then was not different at later time points. Only one antioxidant protein, GPX3, was altered by cancer development (~70% lower in experimental groups). Protein synthesis, measured by a fractional synthesis rate, appeared to become progressively lower with the cancer duration, but the mean difference was not significant. The development and progression of cancer cachexia induces marked alterations to mitochondrial function and ROS production in the diaphragm and may contribute to increased cachexia-associated morbidity and mortality

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

Mitochondrial degeneration precedes the development of muscle atrophy in progression of cancer cachexia in tumour-bearing mice

Background: Cancer cachexia is largely irreversible, at least via nutritional means, and responsible for 20–40% of cancer-related deaths. Therefore, preventive measures are of primary importance; however, little is known about muscle perturbations prior to onset of cachexia. Cancer cachexia is associated with mitochondrial degeneration; yet, it remains to be determined if mitochondrial degeneration precedes muscle wasting in cancer cachexia. Therefore, our purpose was to determine if mitochondrial degeneration precedes cancer-induced muscle wasting in tumour-bearing mice. Methods: First, weight-stable (MinStable) and cachectic (MinCC) ApcMin/+ mice were compared with C57Bl6/J controls for mRNA contents of mitochondrial quality regulators in quadriceps muscle. Next, Lewis lung carcinoma (LLC) cells or PBS (control) were injected into the hind flank of C57Bl6/J mice at 8 week age, and tumour allowed to develop for 1, 2, 3, or 4 weeks to examine time course of cachectic development. Succinate dehydrogenase stain was used to measure oxidative phenotype in tibialis anterior muscle. Mitochondrial quality and function were assessed using the reporter MitoTimer by transfection to flexor digitorum brevis and mitochondrial function/ROS emission in permeabilized adult myofibres from plantaris. RT-qPCR and immunoblot measured the expression of mitochondrial quality control and antioxidant proteins. Data were analysed by one-way ANOVA with Student–Newman–Kuels post hoc test. Results: MinStable mice displayed ~50% lower Pgc-1α, Pparα, and Mfn2 compared with C57Bl6/J controls, whereas MinCC exhibited 10-fold greater Bnip3 content compared with C57Bl6/J controls. In LLC, cachectic muscle loss was evident only at 4 weeks post-tumour implantation. Oxidative capacity and mitochondrial content decreased by ~40% 4 weeks post-tumour implantation. Mitochondrial function decreased by ~25% by 3 weeks after tumour implantation. Mitochondrial degeneration was evident by 2 week LLC compared with PBS control, indicated by MitoTimer red/green ratio and number of pure red puncta. Mitochondrial ROS production was elevated by ~50 to ~100% when compared with PBS at 1–3 weeks post-tumour implantation. Mitochondrial quality control was dysregulated throughout the progression of cancer cachexia in tumour-bearing mice. In contrast, antioxidant proteins were not altered in cachectic muscle wasting. Conclusions: Functional mitochondrial degeneration is evident in LLC tumour-bearing mice prior to muscle atrophy. Contents of mitochondrial quality regulators across ApcMin/+ and LLC mice suggest impaired mitochondrial quality control as a commonality among pre-clinical models of cancer cachexia. Our data provide novel evidence for impaired mitochondrial health prior to cachectic muscle loss and provide a potential therapeutic target to prevent cancer cachexia

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals &lt;1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data