Cancer patients who exhibit cachexia lose weight and have low treatment tolerance and poor outcomes compared to cancer patients without weight loss. Despite the clear increased risk for patients, diagnosing cachexia still often relies on self-reported weight loss. A reliable biomarker to identify patients with cancer cachexia would be a valuable tool to improve clinical decision making and identification of patients at risk of adverse outcomes. Targeted metabolomics, that included panels of amino acids, tricarboxylic acids, fatty acids, acylcarnitines, and sphingolipids, were conducted on plasma samples from patients with confirmed pancreatic ductal adenocarcinoma (PDAC) with and without cachexia and control patients without cancer (n=10/group, equally divided by sex). Additional patient samples were analyzed (total n=95) and Receiver Operating Characteristic (ROC) analyses were performed to establish if any metabolite could effectively serve as a biomarker of cachexia. Targeted profiling revealed that cachectic patients had decreased circulating levels of three sphingolipids compared to either non-cachectic PDAC patients or patients without cancer. The ratio of C18-ceramide to C24-ceramide (C18:C24) outperformed a number of other previously proposed biomarkers of cachexia (area under ROC = 0.810). It was notable that some biomarkers, including C18:C24, were only altered in cachectic males. Our findings identify C18:C24 as a potentially new biomarker of PDAC-induced cachexia that also highlight a previously unappreciated sexual dimorphism in cancer cachexia.

Abstract BackgroundPreoperative skeletal muscle deficiency is an established risk factor for poor survival outcomes in patients with renal cell carcinoma (RCC). However, given the dynamic nature of skeletal muscle associated with malignancy, there is a need to evaluate the prognostic benefit of muscle area change from the preoperative to postoperative period. We hypothesize that an improvement in muscle area following nephrectomy, measured by linear segmentation of L3 psoas and paraspinal musculature, is associated with improvement in overall survival (OS) and cancer specific survival (CSS) for patients with pT3 and pT4 RCC.MethodsWe retrospectively analysed 270 pT3 and pT4 RCC patients who underwent nephrectomy from March 2004 to February 2020 with available preoperative and postoperative axial CT or MRI studies segmented at the L3 vertebrae. The majority were N0 (79%) and M0 (68%). Psoas and paraspinal muscles were measured bilaterally using a validated digital ruler tool. Total muscle area (TMA) was calculated by aggregating the area of all four muscles and total muscle area index (TMI) by dividing the TMA by height squared (m2). The prognostic value of postoperative muscle improvement, defined as any increase in muscle area index, was analysed using Kaplan–Meier and Cox proportional stepwise hazard models.ResultsMedian time between preoperative scans and surgery was approximately 22 days and between surgery and postoperative scans 172 days. One hundred twenty‐one patients (44.8%) had an increase in total muscle area index post‐nephrectomy (IQR = 33.4; P ≤ 0.0001). On Kaplan–Meier analysis, postoperative improvement in TMI was associated with decreased odds of mortality (P = 0.0024) with a median follow‐up of 38.6 months. In a multivariable Cox regression analysis, improvement of TMI was associated with increased OS (HR = 0.52, 95% CI 0.35–0.78, P < 0.001) and increased CSS (HR = 0.55, 95% CI 0.32–0.94, P = 0.030). A 5% or more improvement in TMI was also associated with increased OS (HR = 0.53, 95% CI 0.34–0.84, P = 0.006) and increased CSS (HR = 0.46, 95% CI 0.24–0.86, P = 0.015).ConclusionsAny improvement in TMI between preoperative and initial postoperative imaging after nephrectomy was associated with increased OS and CSS in patients with pT3 and pT4 RCC. Perioperative linear segmentation is an efficient tool that may improve current prognostication methods and can be performed on any imaging software platform.

Abstract BackgroundThe age‐related loss of muscle mass and quality, sarcopenia, has many contributing factors, one of which may be cellular senescence, but this is not well defined in human skeletal muscle.MethodPrimary cells were isolated from biopsy samples of the vastus lateralis muscle from healthy adult males (n = 6, 22 ± 1 years), sorted (magnetic activated cell sorting) and chemically induced (doxorubicin, DOX, 0.2 μM) to a senescent state. This allowed the parallel and simultaneous investigation of the two main skeletal muscle‐derived cell types: satellite cell‐derived CD56+ve/desmin+ve myoblasts (muscle precursor cells) and CD56− ve/TE7+ve fibroblasts (at >95% purity). Both cell types were followed for up to 35 days post DOX treatment with a combination of quantitative immunocytochemistry and qRT‐PCR for senescent markers and senescence‐associated secretory phenotype (SASP) factors.ResultsMyoblasts and fibroblasts showed temporal and quantitative differences in many of the senescence markers studied. p16 protein expression increased across the time course (P < 0.0001) with no difference between cell types, whereas at the mRNA level, myoblasts showed increased p16 expression from 4 days post treatment (FC = 3.03 ± 0.99), and in fibroblasts, this appeared later at 10 and 35 days post DOX treatment (FC = 8.09 ± 2.46, P < 0.0001). Both myoblasts (FC = 8.83 ± 1.72) and fibroblasts (FC = 2.33 ± 1.10) showed significant increases in p21 mRNA (P < 0.0001), which remained elevated in the myoblast cell populations across the 35‐day time course but returned to baseline in the fibroblasts from 4 days post DOX treatment. Within 35 days post DOX treatment, all cell populations of both myoblasts and fibroblasts had reached 100% SA‐β‐Gal‐positive cells (P < 0.05). γH2aX expression (a marker of DNA damage) increased 1 day after DOX treatment in the myoblasts (FC = 3.3 ± 1.1, P < 0.05) but returned to baseline within 4 days post DOX treatment, whereas fibroblasts showed a similar trend that did not reach statistical significance. Significant reductions in expression of the proliferation marker Ki67 1 day post DOX treatment were seen in both cell types and were maintained throughout the time course (FC = 0.11 ± 0.07, P < 0.0001). Significant changes over the time course were also observed in mRNA expression of selected SASP factors (e.g. PAI‐1, MMP3, and IGFBP3, P < 0.05).ConclusionsNeither cellular senescence nor sarcopenia is fully understood. The present data on human primary myoblasts and fibroblasts obtained from the same tissue sample show that senescence is a complex, non‐linear, and dynamic cellular process which shows intra‐ and inter‐cell variability.

Abstract BackgroundComputed tomography (CT)‐derived measures of tissue quality can add to frailty assessment and improve selection of candidates for heart transplant. We investigated the prognostic value of CT measures of tissue density for predicting hospital length of stay (LOS) and mortality post‐transplant.MethodsAll patients at a quaternary care hospital between 1999 to 2018 with preheart transplant CT scans and available data on transplant outcomes were eligible (n = 189), including a subset within the total cohort with scans 6‐month pretransplant (n = 103). Axial chest CT scans were analysed for liver and muscle density at the 12th thoracic vertebrae and aortic arch landmarks, respectively. Cox and linear regression models examined the risk of death and LOS, respectively, according to median (above or below) pectoral muscle density. Low‐density muscle (LDM) area and liver density were analysed as continuous variables.ResultsOut of 157 patients with readable CT scans (median age 55 years, interquartile range [50–60] 10% women), 31 died on 1‐year follow up. Patients with higher than and at median pectoral muscle density (39.5 Hounsfield Unit [HU]) had better 1‐year survival in the overall cohort (hazard ratio [HR] 0.82, 95% confidence interval [CI] 0.673, 0.989; p = 0.039), with the 6‐month cohort showing a trend (HR 0.79, 95% CI 0.603, 1.023; p = 0.074) towards improved survival. Conversely, every 5‐cm2 increase in pectoral LDM area was associated with 2.4‐day lower LOS (p = 0.045) in the overall cohort, and a 2.6‐day lower LOS in the 6‐month cohort (p = 0.05). Patients with higher ratio of normal‐density muscle to LDM had higher LOS (p < 0.01). Every 5‐HU increase in liver density at a region of interest was associated with 0.24‐day higher post‐transplant LOS in the overall cohort, and a 0.41 higher LOS in the 6‐month cohort (p ≤ 0.05).ConclusionsPatients with higher preheart transplant pectoral muscle density had greater 1‐year survival. Higher pectoral LDM area was associated with decreased LOS post‐transplant and higher liver density was associated with increased LOS. These findings raise possibilities that measures of muscle density as they reflect to quality of muscle may have prognostic implications. Future studies with prospective design are needed to confirm these findings.

Abstract BackgroundPsoas muscle area (PMA) has recently been found to be an appropriate surrogate for whole‐body skeletal muscle mass and a measure of sarcopenia. Sarcopenia, which includes a decreased muscle mass and correlated with physical disability, morbidity, and mortality, is prevalent in and has deleterious consequences for patients with chronic kidney disease (CKD). The current study investigates the association of PMA, as a marker of sarcopenia, with baseline kidney function and CKD progression.MethodsA retrospective cohort study was conducted in a community hospital nephrology clinic setting. For this study, sarcopenia was defined as standardized PMA measured at the L3 level either as below 25th percentile or below the median. Progression of CKD was measured by estimated glomerular filtration rate (eGFR) decline rate and change in proteinuria. To assess sarcopenia as a predictor of baseline CKD Grade 4 or 5, a multivariate logistic regression model was applied using standardized PMA < 25th percentile. To assess sarcopenia as a predictor for eGFR slope, a multivariate generalized linear model was applied using PMA < median.ResultsAmong the 230 patients studied, the median age was 74 (31–92) years, 56.52% (130/230) were male patients, and the majority was Caucasian 62.61% (144/230). The body mass index classes of <18.5, 18.5–24.9, 25.0–29.9, and >29.9 kg/m2 were categorized by 4.8% (11/229), 24.89% (57/229), 33.19% (76/229), and 37.12% (85/229), respectively. Sarcopenia as a predictor of baseline CKD Grade 4 or 5 had an unadjusted odds ratio of 1.46 (0.87–2.63) and an adjusted odds ratio of 2.45 (1.13–5.31). Sarcopenia as a predictor for eGFR slope had an unadjusted odds ratio of 1.01 (1.00–1.04) and an adjusted odds ratio 1.03 (1.00–1.05).ConclusionsSarcopenia, as measured by PMA, predicts a significant risk of more severe baseline CKD grade and progression. Future studies should examine sex‐specific muscle mass tools as a predictor of renal function to create more targeted interventions.

Abstract BackgroundThe relationships between cachexia stages and the Functional Assessment of Anorexia/Cachexia Therapy Anorexia Cachexia Subscale (FAACT ACS) 12‐item, 5‐item anorexia symptoms, and 4‐item anorexia concerns have not been investigated in Asian patients with advanced cancer.MethodsThis is a multicentre questionnaire survey conducted in palliative and supportive care settings across Japan. Consecutive patients were enrolled. Patient characteristics and anthropometric measurements were obtained. Dietary intakes and nutrition impact symptoms were also assessed. Patients evaluated their quality of life (QOL) using FAACT ACS. Subjects were divided into two groups, that is, pre‐cachexia (non‐cachexia) and cachexia and refractory cachexia (cachexia), based on cancer cachexia criteria from the international consensus. Comparisons were performed using the Mann–Whitney U test or chi‐squared test. To evaluate the relationship between cachexia stages and FAACT ACS 12‐item, 5‐item anorexia symptoms, and 4‐item anorexia concerns, adjusted odd ratios (ORs) and 95% confidence intervals (CIs) were calculated in the logistic models.ResultsAmong 495 patients, 378 (76.4%) responded. Due to missing data, 344 patients were classified into the non‐cachexia group (n = 174) and cachexia group (n = 170), and 318 remained in the analysis of FAACT ACS. The cachexia group had a more impaired performance status, a lower body mass index, and a higher frequency of weight loss in 1 month (P = 0.021, <0.001, and <0.001, respectively). Advancing stages were associated with lack of appetite and reduced dietary intakes (P < 0.001 and P < 0.001, respectively). QOL scores were significantly worse in the cachexia group in FAACT ACS 12‐item, 5‐item anorexia symptoms, and 4‐item anorexia concerns (P < 0.001, P = 0.001, and P < 0.001, respectively). In the models of FAACT ACS 12‐item, 5‐item anorexia symptoms, and 4‐item anorexia concerns, significantly higher adjusted ORs than in the non‐cachexia group were observed in the cachexia group [2.24 (95% CI 1.34–3.77), P = 0.002; 1.77 (95% CI 1.08–2.92), P = 0.024; and 2.18 (95% CI 1.29–3.70), P = 0.004, respectively].ConclusionsFAACT ACS 12‐item, 5‐item anorexia symptoms, and 4‐item anorexia concerns are useful for identifying patients at risk of QOL that deteriorates with advancing stages in this population.

Abstract BackgroundChronic obstructive pulmonary disease (COPD) is associated with skeletal muscle mitochondrial dysfunction. Resistance exercise training (RT) is a training modality with a relatively small pulmonary demand that has been suggested to increase skeletal muscle oxidative enzyme activity in COPD. Whether a shift into a more oxidative profile following RT also translates into increased mitochondrial respiratory capacity in COPD is yet to be established.MethodsThis study investigated the effects of 13 weeks of RT on m. vastus lateralis mitochondrial capacity in 11 persons with moderate COPD [45% females, age: 69 ± 4 years (mean ± SD), predicted forced expiratory volume in 1 s (FEV1): 56 ± 7%] and 12 healthy controls (75% females, age: 66 ± 5 years, predicted FEV1: 110 ± 16%). RT was supervised and carried out two times per week. Leg exercises included leg press, knee extension, and knee flexion and were performed unilaterally with one leg conducting high‐load training (10 repetitions maximum, 10RM) and the other leg conducting low‐load training (30 repetitions maximum, 30RM). One‐legged muscle mass, maximal muscle strength, and endurance performance were determined prior to and after the RT period, together with mitochondrial respiratory capacity using high‐resolution respirometry and citrate synthase (CS) activity (a marker for mitochondrial volume density). Transcriptome analysis of genes associated with mitochondrial function was performed.ResultsResistance exercise training led to similar improvements in one‐legged muscle mass, muscle strength, and endurance performance in COPD and healthy individuals. In COPD, mitochondrial fatty acid oxidation capacity and oxidative phosphorylation increased following RT (+13 ± 22%, P = 0.033 and +9 ± 23%, P = 0.035, respectively). Marked increases were also seen in COPD for mitochondrial volume density (CS activity, +39 ± 35%, P = 0.001), which increased more than mitochondrial respiration, leading to lowered intrinsic mitochondrial function (respiration/CS activity) for complex‐1‐supported respiration (−12 ± 43%, P = 0.033), oxidative phosphorylation (−10 ± 42%, P = 0.037), and electron transfer system capacity (−6 ± 52%, P = 0.027). No differences were observed between 10RM and 30RM RT, nor were there any adaptations in mitochondrial function following RT in healthy controls. RT led to differential expression of numerous genes related to mitochondrial function in both COPD and healthy controls, with no difference being observed between groups.ConclusionsThirteen weeks of RT resulted in augmented skeletal muscle mitochondrial respiratory capacity in COPD, accompanied by alterations in the transcriptome and driven by an increase in mitochondrial quantity rather than improved mitochondrial quality.

Abstract BackgroundLong‐term exposure to microgravity during spaceflight has adverse effects on human health including muscle atrophy, impaired immune function, and alterations in gut microbiome profile. Gut microorganisms influence a wide range of host biological processes, but their interactions with skeletal muscle and the immune system under microgravity have yet to be elucidated.MethodsRhesus macaques (Macaca mulatta) were subjected to −6° head‐down tilted bed rest (HDBR) for 6 weeks. Faecal samples, skeletal muscle tissue, and peripheral blood mononuclear cells (PBMCs) were collected for metagenomic, metabolomic, and transcriptomic analyses, respectively, and further integrated for a multi‐omics analysis.ResultsHead‐down tilted bed rest significantly altered taxon abundance (P < 0.05) in 1 class, 5 orders, 11 families, 55 genera, and 122 species of microbes. We also identified the significantly changed metabolites (P < 0.05, fold change >1.2, variable importance in projection >1) in atrophied muscles, including some crucial metabolites (such as l‐alanine and l‐carnitine) and hub metabolites (such as pyridoxamine and epinephrine) involved in energy metabolism. Transcriptomic analysis of PBMCs revealed genes related to leucocyte activation, differentiation, and interleukin‐2 production that were differentially expressed as a result of HDBR exposure (fold change >2 and P < 0.05). By integrating multi‐omics analysis, we identified three bacterial genera (Klebsiella, Kluyvera, and Bifidobacterium) that were closely associated with immune dysfunction and five (including Oligella, Sporosarcina, Citrobacter, Weissella, and Myroides) that were associated with abnormal metabolism of amino acids in atrophied muscles induced by HDBR. The reduced abundance of butyrate‐producing colon bacteria Eubacterium, Roseburia, and their cross‐feeding bacteria Bifidobacteria may contribute to the impaired immune function and muscle atrophy caused by HDBR.ConclusionsThis is the first report of the HDBR‐associated changes in gut microbiota composition, metabolomics of skeletal muscle, and transcripts of PBMCs in a non‐human primate. The underlying microbiota–muscle and microbiota–immune interactions during simulated microgravity imply that modulation of gut microbiota may represent a novel strategy for enhancing the health and safety of crew members during long‐term space expeditions.

The menopause transition is marked by hormonal shifts leading to body composition changes, such as fat mass gain and lean mass loss. Weight-bearing and resistance exercise can help maintain lean mass during the menopause transition; however, uptake is low. Pre-clinical research points to bisphosphonates as also being effective in preventing loss of lean mass. Thus, we sought to investigate whether bisphosphonate therapy can mitigate loss of lean mass and outperform weight-bearing exercise in the years immediately following menopause. Data come from the Heartland Osteoporosis Prevention Study (NCT02186600), where osteopenic, postmenopausal women were randomized to bisphosphonate (n=91), weight-bearing/resistance exercise (n=92), or control (n=93) conditions over a one-year period. Dual energy X-ray absorptiometry (DXA)-derived body composition measures (including total lean mass, total fat mass, lean mass index, and lean mass-to-fat mass ratio) were ascertained at baseline, six, and 12-months. Adherence to risedronate and weight-bearing exercise was defined as the percentage of dosages taken and exercise sessions attended. Intent-to-treat analysis using linear modeling was used to generate treatment effects on body composition. Secondary analysis utilized per-protocol analysis and included adjustment for weight change. 276 women (age: 54.5 years; 83.3% Caucasian; BMI: 25.7 kg/m2) were included in the analyses. 12-month adherence to the risedronate and exercise interventions was 89% and 64%, respectively. Group-by-time interactions were observed for lean mass, revealing exercise (0.43±1.49kg) and risedronate groups (0.31±1.68 kg) gained significantly more lean mass than control (-0.15±1.27 kg) over 12-months. However, after controlling for weight change in secondary analysis, the difference in lean mass change between control and risedronate became non-significant (p=0.059). Results suggest both 12 months of oral risedronate and 12 months of weight-bearing exercise may diminish lean mass loss experienced during the menopause transition as compared to control. The lean mass sparing effect for risedronate may be driven by overall weight change.

Skeletal muscle mass is regulated by intracellular anabolic and catabolic activities. Increased catabolic activity can shift the balance towards net protein breakdown and muscle atrophy. Mitochondrial oxidative stress activates catabolism and is linked to muscle loss. Reducing mitochondrial oxidative stress is thus a plausible approach to prevent muscle atrophy. We tested this concept in age-dependent muscle atrophy by genetically overexpressing the mitochondrial antioxidant thioredoxin-2 (TXN2). We tested the functional role of TXN2 using ageing (n=7-10 per group) and denervation (n=3 per group) models in a transgenic mouse line that overexpresses TXN2. We investigated if overexpression of TXN2 blocks muscle loss in these models by examination of muscle weight, fibre size, and fibre number in young (~7months) and aged (~26months) TXN2-transgenic mice and controls. We studied the underlying mechanisms by mRNA and protein assays including transcriptomic profiling, western blot analysis, immunostaining, as well as succinate dehydrogenase, dihydroethidium, and terminal deoxynucleotidyl transferase dUTP nick end labelling staining. Overexpression of TXN2 did not significantly alter the baseline skeletal muscle size, weight, fibre type distribution, or expression of mitochondrial respiratory chain components, but it did preserve muscle mass during ageing. The hindlimb muscle mass in aged TXN-transgenic mice was ~21-24% greater (in tibialis anterior, gastrocnemius/soleus combined, and tibialis anterior/extensor digitorum longus combined) than in age-matched controls (all P<0.05). The reduction in both muscle fibre number (872±206 vs, 637±256 fibres in extensor digitorum longus muscle, P<0.05) and muscle fibre size (1959±296 vs. 1477±564μm2 in tibialis anterior muscle, P<0.05) seen in young vs. aged control muscles was not significant in young vs. aged TXN-transgenic mice (both P>0.05). Transcriptomic analysis revealed that catabolic genes that are up-regulated in ageing muscle, including those subserving apoptosis and the ubiquitin-like conjugation system, were normalized by TXN2 overexpression. Further, overexpressing TXN2 suppressed oxidative stress and caspase-9/3-mediated apoptotic signalling in the aged muscle at the protein level. Although denervation and its effects have been considered a component of age-related muscle atrophy, TXN2 overexpression failed to attenuate atrophy in an acute denervation model (TXN-transgenic vs. control mice, P>0.05), despite preventing denervation-induced oxidative stress and apoptosis. Mitochondrial oxidative stress appears to play a crucial role in effecting chronic age-dependent, but not acute neurogenic, muscle atrophy. Increased TXN2 protects muscle against oxidative stress-associated catabolic activity in ageing muscle and thus is a potential therapeutic approach to attenuate age-related muscle atrophy.