Anabolic Signaling Research Articles

Physicochemical, technofunctional, in vitro antioxidant, and in situ muscle protein synthesis properties of a sprat (Sprattus sprattus) protein hydrolysate.

Sprat (Sprattus sprattus) is an underutilized fish species that may act as an economic and sustainable alternative source of protein due to its good amino acid (AA) profile along with its potential to act as a source of multiple bioactive peptide sequences. This study characterized the physicochemical, technofunctional, and in vitro antioxidant properties along with the AA profile and score of a sprat protein enzymatic hydrolysate (SPH). Furthermore, the impact of the SPH on the growth, proliferation, and muscle protein synthesis (MPS) in skeletal muscle (C2C12) myotubes was examined. The SPH displayed good solubility and emulsion stabilization properties containing all essential and non-essential AAs. Limited additional hydrolysis was observed following in vitro-simulated gastrointestinal digestion (SGID) of the SPH. The SGID-treated SPH (SPH-SGID) displayed in vitro oxygen radical antioxidant capacity (ORAC) activity (549.42 μmol TE/g sample) and the ability to reduce (68%) reactive oxygen species (ROS) production in C2C12 myotubes. Muscle growth and myotube thickness were analyzed using an xCELLigence™ platform in C2C12 myotubes treated with 1 mg protein equivalent.mL-1 of SPH-SGID for 4 h. Anabolic signaling (phosphorylation of mTOR, rpS6, and 4E-BP1) and MPS (measured by puromycin incorporation) were assessed using immunoblotting. SPH-SGID significantly increased myotube thickness (p < 0.0001) compared to the negative control (cells grown in AA and serum-free medium). MPS was also significantly higher after incubation with SPH-SGID compared with the negative control (p < 0.05). These preliminary in situ results indicate that SPH may have the ability to promote muscle enhancement. In vivo human studies are required to verify these findings.

Creatine Supplementation and Akt/mTOR Pathway: Unraveling the Connection for Optimal Muscle Performance

This review paper investigates the relationship between creatine supplementation and the Akt/mTOR pathway, focusing on their impact on muscle performance. The Akt/mTOR pathway is a crucial signaling pathway that regulates muscle protein synthesis and hypertrophy in response to growth factors, nutrients, and mechanical stimuli. Recent evidence suggests that creatine supplementation can influence anabolic signaling pathways, including the phosphorylation of p70S6K, a downstream target of mTOR, leading to enhanced activation of the Akt/mTOR pathway. Additionally, creatine supplementation has been shown to increase intramuscular creatine and phosphocreatine levels, improving ATP availability during exercise and enhancing high-intensity muscle contractions. Understanding the complex regulatory mechanisms of the Akt/mTOR pathway is vital for optimizing muscle performance, as dysregulated signaling can hinder muscle protein synthesis and hypertrophic responses. This review highlights the potential of creatine supplementation to modulate the Akt/mTOR pathway, offering insights into its mechanisms and implications for muscle performance enhancement. By unraveling this connection, researchers and practitioners can develop targeted strategies to maximize muscle performance and promote adaptive responses in various exercise and athletic contexts.

Computational and experimental characterizations of the spatiotemporal activity and functional role of TGF-β in the synovial joint

TGF-β is a prominent anabolic signaling molecule associated with synovial joint health. Recent work has uncovered mechanochemical mechanisms that activate the latent form of TGF-β (LTGF-β) in the synovial joint—synovial fluid (SF) shearing or cartilage compression—pointing to mechanobiological phenomena, whereby enhanced TGF-β activity occurs during joint stimulation. Here, we implement computational and experimental models to better understand the role of mechanochemical-activated TGF-β (aTGF-β) in regulating the functional biosynthetic activities of synovial joint tissues. Reaction-diffusion models describe the pronounced role of extracellular chemical reactions—load-induced activation, reversible ECM-binding, and cell-mediated internalization—in modulating the spatiotemporal distribution of aTGF-β in joint tissues. Of note, aTGF-β from SF shearing predominantly acts on cells in peripheral tissue regions (superficial zone [SZ] chondrocytes and synoviocytes) and aTGF-β from cartilage compression acts on chondrocytes through all cartilage layers. Further, ECM reversible binding sites in cartilage act to modulate the temporal delivery of aTGF-β to cells, creating a dynamic where short durations of joint activity give rise to extended periods of aTGF-β exposure at moderated doses. Ex vivo tissue models were subsequently utilized to characterize the influence of physiologic aTGF-β activity regimens in regulating functional biosynthetic activities. Physiologic exposure regimens of aTGF-β in SF induce strong 4-fold to 9-fold enhancements in the secretion rate of the synovial biolubricant, PRG4, from SZ cartilage and synovium explants. Further, aTGF-β inhibition in cartilage over 1-month culture leads to a pronounced loss of GAG content (30–35% decrease) and tissue softening (60–65% EY reduction). Overall, this work advances a novel perspective on the regulation of TGF-β in the synovial joint and its role in maintaining synovial joint health.

Sex differences in cachexia outcomes, anabolic signalling and BCAA metabolism following chemotherapy

Sex is a risk factor for cancer. Affecting nearly 80% of cancer patients is cachexia, a body and skeletal muscle-wasting syndrome. Poor nutritional status, tumour related factors and chemotherapy contribute to cachexia. Although negative effects of chemotherapy on skeletal muscle are documented, the majority of studies are completed in male animals. Although the branched-chain amino acids (BCAA: leucine, isoleucine and valine) activate anabolic signalling in skeletal muscle and have been shown to reduce some of the effects of cachexia, BCAA nutritional support does not reverse cachexia and few studies have investigated the effects of chemotherapy on BCAA metabolism in skeletal muscle. Therefore, the objective of this study is to compare the effects of chemotherapy on cachexia outcomes and BCAA metabolism between male and female mice. Three-month-old CD2F1 male and female mice were treated with either the chemotherapy drug cocktail folfiri (50mg/kg 5-fluorouracil (5FU), 90mg/kg Leucovorin and 24mg/kg CPT11) (drug) or vehicle (10% DMSO in saline) for 6-weeks. Within sex, drug treatment led to reductions in body and skeletal muscle weights. Between sex, drug-treated female mice lost more body and gastrocnemius muscle weight compared to drug-treated males. Within sex, drug treated animals exhibited reductions in anabolic signalling, specifically AKT, S6K1, S6 and 4E-BP1. Between sex, drug-treated females mice showed greater loss of anabolic signalling compared to drug-treated males. Within sex, the drug cocktail reduced amino acid transporters SNAT1 and LAT1, while reducing BCAA concentrations in the skeletal muscle. Drug-treated female mice exhibited greater loss of LAT1 compared to male-drug treated mice. However, male-drug treated mice had greater decreases in BCAA concentrations compared to female drug-treated mice. In addition, minimal differences were found within sex for key enzymes involved in BCAA metabolism. However, the inhibitory phosphorylation of branched-chain alpha-ketoacid dehydrogenase complex (BCKD) was elevated only in drug-treated male animals. Lastly, within sex, drug treatment showed elevated levels of BCAA concentrations in the liver, but only male drug-treated mice showed elevated plasma BCAAs compared to male vehicle-treated mice. Our data suggests a link between muscle atrophy and decreased BCAA metabolism during chemotherapy. We found that drug-treated female mice had worsened outcomes for cachexia measures such as body, skeletal muscle weight and anabolic signalling compared to drug-treated male mice. However, drug-treated males exhibited greater decreases in BCAA concentrations and their metabolism compared to drug-treated females. Findings from this study suggest that altered availability and metabolism of the BCAAs may contribute to muscle wasting differently in males and females during chemotherapy and interventions that correct such alterations may need to consider sex in order to better manage cachexia. Nserc, York University This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract LB022: A novel first-in-class USP19 inhibitor for the treatment of cancer-induced muscle atrophy

Abstract Cancer cachexia is a metabolic wasting syndrome characterized by weight loss, anorexia and anemia as a result of tumor burden, and affects up to 80% of advanced cancer patients #1. Cachexia is particularly prevalent in pancreatic, lung, colorectal and gastro-intestinal cancers and can lead to reduced tolerance and responsiveness to chemotherapy, increased treatment-related toxicity and morbidity, and poor overall quality of life. There are currently no approved therapies for cancer cachexia. The development and maintenance of muscle tissue is dependent on the balance between protein synthesis and protein degradation, controlled through various anabolic and catabolic signaling pathways. Dysregulation of these pathways can result in muscle atrophy, which arises in many chronic illnesses. The ubiquitin proteasome system (UPS) has a central role in regulating skeletal muscle physiology. Previous work utilizing USP19 knock out mouse models has demonstrated that USP19 plays an important role in muscle wasting and can protect against denervation-induced muscle atrophy #2. We have previously demonstrated that inhibition of USP19 enzymatic activity spares the muscle wasting observed in limb-casted and denervated mouse models of muscle wasting. Here, we report the discovery of a novel, highly potent and selective inhibitor of USP19 (ADC-846) and demonstrate its utility in a cancer-induced muscle atrophy model in vivo. Pharmacological inhibition of USP19 by ADC-846 increased lean muscle and fat mass following oral dosing in a Lewis Lung Carcinoma-induced cachexia model and reduced the cachexic index by &amp;gt;60% compared to controls. This data, in combination with our previous work detailing the effect of USP19 inhibition on muscle force and function, provides a much-needed novel pharmacological strategy for therapeutic intervention in muscle wasting conditions. Citation Format: Natalie Page, Vignesh Karthikaisamy, Darren O'Hara, Aaron N. Cranston, Colin R. O'Dowd, Xavier Jacq, Richard Wilkinson, Stephanie Burton, Hayley Gibson, Joana Costa, Daniel Longley, Matthew Helm, Chris McGivern, Steven Shepherd, Christina Bell, Peter Hewitt, Mary McFarland, Hugues Miel, Steven Whitehead, Lauren Proctor, Shane J. Rountree, Mark Wappett, Mauricio Berriel Diaz, Stephan Herzig, Timothy Harrison. A novel first-in-class USP19 inhibitor for the treatment of cancer-induced muscle atrophy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB022.

Investigating the effect of Resistance Training on the Axis / IGF-1 PI3K in Skeletal Muscle and Insulin Sensitivity in Type 2 Diabetic Rats

Introduction: IGF-1 is a polypeptide hormone that plays a vital role in the development and regeneration of skeletal muscles.&#x0D; IGF-1 activates anabolic pathways such as PI3K and IRS-1/2 to activate a series of intracellular anabolic signals. The purpose of this study was to investigate the effect of 8 weeks resistance training on IGF1, PI3K and insulin sensitivity in neural muscle in type 2 diabetic rats.&#x0D; Methods: In this experimental study, 14 male Wistar mice aged 8 to 10 weeks and induction of type 2 diabetes (type 2 diabetes was induced by intraperitoneal injection of nicotinamide and STZ in diabetic groups) and were divided into two groups of resistance training and control groups. The mice in the control group did not exercise during the study, while the resistance group performed 8 weeks of training for 5 sessions per week. Resistance exercises in each session were performed in the form of 10 repetitions with 90-second rest intervals in the form of climbing a 26-step ladder to a height of one meter with a vertical slope of 85%, and weight was applied to the base of the rat's tail. 48 hours after the last training session, blood samples were taken from the heart and neural muscle tissue was extracted. Real Time-PCR method was used to determine the genes and one-way ANOVA test was used to compare the variables. All statistical studies were done using SPSS version 16 software.&#x0D; Results: The results of this study showed that expression of IGF-1 and PI3K in the duodenal axis of diabetic rats in resistance training group was significantly higher than that of the control group (P = 0.001) (P = 0.11). Serum insulin sensitivity in diabetic rats was not significantly different in diabetic rats compared to the diabetic rats (P=0.778).&#x0D; Conclusion: It seems that eight weeks of resistance training can be a strong stimulus on IGF-1 and PI3K gene expression and no change in insulin sensitivity in neural muscle in type 2 diabetic rats.

A Cell-Based Assessment of the Muscle Anabolic Potential of Blue Whiting (Micromesistius poutassou) Protein Hydrolysates.

Blue whiting (BW) represents an underutilised fish species containing a high-quality protein and amino acid (AA) profile with numerous potentially bioactive peptide sequences, making BW an economic and sustainable alternative source of protein. This study investigated the impact of three different BW protein hydrolysates (BWPH-X, Y and Z) on growth, proliferation and muscle protein synthesis (MPS) in skeletal muscle (C2C12) myotubes. BWPHs were hydrolysed using different enzymatic and heat exposures and underwent simulated gastrointestinal digestion (SGID), each resulting in a high degree of hydrolysis (33.41-37.29%) and high quantities of low molecular mass peptides (86.17-97.12% <1 kDa). C2C12 myotubes were treated with 1 mg protein equivalent/mL of SGID-BWPHs for 4 h. Muscle growth and myotube thickness were analysed using an xCelligence™ platform. Anabolic signalling (phosphorylation of mTOR, rpS6 and 4E-BP1) and MPS measured by puromycin incorporation were assessed using immunoblotting. BWPH-X significantly increased muscle growth (p < 0.01) and myotube thickness (p < 0.0001) compared to the negative control (amino acid and serum free media). Muscle protein synthesis (MPS), as measured by puromycin incorporation, was significantly higher after incubation with BWPH-X compared with the negative control, but did not significantly change in response to BWPH-Y and Z treatments. Taken together, these preliminary findings demonstrate the anabolic potential of some but not all BWPHs on muscle enhancement, thus providing justification for human dietary intervention studies to confirm and translate the results of such investigations to dietary recommendations and practices.

Characterization of sclerostin's response within white adipose tissue to an obesogenic diet at rest and in response to acute exercise in male mice.

Introduction: It is well established that sclerostin antagonizes the anabolic Wnt signalling pathway in bone, however, its physiological role in other tissues remains less clear. This study examined the effect of a high-fat diet (HFD) on sclerostin content and downstream markers of the Wnt signaling pathway (GSK3β and β-catenin) within subcutaneous inguinal white adipose tissue (iWAT), and visceral epididymal WAT (eWAT) depots at rest and in response to acute aerobic exercise. Methods: Male C57BL/6 mice (n = 40, 18weeks of age) underwent 10 weeks of either a low-fat diet (LFD) or HFD. Within each diet group, mice were assigned to either remain sedentary (SED) or perform 2h of endurance treadmill exercise at 15mmin-1 with 5° incline (EX), creating four groups: LFD + SED (N = 10), LFD + EX (N = 10), HFD + SED (N = 10), and HFD + EX (N = 10). Serum and WAT depots were collected 2h post-exercise. Results: Serum sclerostin showed a diet-by-exercise interaction, reflecting HFD + EX mice having higher concentration than HFD + SED (+31%, p = 0.03), and LFD mice being unresponsive to exercise. iWAT sclerostin content decreased post-exercise in both 28kDa (-31%, p = 0.04) and 30kDa bands (-36%, main effect for exercise, p = 0.02). iWAT β-catenin (+44%, p = 0.03) and GSK3β content were higher in HFD mice compared to LFD (+128%, main effect for diet, p = 0.005). Monomeric sclerostin content was abolished in eWAT of HFD mice (-96%, main effect for diet, p < 0.0001), was only detectable as a 30kDa band in LFD mice and was unresponsive to exercise. β-catenin and GSK3β were both unresponsive to diet and exercise within eWAT. Conclusion: These results characterized sclerostin's content to WAT depots in response to acute exercise, which appears to be specific to a reduction in iWAT and identified a differential regulation of sclerostin's form/post-translational modifications depending on diet and WAT depot.

The dose-response effects of arachidonic acid on primary human skeletal myoblasts and myotubes.

Cellular inflammatory response, mediated by arachidonic acid (AA) and cyclooxygenase, is a highly regulated process that leads to the repair of damaged tissue. Recent studies on murine C2C12 cells have demonstrated that AA supplementation leads to myotube hypertrophy. However, AA has not been tested on primary human muscle cells. Therefore, the purpose of this study was to determine whether AA supplementation has similar effects on human muscle cells. Proliferating and differentiating human myoblasts were exposed to AA in a dose-dependent manner (50-0.80µM) for 48 (myoblasts) or 72 (myotubes) hours. Cell viability was tested using a 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay and cell counting; myotube area was determined by immunocytochemistry and confocal microscopy; and anabolic signaling pathways were evaluated by western blot and RT-PCR. Our data show that the treatment of primary human myoblasts treated with 50µM and 25µM of AA led to the release of PGE2 and PGF2α at levels higher than those of control-treated cells (p <0.001 for all concentrations). Additionally, 50µM and 25µM of AA suppressed myoblast proliferation, myotube area, and myotube fusion. Anabolic signaling indicated reductions in total and phosphorylated TSC2, AKT, S6, and 4EBP1 in myoblasts at 50µM of AA (p <0.01 for all), but not in myotubes. These changes were not affected by COX-2 inhibition with celecoxib. Together, our data demonstrate that high concentrations of AA inhibit myoblast proliferation, myotube fusion, and myotube hypertrophy, thus revealing potential deleterious effects of AA on human skeletal muscle cell health and viability.

THE ROLE OF PPARδ-DRIVEN β-OXIDATION IN BONE HEALTH DURING AGING

Abstract Musculoskeletal disorders are a significant complication of aging, leading to increased morbidity and mortality. However, current understanding of the mechanisms by which aging affects skeletal health is limited. Osteocytes are the most numerous and long-lived bone cells and play key roles in maintaining bone mass by responding to anabolic signals such as mechanical loading. Energy metabolism is dysregulated in many cells with aging, however regulation of energy metabolism in osteocytes and how this is affected during aging and by mechanical loading remains undefined. To investigate this, we first used IDG-SW3 osteocyte cells to determine the effects of mechanical loading on osteocytes in vitro by applying fluid flow shear stress (FFSS). FFSS increased Pparδ and Cpt1 expression, key promoters of fatty acid β-oxidation (FAO). Pharmacological antagonism of PPARδ or CPT1 resulted in dysregulated expression of key bone remodeling genes and impaired ATP release in response to FFSS. In vivo, mechanical loading significantly increased FAO in tibia cortical bone. However, FAO was impaired in the bones from aged mice. To further elucidate the role of osteocyte FAO, we deleted PPARδ specifically in osteocytes (PPARδ cKO), which resulted in decreased FAO and bone volume in female PPARδ cKO mice. Lastly, treatment of aging mice with the PPARδ activator GW0742 resulted in significantly increased bone mineral content, density and trabecular bone volume. These findings suggest important functions of osteocyte energy metabolism during aging and with mechanical loading on bone and identify PPARδ-driven FAO as a novel therapeutic target for improving skeletal health with aging.

Intensive Care Unit Acquired Weakness Is Associated with Rapid Changes to Skeletal Muscle Proteostasis.

Intensive care unit (ICU)-acquired weakness is a frequent consequence of critical illness that impacts both the limb and respiratory muscles. The cause of ICU-acquired weakness is multifactorial, but both prolonged limb muscle inactivity and mechanical ventilation are risk factors for muscle wasting, which predisposes ICU patients to both short-term complications and long-term disabilities resulting from muscle weakness. Unfortunately, the current research does not provide a detailed understanding of the cellular etiology of ICU-acquired weakness, and no standard treatment exists. Therefore, improving knowledge of the mechanisms promoting muscle atrophy in critically ill patients is essential to developing therapeutic strategies to protect against ICU-induced skeletal muscle wasting. To advance our understanding of the mechanism(s) responsible for ICU-acquired weakness, we tested the hypothesis that ICU-induced muscle inactivity promotes a rapid decrease in anabolic signaling/protein synthesis and accelerates proteolysis in both limb and respiratory muscles. To investigate ICU-induced changes in skeletal muscle proteostasis, adult Sprague Dawley rats were anesthetized and mechanically ventilated for 12 h to simulate ICU care. Measurements of anabolic signaling, protein synthesis, and proteolytic activity in the limb muscles (plantaris and soleus) and respiratory muscles (parasternal and intercostal) revealed ICU-induced reductions in both anabolic signaling (i.e., AKT/mTOR pathway) and muscle protein synthesis. Moreover, simulated ICU care resulted in increased biomarkers of accelerated proteolysis in both limb and respiratory muscles. These novel findings reveal that disturbances in limb and respiratory muscle proteostasis occur rapidly during ICU-induced muscle inactivity, irrespective of the muscle function or muscle fiber type.

Depressed Protein Synthesis and Anabolic Signaling Potentiate ACL Tear–Resultant Quadriceps Atrophy

Background: Anterior cruciate ligament (ACL) tear (ACLT) leads to protracted quadriceps muscle atrophy. Protein turnover largely dictates muscle size and is highly responsive to injury and loading. Regulation of quadriceps molecular protein synthetic machinery after ACLT has largely been unexplored, limiting development of targeted therapies. Purpose: To define the effect of ACLT on (1) the activation of protein synthetic and catabolic signaling within quadriceps biopsy specimens from human participants and (2) the time course of alterations to protein synthesis and its molecular regulation in a mouse ACL injury model. Study Design: Descriptive laboratory study. Methods: Muscle biopsy specimens were obtained from the ACL-injured and noninjured vastus lateralis of young adult humans after an overnight fast (N = 21; mean ± SD, 19 ± 5 years). Mice had their limbs assigned to ACLT or control, and whole quadriceps were collected 6 hours or 1, 3, or 7 days after injury with puromycin injected before tissue collection for assessment of relative protein synthesis. Muscle fiber size and expression and phosphorylation of protein anabolic and catabolic signaling proteins were assessed at the protein and transcript levels (RNA sequencing). Results: Human quadriceps showed reduced phosphorylation of ribosomal protein S6 (–41%) in the ACL-injured limb (P = .008), in addition to elevated phosphorylation of eukaryotic initiation factor 2α (+98%; P = .006), indicative of depressed protein anabolic signaling in the injured limb. No differences in E3 ubiquitin ligase expression were noted. Protein synthesis was lower at 1 day (P = .01 vs control limb) and 3 days (P = .002 vs control limb) after ACLT in mice. Pathway analyses revealed shared molecular alterations between human and mouse quadriceps after ACLT. Conclusion: (1) Global protein synthesis and anabolic signaling deficits occur in the quadriceps in response to ACL injury, without notable changes in measured markers of muscle protein catabolism. (2) Importantly, these deficits occur before the onset of significant atrophy, underscoring the need for early intervention. Clinical Relevance: These findings suggest that blunted protein anabolism as opposed to increased catabolism likely mediates quadriceps atrophy after ACL injury. Thus, future interventions should aim to restore muscle protein anabolism rapidly after ACLT.

Evaluation of the Timing of Use of Phosphatidic Acid in the Diet on Growth Performance and Breast Meat Yield in Broilers.

With a growing increase in further processing of poultry, there has been an increased interest in factors, including feed additives, that may improve broiler performance, increase growth, and influence dressing percentage. Mammalian target of rapamycin (mTOR) is known to play vital roles in protein synthesis; mTOR controls the anabolic and catabolic signaling of skeletal muscle mass, resulting in the modulation of muscle hypertrophy. Exogenous phosphatidic acid (PA) can stimulate the mTOR pathway via its activation of the substrate S6 kinase. A study with 648 Cobb 500 male broilers, housed in 36 floor pens (1.11 m2) from 1 to 42 days of age was conducted to evaluate the timing of PA (Mediator® 50P, Chemi Nutra, Austin, TX, USA) supplementation on the growth performance and carcass yield of broilers. Dietary treatments included T1, Control (CON), T2, 5 mg/bird/day of PA for 42 days (d0-42, PAA); T3, 5 mg/bird/day of PA for 28 days (d15-42, PAGF); and T4, 5 mg/bird/day of PA for 14 days (d29-42, PAF). All birds were weighed on d14, 28, and 42 to obtain BW (body weight), FCR (feed conversion ratio), and MORT (mortality percentage). On d42, eight birds per pen were processed to determine carcass and breast meat yield. No differences were observed in BW at d14 or d28. At d42, birds fed PAA were heavier (3.73 ± 0.02, p &lt; 0.05) than all dietary treatments (3.68 ± 0.02). From d0 to d28, birds fed PAA had the lowest FCR (1.423 ± 0.005, p &lt; 0.05) compared to all dietary treatments (1.441 ± 0.005). From d0 to d42, birds fed PAA and PAGF had a lower FCR (1.545 ± 0.014, p &lt; 0.05) when compared to the CON (1.609 ± 0.013). No differences were observed in MORT between treatments during growout. Increased BW observed in birds fed PAA translated to increased breast fillet weight (0.772 ± 0.009 kg, p &lt; 0.05) when compared to the CON (0.743 ± 0.008 kg). Carcass yields were increased in birds fed PAA (77.48 ± 0.32 kg, p &lt; 0.05) when compared to all dietary treatments (76.24 ± 0.16 kg). Utilizing PA for 42 days increased live weights, improved FCR, increased carcass yield, and increased breast fillet weight at processing. Results from this study indicate that supplementation of PA during all phases of growth may increase the production efficiency of broilers.

Combining Exercise and Carbohydrate Restriction in Patients with Type 2 Diabetes Mellitus—A Critical Look at Possible Intervention Effects

Combining regular exercise and a healthy diet is recommended in international guidelines to fight type 2 diabetes mellitus (T2DM). Low- and very low-carbohydrate diets have attracted attention in the last years. This article takes a critical look at the possible effects when regular exercise and carbohydrate restriction are combined. An increased intervention effect on the oxidative capacity as well as glucose and lipid profiles can be assumed (at least for a short period of time). However, anabolic signaling pathways might be blunted during a very low-carbohydrate diet and increasing ketosis. Thus, muscle build-up can become difficult or impossible. Furthermore, maximal performance during high-intensity workouts may be attenuated due to a possible reduced anaerobic glycolysis and metabolic inflexibility in T2DM patients. However, more studies are needed to evaluate the effects of this combination in comparison to those of exercise and other types of diet.

Whole-genome RNAsequencing identifies distinct transcriptomic profiles in impingement cartilage between patients with femoroacetabular impingement and hip osteoarthritis.

Femoroacetabular impingement (FAI) has a strong clinical association with the development of hip osteoarthritis (OA); however, the pathobiological mechanisms underlying the transition from focal impingement to global joint degeneration remain poorly understood. The purpose of this study is to use whole-genome RNA sequencing to identify and subsequently validate differentially expressed genes (DEGs) in femoral head articular cartilage samples from patients with FAI and hip OA secondary to FAI. Thirty-seven patients were included in the study with whole-genome RNA sequencing performed on 10 gender-matched patients in the FAI and OA cohorts and the remaining specimens were used for validation analyses. We identified a total of 3531 DEGs between the FAI and OA cohorts with multiple targets for genes implicated in canonical OA pathways.Quantitative reverse transcription-polymerase chain reaction validation confirmed increased expression of FGF18 and WNT16 in the FAI samples, while there was increased expression of MMP13 and ADAMTS4 in the OA samples. Expression levels of FGF18 and WNT16 were also higher in FAI samples with mild cartilage damage compared to FAI samples with severe cartilage damage or OA cartilage. Our study further expands the knowledge regarding distinct genetic reprogramming in the cartilage between FAI and hip OA patients. We independently validated the results of the sequencing analysis and found increased expression of anabolic markers in patients with FAI and minimal histologic cartilage damage, suggesting that anabolic signaling may be increased in early FAI with a transition to catabolic and inflammatory gene expression as FAI progresses towards more severe hip OA.Clinical significance:Cam-typeFAI has a strong clinical association with hip OA; however, the cellular pathophysiology of disease progression remains poorly understood. Several previous studies have demonstrated increased expression of inflammatory markers in FAI cartilage samples, suggesting the involvement of these inflammatory pathways in the disease progression. Our study further expands the knowledge regarding distinct genetic reprogramming in the cartilage between FAI and hip OA patients. In addition to differences in inflammatory gene expression, we also identified differential expression in multiple pathways involved in hip OA progression.

Angiopoietin-like 3-derivative LNA043 for cartilage regeneration in osteoarthritis: a randomized phase 1 trial

Osteoarthritis (OA) is a common, debilitating, chronic disease with no disease-modifying drug approved to date. We discovered LNA043—a derivative of angiopoietin-like 3 (ANGPTL3)—as a potent chondrogenesis inducer using a phenotypic screen with human mesenchymal stem cells. We show that LNA043 promotes chondrogenesis and cartilage matrix synthesis in vitro and regenerates hyaline articular cartilage in preclinical OA and cartilage injury models in vivo. LNA043 exerts at least part of these effects through binding to the fibronectin receptor, integrin α5β1 on mesenchymal stem cells and chondrocytes. In a first-in-human (phase 1), randomized, double-blinded, placebo-controlled, single ascending dose, single-center trial (NCT02491281; sponsored by Novartis Pharmaceuticals), 28 patients with knee OA were injected intra-articularly with LNA043 or placebo (3:1 ratio) either 2 h, 7 d or 21 d before total knee replacement. LNA043 met its primary safety endpoint and showed short serum pharmacokinetics, cartilage penetration and a lack of immunogenicity (secondary endpoints). Post-hoc transcriptomics profiling of cartilage revealed that a single LNA043 injection reverses the OA transcriptome signature over at least 21 d, inducing the expression of hyaline cartilage matrix components and anabolic signaling pathways, while suppressing mediators of OA progression. LNA043 is a novel disease-modifying OA drug candidate that is currently in a phase 2b trial (NCT04864392) in patients with knee OA.

Inter-set stretch: A potential time-efficient strategy for enhancing skeletal muscle adaptations.

Time is considered a primary barrier to exercise adherence. Therefore, developing time-efficient resistance training (RT) strategies that optimize muscular adaptations is of primary interest to practitioners. A novel approach to the problem involves combining intensive stretch protocols with RT. Conceivably, integrating stretch into the inter-set period may provide an added stimulus for muscle growth without increasing session duration. Mechanistically, stretch can regulate anabolic signaling via both active and passive force sensors. Emerging evidence indicates that both lengthening contractions against a high load as well as passive stretch can acutely activate anabolic intracellular signaling pathways involved in muscle hypertrophy. Although longitudinal research investigating the effects of stretching between RT sets is limited, some evidence suggests it may in fact enhance hypertrophic adaptations. Accordingly, the purpose of this paper is threefold: (1) to review how the active force of a muscle contraction and the force of a passive stretched are sensed; (2) to present evidence for the effectiveness of RT with inter-set stretch for muscle hypertrophy (3) to provide practical recommendations for application of inter-set stretch in program design as well as directions for future research.

Nicotinamide riboside kinases regulate skeletal muscle fiber-type specification and are rate-limiting for metabolic adaptations during regeneration.

Nicotinamide riboside kinases (NRKs) control the conversion of dietary Nicotinamide Riboside (NR) to NAD+, but little is known about their contribution to endogenous NAD+ turnover and muscle plasticity during skeletal muscle growth and remodeling. Using NRK1/2 double KO (NRKdKO) mice, we investigated the influence of NRKs on NAD+ metabolism and muscle homeostasis, and on the response to neurogenic muscle atrophy and regeneration following muscle injury. Muscles from NRKdKO animals have altered nicotinamide (NAM) salvage and a decrease in mitochondrial content. In single myonuclei RNAseq of skeletal muscle, NRK2 mRNA expression is restricted to type IIx muscle fibers, and perturbed NAD+ turnover and mitochondrial metabolism shifts the fiber type composition of NRKdKO muscle to fast glycolytic IIB fibers. NRKdKO does not influence muscle atrophy during denervation but alters muscle repair after myofiber injury. During regeneration, muscle stem cells (MuSCs) from NRKdKO animals hyper-proliferate but fail to differentiate. NRKdKO also alters the recovery of NAD+ during muscle regeneration as well as mitochondrial adaptations and extracellular matrix remodeling required for tissue repair. These metabolic perturbations result in a transient delay of muscle regeneration which normalizes during myofiber maturation at late stages of regeneration via over-compensation of anabolic IGF1-Akt signaling. Altogether, we demonstrate that NAD+ synthesis controls mitochondrial metabolism and fiber type composition via NRK1/2 and is rate-limiting for myogenic commitment and mitochondrial maturation during skeletal muscle repair.

Interleukin-6 initiates muscle- and adipose tissue wasting in a novel C57BL/6 model of cancer-associated cachexia.

Cancer-associated cachexia (CAC) is a wasting syndrome drastically reducing efficacy of chemotherapy and life expectancy of patients. CAC affects up to 80% of cancer patients, yet the mechanisms underlying the disease are not well understood and no approved disease-specific medication exists. As a multiorgan disorder, CAC can only be studied on an organismal level. To cover the diverse aetiologies of CAC, researchers rely on the availability of a multifaceted pool of cancer models with varying degrees of cachexia symptoms. So far, no tumour model syngeneic to C57BL/6 mice exists that allows direct comparison between cachexigenic- and non-cachexigenic tumours. MCA207 and CHX207 fibrosarcoma cells were intramuscularly implanted into male or female, 10-11-week-old C57BL/6J mice. Tumour tissues were subjected to magnetic resonance imaging, immunohistochemical-, and transcriptomic analysis. Mice were analysed for tumour growth, body weight and -composition, food- and water intake, locomotor activity, O2 consumption, CO2 production, circulating blood cells, metabolites, and tumourkines. Mice were sacrificed with same tumour weights in all groups. Adipose tissues were examined using high-resolution respirometry, lipolysis measurements in vitro and ex vivo, and radioactive tracer studies in vivo. Gene expression was determined in adipose- and muscle tissues by quantitative PCR and Western blotting analyses. Muscles and cultured myotubes were analysed histologically and by immunofluorescence microscopy for myofibre cross sectional area and myofibre diameter, respectively. Interleukin-6 (Il-6) was deleted from cancer cells using CRISPR/Cas9 mediated gene editing. CHX207, but not MCA207-tumour-bearing mice exhibited major clinical features of CAC, including systemic inflammation, increased plasma IL-6 concentrations (190pg/mL, P≤0.0001), increased energy expenditure (+28%, P≤0.01), adipose tissue loss (-47%, P≤0.0001), skeletal muscle wasting (-18%, P≤0.001), and body weight reduction (-13%, P≤0.01) 13days after cancer cell inoculation. Adipose tissue loss resulted from reduced lipid uptake and -synthesis combined with increased lipolysis but was not associated with elevated beta-adrenergic signalling or adipose tissue browning. Muscle atrophy was evident by reduced myofibre cross sectional area (-21.8%, P≤0.001), increased catabolic- and reduced anabolic signalling. Deletion of IL-6 from CHX207 cancer cells completely protected CHX207IL6KO -tumour-bearing mice from CAC. In this study, we present CHX207 fibrosarcoma cells as a novel tool to investigate the mediators and metabolic consequences of CAC in C57BL/6 mice in comparison to non-cachectic MCA207-tumour-bearing mice. IL-6 represents an essential trigger for CAC development in CHX207-tumour-bearing mice.

The atypical β-blocker S-oxprenolol reduces cachexia and improves survival in a rat cancer cachexia model.

Beta-blockers and selected stereoisomers of beta-blockers, like bisoprolol and S-pindolol (ACM-001), have been shown to be effective in preclinical cancer cachexia models. Here, we tested the efficacy of stereoisomers of oxprenolol in two preclinical models of cancer cachexia-the Yoshida AH-130 rat model and the Lewis lung carcinoma (LLC) mouse model. In the Yoshida AH130 hepatoma rat cancer cachexia model and compared with placebo, 50mg/kg/d S-oxprenolol (HR: 0.49, 95% CI: 0.28-0.85, P=0.012) was superior to 50mg/kg/d R-oxprenolol (HR: 0.83, 95% CI 0.38-1.45, P=0.51) in reducing mortality (=reaching ethical endpoints). Combination of the three doses (12.5, 25 and 50mg/kg/d) that had a significant effect on body weight loss in the S-oxprenolol groups vs the same combination of the R-oxprenolol groups lead to a significantly improved survival of S-oxprenolol vs R-oxprenolol (HR: 1.61, 95% CI: 1.08-2.39, P=0.0185). Interestingly, there is a clear dose dependency in S-oxprenolol-treated (5, 12.5, 25 and 50mg/kg/d) groups, which was not observed in groups treated with R-oxprenolol. A dose-dependent attenuation of weight and lean mass loss by S-oxprenolol was seen in the Yoshida rat model, whereas R-oxprenolol had only had a significant effect on fat mass. S-oxprenolol also non-significantly reduced weight loss in the LLC model and also improved muscle function (grip strength 428±25 and 539±37g/100g body weight for placebo and S-oxprenolol, respectively). However, there was only a minor effect on quality of life indicators food intake and spontaneous activity in the Yoshida model (25mg/kg/S-oxprenolol: 11.9±2.5g vs placebo: 4.9±0.8g, P=0.013 and also vs 25mg/kg/d R-oxprenolol: 7.5±2.6g, P=0.025). Both enantiomers had no effects on cardiac dimensions and function at the doses used in this study. Western blotting of proteins involved in the anabolic/catabolic homoeostasis suggest that anabolic signalling is persevered (IGF-1 receptor, Akt) and catabolic signalling is inhibited (FXBO-10, TRAF-6) by S-pindolol, but not he R-enantiomer. Expression of glucose transporters Glut1 and Glut 4 was similar in all groups, as was AMPK. S-oxprenolol is superior to R-oxprenolol in cancer cachexia animal models and shows promise for a human application in cancer cachexia.