bims-moremu 2022-05-22 papers

bims-moremu

Biomed News

on Molecular regulators of muscle mass

Issue of 2022‒05‒22
thirty-one papers selected by
Anna Vainshtein
Craft Science Inc.

Alterations of Lysine Acetylation Profile in Murine Skeletal Muscles Upon Exercise.
A Simple and Inexpensive Running Wheel Model for Progressive Resistance Training in Mice.
Conservative analysis of Synaptopodin-2 intron sense-overlapping lncRNA reveals its novel function in promoting muscle atrophy.
Ambra1 deficiency impairs mitophagy in skeletal muscle.
Selenium Deficiency Leads to Reduced Skeletal Muscle Cell Differentiation by Oxidative Stress in Mice.
DVL1 and DVL3 require nuclear localisation to regulate proliferation in human myoblasts.
Heat shock protein A4 ablation leads to skeletal muscle myopathy associated with dysregulated autophagy and induced apoptosis.
Effect of 32-Weeks High-Intensity Interval Training and Resistance Training on Delaying Sarcopenia: Focus on Endogenous Apoptosis.
Maternal vitamin D deficiency affects the morphology and function of glycolytic muscle in adult offspring rats.
Sex-Based Differences in the Myogenic Response and Inflammatory Gene Expression Following Eccentric Contractions in Humans.
CTRP11 contributes modestly to systemic metabolism and energy balance.
Acute bioenergetic insulin sensitivity of skeletal muscle cells: ATP-demand-provoked glycolysis contributes to stimulation of ATP supply.
The Vitamin D/Vitamin D receptor (VDR) axis in muscle atrophy and sarcopenia.
A drug and ATP binding site in type 1 ryanodine receptor.
MEF2C Expression Is Regulated by the Post-transcriptional Activation of the METTL3-m6A-YTHDF1 Axis in Myoblast Differentiation.
Benefits of aerobic exercise in myotonic dystrophy type 1.
Fluid shear stress-induced mechanotransduction in myoblasts: Does it depend on the glycocalyx?
Piezo1: opening the way to preventing muscle atrophy.
The cell polarity protein Vangl2 in the muscle shapes the neuromuscular synapse by binding to and regulating the tyrosine kinase MuSK.
A pathogenic mechanism associated with myopathies and structural birth defects involves TPM2 directed myogenesis.
Fasting induces a highly resilient deep quiescent state in muscle stem cells via ketone body signaling.
Docosahexaenoic acid ester of phloridzin reduces inflammation and insulin resistance via AMPK.
Aerobic exercise training-induced follistatin-like 1 secretion in the skeletal muscle is related to arterial stiffness via arterial NO production in obese rats.
What Can We Learn from Single Sarcomere and Myofibril Preparations?
Specialized androgen synthesis in skeletal muscles that actuate elaborate social displays.
Slow motor neurons resist pathological TDP-43 and mediate motor recovery in the rNLS8 model of amyotrophic lateral sclerosis.
Rho-kinase inhibition improves haemodynamic responses and circulating ATP during hypoxia and moderate intensity handgrip exercise in healthy older adults.
Combined overexpression of ATXN1L and mutant ATXN1 knockdown by AAV rescue motor phenotypes and gene signatures in SCA1 mice.
Heterozygous SOD2 deletion selectively impairs SERCA function in the soleus of female mice.
Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults.
Residual force enhancement is affected more by quadriceps muscle length than stretch amplitude.

Front Aging Neurosci. 2022 ;14 859313

Alterations of Lysine Acetylation Profile in Murine Skeletal Muscles Upon Exercise.

Dehuan Liang, Cheng Chen, Song Huang, Sujuan Liu, Li Fu, Yanmei Niu.

  Objective: Regular exercise is a powerful tool that enhances skeletal muscle mass and strength. Lysine acetylation is an important post-translational modification (PTM) involved in a broad array of cellular functions. Skeletal muscle protein contains a considerable number of lysine-acetylated (Kac) sites, so we aimed to investigate the effects of exercise-induced lysine acetylation on skeletal muscle proteins.Methods: We randomly divided 20 male C57BL/6 mice into exercise and control groups. After 6 weeks of treadmill exercise, a lysine acetylation proteomics analysis of the gastrocnemius muscles of mice was performed.
Results: A total of 2,254 lysine acetylation sites in 693 protein groups were identified, among which 1,916 sites in 528 proteins were quantified. The enrichment analysis suggested that protein acetylation could influence both structural and functional muscle protein properties. Moreover, molecular docking revealed that mimicking protein deacetylation primarily influenced the interaction between substrates and enzymes.
Conclusion: Exercise-induced lysine acetylation appears to be a crucial contributor to the alteration of skeletal muscle protein binding free energy, suggesting that its modulation is a potential approach for improving exercise performance.

Keywords:  aerobic exercise; bioinformatics analysis; lysine acetylation; muscle; proteins

DOI:  https://doi.org/10.3389/fnagi.2022.859313
J Vis Exp. 2022 Apr 28.

A Simple and Inexpensive Running Wheel Model for Progressive Resistance Training in Mice.

Pieter J Koopmans, Kevin A Zwetsloot.

Previously developed rodent resistance-based exercise models, including synergistic ablation, electrical stimulation, weighted-ladder climbing, and most recently, weighted-sled pulling, are highly effective at providing a hypertrophic stimulus to induce skeletal muscle adaptations. While these models have proven invaluable for skeletal muscle research, they are either invasive or involuntary and labor-intensive. Fortunately, many rodent strains voluntarily run long distances when given access to a running wheel. Loaded wheel running (LWR) models in rodents are capable of inducing adaptations commonly observed with resistance training in humans, such as increased muscle mass and fiber hypertrophy, as well as stimulation of muscle protein synthesis. However, the addition of moderate wheel load either fails to deter mice from running great distances, which is more reflective of an endurance/resistance training model, or the mice discontinue running nearly entirely due to the method of load application. Therefore, a novel high-load wheel running model (HLWR) has been developed for mice where external resistance is applied and progressively increased, enabling mice to continue running with much higher loads than previously utilized. Preliminary results from this novel HLWR model suggest it provides sufficient stimulus to induce hypertrophic adaptations over the 9 week training protocol. Herein, the specific procedures to execute this simple yet inexpensive progressive resistance-based exercise training model in mice are described.

DOI: https://doi.org/10.3791/63933
J Cachexia Sarcopenia Muscle. 2022 May 19.

Conservative analysis of Synaptopodin-2 intron sense-overlapping lncRNA reveals its novel function in promoting muscle atrophy.

Jianjun Jin, Mengmeng Du, Jian Wang, Yubo Guo, Jiali Zhang, Hao Zuo, Yunqing Hou, Shanshan Wang, Wei Lv, Wei Bai, Jin Wang, Xizhen Zhan, Yaxin Peng, Qian Tong, Jin Chai, Zaiyan Xu, Bo Zuo.

  BACKGROUND: Dissection of the regulatory pathways that control skeletal muscle development and atrophy is important for the treatment of muscle wasting. Long noncoding RNA (lncRNA) play important roles in various stages of muscle development. We previously reported that Synaptopodin-2 (SYNPO2) intron sense-overlapping lncRNA (SYISL) regulates myogenesis through an interaction with enhancer of zeste homologue 2 (EZH2). However, it remains unclear whether SYISL homologues exist in humans and pigs, and whether the functions and mechanisms of these homologues are conserved among species.METHODS: Bioinformatics, cell fractionation, and quantitative real-time polymerase chain reaction (qRT-PCR) analyses were used for the identification and molecular characterization of SYISL homologues in humans and pigs. Effects on myogenesis and muscle atrophy were determined via loss-of-function or gain-of-function experiments using C2C12 myoblasts, myogenic progenitor cells, dexamethasone (DEX), and aging-induced muscle atrophy models. RNA pulldown, RNA immunoprecipitation, dual luciferase reporting, and co-transfection experiments were used to explore the mechanisms of SYISL interactions with proteins and miRNAs.
RESULTS: We identified SYISL homologues in humans (designated hSYISL) and pigs (designated pSYISL). Functional experiments demonstrated that hSYISL and pSYISL regulate myogenesis through interactions with EZH2. Interestingly, we showed that SYISL functions to regulate muscle atrophy and sarcopenia through comparative analysis. SYISL is significantly up-regulated after muscle atrophy (P < 0.01); it significantly promotes muscle atrophy in DEX-induced muscle atrophy models (P < 0.01). SYISL knockdown or knockout alleviates muscle atrophy and sarcopenia in DEX-induced and aged mice. The tibialis anterior (TA) muscle weight of 3-month-old wild-type (WT) mice decreased by 33.24% after DEX treatment (P < 0.001), while the muscle weight loss of 3-month-old SYISL knockout mice was only 18.20% after DEX treatment (P < 0.001). SYISL knockout in 18-month-old WT mice significantly increased the weights of quadriceps (Qu), gastrocnemius (Gas), and TA muscles by 10.45% (P < 0.05), 13.95% (P < 0.01), and 24.82% (P < 0.05), respectively. Mechanistically, SYISL increases the expression levels of the muscle atrophy genes forkhead box protein O3a (FoxO3a), muscle ring finger 1 (MuRF1), and muscle atrophy-related F-box (Atrogin-1) via sponging of miR-23a-3p/miR-103-3p/miR-205-5p and thus promotes muscle atrophy. Additionally, we verified that human SYISL overexpression in muscles of 18-month-old WT mice significantly decreased the weights of Gas, Qu, and TA muscles by 7.76% (P < 0.01), 12.26% (P < 0.05), and 13.44% (P < 0.01), respectively, and accelerates muscle atrophy through conserved mechanisms.
CONCLUSIONS: Our results identify SYISL as a conserved lncRNA that modulates myogenesis in mice, pigs, and humans. We also demonstrated its previously unknown ability to promote muscle atrophy.

Keywords:  Muscle atrophy; Myogenesis; SYISL; miRNA sponging

DOI:  https://doi.org/10.1002/jcsm.13012
J Cachexia Sarcopenia Muscle. 2022 May 20.

Ambra1 deficiency impairs mitophagy in skeletal muscle.

Lisa Gambarotto, Samuele Metti, Martina Chrisam, Cristina Cerqua, Patrizia Sabatelli, Andrea Armani, Carlo Zanon, Marianna Spizzotin, Silvia Castagnaro, Flavie Strappazzon, Paolo Grumati, Matilde Cescon, Paola Braghetta, Eva Trevisson, Francesco Cecconi, Paolo Bonaldo.

  BACKGROUND: Maintaining healthy mitochondria is mandatory for muscle viability and function. An essential surveillance mechanism targeting defective and harmful mitochondria to degradation is the selective form of autophagy called mitophagy. Ambra1 is a multifaceted protein with well-known autophagic and mitophagic functions. However, the study of its role in adult tissues has been extremely limited due to the embryonic lethality caused by full-body Ambra1 deficiency.METHODS: To establish the role of Ambra1 as a positive regulator of mitophagy, we exploited in vivo overexpression of a mitochondria-targeted form of Ambra1 in skeletal muscle. To dissect the consequence of Ambra1 inactivation in skeletal muscle, we generated muscle-specific Ambra1 knockout (Ambra1fl/fl :Mlc1f-Cre) mice. Mitochondria-enriched fractions were obtained from muscles of fed and starved animals to investigate the dynamics of the mitophagic flux.
RESULTS: Our data show that Ambra1 has a critical role in the mitophagic flux of adult murine skeletal muscle and that its genetic inactivation leads to mitochondria alterations and myofibre remodelling. Ambra1 overexpression in wild-type muscles is sufficient to enhance mitochondria clearance through the autophagy-lysosome system. Consistently with this, Ambra1-deficient muscles display an abnormal accumulation of the mitochondrial marker TOMM20 by +76% (n = 6-7; P < 0.05), a higher presence of myofibres with swollen mitochondria by +173% (n = 4; P < 0.05), and an alteration in the maintenance of the mitochondrial membrane potential and a 34% reduction in the mitochondrial respiratory complex I activity (n = 4; P < 0.05). Lack of Ambra1 in skeletal muscle leads to impaired mitophagic flux, without affecting the bulk autophagic process. This is due to a significantly decreased recruitment of DRP1 (n = 6-7 mice; P < 0.01) and Parkin (n = 6-7 mice; P < 0.05) to the mitochondrial compartment, when compared with controls. Ambra1-deficient muscles also show a marked dysregulation of the endolysosome compartment, as the incidence of myofibres with lysosomal accumulation is 20 times higher than wild-type muscles (n = 4; P < 0.05). Histologically, Ambra1-deficient muscles of both 3- and 6-month-old animals display a significant decrease of myofibre cross-sectional area and a 52% reduction in oxidative fibres (n = 6-7; P < 0.05), thus highlighting a role for Ambra1 in the proper structure and activity of skeletal muscle.
CONCLUSIONS: Our study indicates that Ambra1 is critical for skeletal muscle mitophagy and for the proper maintenance of functional mitochondria.

Keywords:  Ambra1; Mitochondria; Mitophagy; Skeletal muscle

DOI:  https://doi.org/10.1002/jcsm.13010
Biol Trace Elem Res. 2022 May 16.

Selenium Deficiency Leads to Reduced Skeletal Muscle Cell Differentiation by Oxidative Stress in Mice.

Bo-Wen Bao, Zibo Kang, Yu Zhang, Kan Li, Ran Xu, Meng-Yao Guo.

  Selenium (Se) is one of the essential trace elements in animal organisms with good antioxidant and immune-enhancing abilities. In this study, we investigated the effect and mechanism of Se deficiency on skeletal muscle cell differentiation. A selenium-deficient skeletal muscle model was established. The skeletal muscle tissue and blood Se content were significantly reduced in the Se deficiency group. HE staining showed that the skeletal muscle tissue had a reduced myofiber area and nuclei and an increased myofascicular membrane with Se deficiency. The TUNEL test showed massive apoptosis of skeletal muscle cells in Se deficiency. With Se deficiency, reactive oxygen species (ROS) and malondialdehyde (MDA) increased, and the activities of glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and catalase (CAT) were inhibited. In in vitro experiments, microscopic observations showed that the low-Se group had reduced C2C12 cell fusion and a reduced number of differentiated myotubes. In addition, qPCR results showed that differentiation genes (Myog, Myod, Myh2, Myh3, and Myf5) were significantly reduced in the low Se group. Meanwhile, Western blot analysis showed that the levels of differentiation proteins (Myog, Myod, and Myhc) were significantly reduced in the low-Se group. This finding indicates that Se deficiency reduces the expression of skeletal muscle cell differentiation factors. All the above data suggest that Se deficiency can lead to oxidative stress in skeletal muscle, resulting in a reduction in the differentiation capacity of muscle cells.

Keywords:  Myogenic differentiation; Oxidative stress; ROS; Se deficiency; Skeletal muscle

DOI:  https://doi.org/10.1007/s12011-022-03288-2
Sci Rep. 2022 May 19. 12(1): 8388

DVL1 and DVL3 require nuclear localisation to regulate proliferation in human myoblasts.

Johanna Pruller, Nicolas Figeac, Peter S Zammit.

WNT signalling is essential for regulating a diverse range of cellular processes. In skeletal muscle, the WNT pathway plays crucial roles in maintenance of the stem cell pool and myogenic differentiation. Focus is usually directed at examining the function of central components of the WNT pathway, including β-CATENIN and the GSK3β complex and TCF/LEF transcription factors, in tissue homeostasis and cancer. Other core components of the WNT pathway though, are three dishevelled (DVL) proteins: membrane associated proteins that propagate WNT signalling from membrane to nucleus. Here we examined DVL function in human myogenesis and the muscle-related cancer alveolar rhabdomyosarcoma. We demonstrate that DVL1 and DVL3 are necessary for efficient proliferation in human myoblasts and are important for timely myogenic differentiation. DVL1 and DVL3 also contribute to regulation of proliferation in rhabdomyosarcoma. DVL1 or DVL3 must be present in the nucleus to regulate proliferation, but they operate through different protein domains: DVL3 requires the DIX and PDZ domains, while DVL1 does not. Importantly, DVL1 and DVL3 activity is independent of markedly increased translocation of β-CATENIN to the nucleus, normally a hallmark of active canonical WNT signalling.

DOI: https://doi.org/10.1038/s41598-022-10536-x
J Transl Med. 2022 05 14. 20(1): 229

Heat shock protein A4 ablation leads to skeletal muscle myopathy associated with dysregulated autophagy and induced apoptosis.

Manar Elkenani, Amal Z Barakat, Torsten Held, Daniel Marques Rodrigues, Sherok Mobarak, Surabhi Swarnka, Ibrahim M Adham, Belal A Mohamed.

  BACKGROUND: Molecular chaperones assist protein folding, facilitate degradation of misfolded polypeptides, and thereby maintain protein homeostasis. Impaired chaperone activity leads to defective protein quality control that is implicated in multiple skeletal muscle diseases. The heat shock protein A4 (HSPA4) acts as a co-chaperone for HSP70. Previously, we showed that Hspa4 deletion causes impaired protein homeostasis in the heart. However, its functional role in skeletal muscle has not been explored.METHODS: We performed a comparative phenotypic and biochemical analyses of Hspa4 knockout (KO) mice with wild-type (WT) littermates.
RESULTS: HSPA4 is markedly upregulated in regenerating WT muscle in vivo, and in differentiated myoblasts in vitro. Hspa4-KO mice are marked by growth retardation and increased variability in body weight, accompanied by 35% mortality rates during the peri-weaning period. The surviving Hspa4-KO mice experienced progressive skeletal muscle myopathy, characterized by increased number of muscle fibers with centralized nuclei, heterogeneous myofiber size distribution, inflammatory cell infiltrates and upregulation of embryonic and perinatal myosin heavy chain transcripts. Hspa4-KO muscles demonstrated an accumulation of autophagosome-associated proteins including microtubule associated protein1 light chain 3-II (LC3-II) and p62/sequestosome accompanied by increased number of TUNEL-positive nuclei.
CONCLUSIONS: Our findings underscore the indispensable role of HSPA4 in maintenance of muscle integrity through contribution in skeletal muscle autophagy and apoptosis, which might provide a novel therapeutic strategy for skeletal muscle morbidities.

Keywords:  Autophagy; HSPs; Myopathy

DOI:  https://doi.org/10.1186/s12967-022-03418-3
Front Physiol. 2022 ;13 811369

Effect of 32-Weeks High-Intensity Interval Training and Resistance Training on Delaying Sarcopenia: Focus on Endogenous Apoptosis.

Hao Su, Tianhao Wen, Dongsen Liu, Jia Shao, Lei Zhao, Qi Gao.

  Sarcopenia caused by aging is an important factor leading to a decline in the quality of life of older people. Apoptosis in muscle atrophy accelerates the process of muscle loss in older populations. The present study aimed to investigate the effects of 32 weeks of high-intensity interval training (HIIT) and resistance training (RT) on the skeletal muscle-related indices and provide a theoretical basis for regulating the mitochondrial-mediated pathway to delay sarcopenia. We randomly selected 10 from eight-month-old male SD rats (N = 130) as the baseline group; after 1 week of adaptive feeding, the rats were sacrificed. The remaining rats were randomly assigned to one of three groups: control group (C, N = 40, natural aging for 32 weeks), HIIT group (H, N = 40, performed six loops of 3 min at 90% and 3 min at 50% VO2 max speed treadmill running, with 5 min at 70% VO2 max speed at the beginning and the end of the training, 3 times a week for 32 weeks), and resistance group (R, n = 40, 46 min per day, 3 days per week, with a 30% maximum load on a treadmill with a slope of 35°, 15 m/min). The soleus muscles were collected for analysis at baseline and every 8 weeks. Aging resulted in decreased soleus muscle mass and Bcl-2 levels in the mitochondria, while the levels of reactive oxygen species (ROS) and Bax did not change. HIIT reversed the age-associated activation of pro-apoptotic processes, but RT did not. In addition, when rats were aged from 8 to 16 months, the level of Cyt-C did not change, the Caspase-9 levels and Caspase-3 levels decreased gradually in the soleus muscles, the rats of both the HIIT and RT groups had these indices decreased at 32 weeks. The results suggest that the age-associated loss of muscle mass was reversed by training, and the effect of RT was better than that of HIIT. Both the HIIT and RT rats showed a decrease in the apoptosis of skeletal muscle cells after 32 weeks of intervention. HIIT performed better for long-term intervention regarding the pro-apoptotic factors. This study warranted further research to delineate the underlying mechanism of effects of different exercise methods on the changes of aging skeletal muscle at in vivo level.

Keywords:  HIIT; aging; endogenous apoptosis; resistance exercise; sarcopenia

DOI:  https://doi.org/10.3389/fphys.2022.811369
J Cachexia Sarcopenia Muscle. 2022 May 18.

Maternal vitamin D deficiency affects the morphology and function of glycolytic muscle in adult offspring rats.

Natany G Reis, Ana P Assis, Natália Lautherbach, Dawit A Gonçalves, Wilian A Silveira, Henrique J N Morgan, Rafael R Valentim, Lucas F Almeida, Lilian C Heck, Neusa M Zanon, Tatiana E Koike, Audrei R Santos, Elen H Miyabara, Isis C Kettelhut, Luiz C Navegantes.

  BACKGROUND: Fetal stage is a critical developmental window for the skeletal muscle, but little information is available about the impact of maternal vitamin D (Vit. D) deficiency (VDD) on offspring lean mass development in the adult life of male and female animals.METHODS: Female rats (Wistar Hannover) were fed either a control (1000 IU Vit. D3/kg) or a VDD diet (0 IU Vit. D3/kg) for 6 weeks and during gestation and lactation. At weaning, male and female offspring were randomly separated and received a standard diet up to 180 days old.
RESULTS: Vitamin D deficiency induced muscle atrophy in the male (M-VDD) offspring at the end of weaning, an effect that was reverted along the time. Following 180 days, fast-twitch skeletal muscles [extensor digitorum longus (EDL)] from the M-VDD showed a decrease (20%; P < 0.05) in the number of total fibres but an increase in the cross-sectional area of IIB (17%; P < 0.05), IIA (19%; P < 0.05) and IIAX (21%; P < 0.05) fibres. The fibre hypertrophy was associated with the higher protein levels of MyoD (73%; P < 0.05) and myogenin (55% %; P < 0.05) and in the number of satellite cells (128.8 ± 14 vs. 91 ± 7.6 nuclei Pax7 + in the M-CTRL; P < 0.05). M-VDD increased time to fatigue during ex vivo contractions of EDL muscles and showed an increase in the phosphorylation levels of IGF-1/insulin receptor and their downstream targets related to anabolic processes and myogenic activation, including Ser 473 Akt and Ser 21/9 GSK-3β. In such muscles, maternal VDD induced a compensatory increase in the content of calcitriol (two-fold; P < 0.05) and CYP27B1 (58%; P < 0.05), a metabolizing enzyme that converts calcidiol to calcitriol. Interestingly, most morphological and biochemical changes found in EDL were not observed in slow-twitch skeletal muscles (soleus) from the M-VDD group as well as in both EDL and soleus muscles from the female offspring.
CONCLUSIONS: These data show that maternal VDD selectively affects the development of type-II muscle fibres in male offspring rats but not in female offspring rats and suggest that the enhancement of their size and fatigue resistance in fast-twitch skeletal muscle (EDL) is probably due to a compensatory increase in the muscle content of Vit. D in the adult age.

Keywords:  Fetal programming; Hypertrophy; Muscle development; Skeletal muscle; Vitamin D

DOI:  https://doi.org/10.1002/jcsm.12986
Front Physiol. 2022 ;13 880625

Sex-Based Differences in the Myogenic Response and Inflammatory Gene Expression Following Eccentric Contractions in Humans.

Stephen A Fortino, Mai Wageh, Riley Pontello, Chris McGlory, Dinesh Kumbhare, Stuart M Phillips, Gianni Parise.

  After muscle injury, the interaction between muscle satellite cells (SC) and the immune response is instrumental for the repair and regeneration of skeletal muscle tissue. Studies have reported sex-based differences in the skeletal muscle inflammatory and regenerative response following injury. However, many of these studies investigated such differences by manipulating the concentration of estradiol, in rodents and humans, without directly comparing males to females. We sought to explore differences in the myogenic and inflammatory response following unaccustomed eccentric exercise in males and females. We hypothesized that females would have a blunted myogenic and inflammatory response as compared to males. Methods: 26 (13 male, 13 female) healthy young adults (22 ± 0.4 years [mean ± SEM]) performed 300 maximal eccentric contractions (180°/s) of the knee extensors. Muscle biopsies were taken before (pre) and 48 h (post) following eccentric damage. SC content and activation were determined by immunohistochemical and real time-polymerase chain reaction (rt-PCR) analysis. Inflammatory markers were analyzed using rt-PCR. Results: Following eccentric damage, males had a greater expansion of type I-associated SC (p < 0.05), and there was a trend for a greater expansion in total SC (type I + II fibers) (p = 0.06) compared to females. There was a trend for a greater increase in Pax7 and CCL2 gene expression in males compared to females (p = 0.09). Conclusion: We conclude that there are sex-based differences in the myogenic and inflammatory response, where females have a blunted SC and inflammatory response.

Keywords:  inflammation; myogenesis; regeneration; repair; satellite cell

DOI:  https://doi.org/10.3389/fphys.2022.880625
FASEB J. 2022 Jun;36(6): e22347

CTRP11 contributes modestly to systemic metabolism and energy balance.

Dylan C Sarver, Cheng Xu, Dana Carreno, Alexander Arking, Chantelle E Terrillion, Susan Aja, G William Wong.

  C1q/TNF-related proteins (CTRP1-15) constitute a conserved group of secreted proteins of the C1q family with diverse functions. In vitro studies have shown that CTRP11/C1QL4 can inhibit adipogenesis, antagonize myoblast fusion, and promote testosterone synthesis and secretion. Whether CTRP11 is required for these processes in vivo remains unknown. Here, we show that knockout (KO) mice lacking CTRP11 have normal skeletal muscle mass and function, and testosterone level, suggesting that CTRP11 is dispensable for skeletal muscle development and testosterone production. We focused our analysis on whether this nutrient-responsive secreted protein plays a role in controlling sugar and fat metabolism. At baseline when mice are fed a standard chow, CTRP11 deficiency affects metabolic parameters in a sexually dimorphic manner. Only Ctrp11-KO female mice have significantly higher fasting serum ketones and reduced physical activity. In the refeeding phase following food withdrawal, Ctrp11-KO female mice have reduced food intake and increased metabolic rate and energy expenditure, highlighting CTRP11's role in fasting-refeeding response. When challenged with a high-fat diet to induce obesity and metabolic dysfunction, CTRP11 deficiency modestly exacerbates obesity-induced glucose intolerance, with more pronounced effects seen in Ctrp11-KO male mice. Switching to a low-fat diet after obesity induction results in greater fat loss in wild type relative to KO male mice, suggesting impaired response to obesity reversal and reduced metabolic flexibility in the absence of CTRP11. Collectively, our data provide genetic evidence for novel sex-dependent metabolic regulation by CTRP11, but note the overall modest contribution of CTRP11 to systemic energy homeostasis.

Keywords:  C1QL4; C1QTNF11; metabolism; obesity reversal; secreted hormone

DOI:  https://doi.org/10.1096/fj.202200189RR
Biochem Biophys Rep. 2022 Jul;30 101274

Acute bioenergetic insulin sensitivity of skeletal muscle cells: ATP-demand-provoked glycolysis contributes to stimulation of ATP supply.

Rosie A Donnell, Jane E Carré, Charles Affourtit.

  Skeletal muscle takes up glucose in an insulin-sensitive manner and is thus important for the maintenance of blood glucose homeostasis. Insulin resistance during development of type 2 diabetes is associated with decreased ATP synthesis, but the causality of this association is controversial. In this paper, we report real-time oxygen uptake and medium acidification data that we use to quantify acute insulin effects on intracellular ATP supply and ATP demand in rat and human skeletal muscle cells. We demonstrate that insulin increases overall cellular ATP supply by stimulating the rate of glycolytic ATP synthesis. Stimulation is immediate and achieved directly by increased glycolytic capacity, and indirectly by elevated ATP demand from protein synthesis. Raised glycolytic capacity does not result from augmented glucose uptake. Notably, insulin-sensitive glucose uptake is increased synergistically by nitrite. While nitrite has a similar stimulatory effect on glycolytic ATP supply as insulin, it does not amplify insulin stimulation. These data highlight the multifarious nature of acute bioenergetic insulin sensitivity of skeletal muscle cells, and are thus important for the interpretation of changes in energy metabolism that are seen in insulin-resistant muscle.

Keywords:  ATP demand; Cellular energy metabolism; Efficiency of mitochondrial ATP synthesis; Oxidative phosphorylation; Skeletal muscle insulin resistance; Type 2 diabetes

DOI:  https://doi.org/10.1016/j.bbrep.2022.101274
Cell Signal. 2022 May 17. pii: S0898-6568(22)00117-6. [Epub ahead of print] 110355

The Vitamin D/Vitamin D receptor (VDR) axis in muscle atrophy and sarcopenia.

Shelby E Bollen, Joseph J Bass, Satoshi Fujita, Daniel Wilkinson, Martin Hewison, Philip J Atherton.

  Muscle atrophy and sarcopenia (the term given to the age-related decline in muscle mass and function), influence an individuals risk of falls, frailty, functional decline, and, ultimately, impaired quality of life. Vitamin D deficiency (low serum levels of 25-hydroxyvitamin D (25(OH)D3)) has been reported to impair muscle strength and increase risk of sarcopenia. The mechanisms that underpin the link between low 25(OH)D3 and sarcopenia are yet to be fully understood but several lines of evidence have highlighted the importance of both genomic and non-genomic effects of active vitamin D (1,25-dihydroxyvitamin D (1,25(OH)2D3)) and its nuclear vitamin D receptor (VDR), in skeletal muscle functioning. Studies in vitro have demonstrated a key role for the vitamin D/VDR axis in regulating biological processes central to sarcopenic muscle atrophy, such as proteolysis, mitochondrial function, cellular senescence, and adiposity. The aim of this review is to provide a mechanistic overview of the proposed mechanisms for the vitamin D/VDR axis in sarcopenic muscle atrophy.

Keywords:  Ageing; Metabolism; Muscle; Muscle atrophy; Sarcopenia; Vitamin D; Vitamin D receptor

DOI:  https://doi.org/10.1016/j.cellsig.2022.110355
Structure. 2022 May 12. pii: S0969-2126(22)00137-X. [Epub ahead of print]

A drug and ATP binding site in type 1 ryanodine receptor.

Zephan Melville, Haikel Dridi, Qi Yuan, Steven Reiken, Anetta Wronska, Yang Liu, Oliver B Clarke, Andrew R Marks.

  The ryanodine receptor (RyR)/calcium release channel on the sarcoplasmic reticulum (SR) is required for excitation-contraction coupling in skeletal and cardiac muscle. Inherited mutations and stress-induced post-translational modifications result in an SR Ca2+ leak that causes skeletal myopathies, heart failure, and exercise-induced sudden death. A class of therapeutics known as Rycals prevent the RyR-mediated leak, are effective in preventing disease progression and restoring function in animal models, and are in clinical trials for patients with muscle and heart disorders. Using cryogenic-electron microscopy, we present a model of RyR1 with a 2.45-Å resolution before local refinement, revealing a binding site in the RY1&2 domain (3.10 Å local resolution), where the Rycal ARM210 binds cooperatively with ATP and stabilizes the closed state of RyR1.

Keywords:  Rycals; calcium channels; calmodulin; calstabin; cryoEM; electrophysiology; ion channels; muscular dystrophy; ryanodine receptor; sarcoplasmic reticulum

DOI:  https://doi.org/10.1016/j.str.2022.04.010
Front Vet Sci. 2022 ;9 900924

MEF2C Expression Is Regulated by the Post-transcriptional Activation of the METTL3-m6A-YTHDF1 Axis in Myoblast Differentiation.

Xinran Yang, Yue Ning, Sayed Haidar Abbas Raza, Chugang Mei, Linsen Zan.

  N 6-methyladenosine (m6A) plays an essential role in regulating gene expression. However, the effect of m6A on skeletal myoblast differentiation and the underlying mechanisms are still unclear. Here, we ascertained mRNA m6A methylation exhibited declined changes during bovine skeletal myoblast differentiation, and both MEF2C mRNA expression and m6A levels were significantly increased during myoblast differentiation. We found that MEF2C with mutated m6A sites significantly inhibited myoblast differentiation compared with wild-type MEF2C. METTL3 promoted MEF2C protein expression through posttranscriptional modification in an m6A-YTHDF1-dependent manner. Moreover, MEF2C promoted the expression of METTL3 by binding to its promoter. These results revealed that there is a positive feedback loop between these molecules in myoblast differentiation. Our study provided new insights into skeletal muscle differentiation and fusion, which may provide an RNA methylation-based approach for molecular genetics and breeding in livestock as well as for the treatment of muscle-related diseases.

Keywords:  MEF2C; METTL3; N6-methyladenosine; cattle; myoblast differentiation

DOI:  https://doi.org/10.3389/fvets.2022.900924
J Clin Invest. 2022 May 16. pii: e160229. [Epub ahead of print]132(10):

Benefits of aerobic exercise in myotonic dystrophy type 1.

Samuel J Mackenzie, Johanna Hamel, Charles A Thornton.

Myotonic dystrophy type 1 (DM1) is a multisystem trinucleotide repeat expansion disorder characterized by the misregulated alternative splicing of critical mRNAs. Previous work in a transgenic mouse model indicated that aerobic exercise effectively improves splicing regulation and function in skeletal muscle. In this issue of the JCI, Mikhail et al. describe the safety and benefits of applying this approach in individuals affected by DM1. A 12-week aerobic exercise program improved aerobic capacity and mobility, but not by the mechanism observed in transgenic mice. Here, we consider the possible reasons for this disparity and review other salient findings of the study in the context of evolving DM1 research.

DOI: https://doi.org/10.1172/JCI160229
Exp Cell Res. 2022 May 16. pii: S0014-4827(22)00197-5. [Epub ahead of print] 113204

Fluid shear stress-induced mechanotransduction in myoblasts: Does it depend on the glycocalyx?

Mohammad Haroon, Niek G C Bloks, Louise Deldicque, Katrien Koppo, Hadi Seddiqi, Astrid D Bakker, Jenneke Klein-Nulend, Richard T Jaspers.

  Muscle stem cells (MuSCs) are involved in muscle maintenance and regeneration. Mechanically loaded MuSCs within their native niche undergo tensile and shear deformations, but how MuSCs sense mechanical stimuli and translate these into biochemical signals regulating function and fate is still poorly understood. We aimed to investigate whether the glycocalyx is involved in the MuSC mechanoresponse, and whether MuSC morphology affects mechanical loading-induced pressure, shear stress, and fluid velocity distribution. FSS-induced deformation of active proliferating MuSCs (myoblasts) with intact or degraded glycocalyx was assessed by live-cell imaging. Glycocalyx-degradation did not significantly affect nitric oxide production, but reduced FSS-induced myoblast deformation and modulated gene expression. Finite-element analysis revealed that the distribution of FSS-induced pressure, shear stress, and fluid velocity on myoblasts was non-uniform, and the magnitude depended on myoblast morphology and apex-height. In conclusion, our results suggest that the glycocalyx does not play a role in NO production in myoblasts but might impact mechanotransduction and gene expression, which needs further investigation. Future studies will unravel the underlying mechanism by which the glycocalyx affects FSS-induced myoblast deformation, which might be related to increased drag forces. Moreover, MuSCs with varying apex-height experience different levels of FSS-induced pressure, shear stress, and fluid velocity, suggesting differential responsiveness to fluid shear forces.

Keywords:  Cell deformation; Fluid shear stress; Glycocalyx; Mechanotransduction; Myoblast; Nitric oxide

DOI:  https://doi.org/10.1016/j.yexcr.2022.113204
J Clin Invest. 2022 May 16. pii: e159668. [Epub ahead of print]132(10):

Piezo1: opening the way to preventing muscle atrophy.

Ravi Jagasia, Kathryn R Wagner.

The loss of skeletal muscle mass and size, or muscle atrophy, is a common human experience, linked to disability, for which there are no widely accepted pharmacological therapies. Piezo1 is a mechanosensitive cation channel that opens upon alteration of the plasma membrane lipid bilayer, such as through increased membrane tension. In this issue of the JCI, Hirata et al. identified Piezo1 and its downstream effectors, Krüppel-like factor 15 (KLF15) and interleukin-6 (IL-6), as an important signaling pathway in a murine model of disuse atrophy. Through genetic and pharmacological modulation of the pathway, the authors demonstrated that immobilization resulted in downregulation of Piezo1 and basal intracellular calcium concentration ([Ca2+]i), increasing expression of Klf15 and its downstream target Il6 and thereby inducing muscle atrophy. Piezo1 has been considered a therapeutic target for diverse disorders, including atherosclerosis and kidney fibrosis, and with this publication should now also be considered a viable target for disuse atrophy.

DOI: https://doi.org/10.1172/JCI159668
Sci Signal. 2022 May 17. 15(734): eabg4982

The cell polarity protein Vangl2 in the muscle shapes the neuromuscular synapse by binding to and regulating the tyrosine kinase MuSK.

Myriam Boëx, Steve Cottin, Marius Halliez, Stéphanie Bauché, Céline Buon, Nathalie Sans, Mireille Montcouquiol, Jordi Molgó, Muriel Amar, Arnaud Ferry, Mégane Lemaitre, Andrée Rouche, Dominique Langui, Asha Baskaran, Bertrand Fontaine, Julien Messéant, Laure Strochlic.

The development of the neuromuscular junction (NMJ) requires dynamic trans-synaptic coordination orchestrated by secreted factors, including Wnt family morphogens. To investigate how these synaptic cues in NMJ development are transduced, particularly in the regulation of acetylcholine receptor (AChR) accumulation in the postsynaptic membrane, we explored the function of Van Gogh-like protein 2 (Vangl2), a core component of Wnt planar cell polarity signaling. We found that conditional, muscle-specific ablation of Vangl2 in mice reproduced the NMJ differentiation defects seen in mice with global Vangl2 deletion. These alterations persisted into adulthood and led to NMJ disassembly, impaired neurotransmission, and deficits in motor function. Vangl2 and the muscle-specific receptor tyrosine kinase MuSK were functionally associated in Wnt signaling in the muscle. Vangl2 bound to and promoted the signaling activity of MuSK in response to Wnt11. The loss of Vangl2 impaired RhoA activation in cultured mouse myotubes and caused dispersed, rather than clustered, organization of AChRs at the postsynaptic or muscle cell side of NMJs in vivo. Our results identify Vangl2 as a key player of the core complex of molecules shaping neuromuscular synapses and thus shed light on the molecular mechanisms underlying NMJ assembly.

DOI: https://doi.org/10.1126/scisignal.abg4982
JCI Insight. 2022 May 17. pii: e152466. [Epub ahead of print]

A pathogenic mechanism associated with myopathies and structural birth defects involves TPM2 directed myogenesis.

Jennifer McAdow, Shuo Yang, Tiffany Ou, Gary Huang, Matthew B Dobbs, Christina A Gurnett, Michael J Greenberg, Aaron N Johnson.

  Nemaline Myopathy (NM) is the most common congenital myopathy, characterized by extreme weakness of the respiratory, limb, and facial muscles. Pathogenic variants in Tropomyosin 2 (TPM2), which encodes a skeletal muscle specific actin binding protein essential for sarcomere function, cause a spectrum of musculoskeletal disorders that include NM as well as Cap Myopathy, congenital fiber type disproportion, and distal arthrogryposis (DA). The in vivo pathomechanisms underlying TPM2-related disorders are unknown, so we expressed a series of dominant, pathogenic TPM2 variants in Drosophila embryos and found four variants significantly affected muscle development and muscle function. Transient overexpression of the four variants also disrupted the morphogenesis of mouse myotubes in vitro, and negatively affected zebrafish muscle development in vivo. We used transient overexpression assays in zebrafish to characterize two novel TPM2 variants and one recurring variant that we identified in DA patients (V129A, E139K, A155T), and found these variants caused musculoskeletal defects similar to those of known pathogenic variants. The consistency of musculoskeletal phenotypes in our assays correlated with the severity of clinical phenotypes observed in our DA patients, suggesting disrupted myogenesis is a novel pathomechanism of TPM2 disorders, and that our myogenic assays can predict the clinical severity of TPM2 variants.

Keywords:  Molecular genetics; Movement disorders; Muscle; Muscle Biology

DOI:  https://doi.org/10.1172/jci.insight.152466
Cell Metab. 2022 May 12. pii: S1550-4131(22)00182-6. [Epub ahead of print]

Fasting induces a highly resilient deep quiescent state in muscle stem cells via ketone body signaling.

Daniel I Benjamin, Pieter Both, Joel S Benjamin, Christopher W Nutter, Jenna H Tan, Jengmin Kang, Leo A Machado, Julian D D Klein, Antoine de Morree, Soochi Kim, Ling Liu, Hunter Dulay, Ludovica Feraboli, Sharon M Louie, Daniel K Nomura, Thomas A Rando.

  Short-term fasting is beneficial for the regeneration of multiple tissue types. However, the effects of fasting on muscle regeneration are largely unknown. Here, we report that fasting slows muscle repair both immediately after the conclusion of fasting as well as after multiple days of refeeding. We show that ketosis, either endogenously produced during fasting or a ketogenic diet or exogenously administered, promotes a deep quiescent state in muscle stem cells (MuSCs). Although deep quiescent MuSCs are less poised to activate, slowing muscle regeneration, they have markedly improved survival when facing sources of cellular stress. Furthermore, we show that ketone bodies, specifically β-hydroxybutyrate, directly promote MuSC deep quiescence via a nonmetabolic mechanism. We show that β-hydroxybutyrate functions as an HDAC inhibitor within MuSCs, leading to acetylation and activation of an HDAC1 target protein p53. Finally, we demonstrate that p53 activation contributes to the deep quiescence and enhanced resilience observed during fasting.

Keywords:  BHB; HDAC; MuSC; diet; fasting; ketosis; muscle; p53; quiescence; stem cells

DOI:  https://doi.org/10.1016/j.cmet.2022.04.012
Curr Pharm Des. 2022 May 18.

Docosahexaenoic acid ester of phloridzin reduces inflammation and insulin resistance via AMPK.

Jingqing Chen, Zhenlong Wu, Jin Wang, Xuemeng Si, Rui Zhang, Tianqi Sun, Qiaoyan Dong, Wenqing Wu, Yefeng Qiu.

  BACKGROUND: Docosahexaenoic acid-acylated phloridzin (PZ-DHA), a novel polyphenol fatty acid ester derivative, is synthesized through an acylation reaction of phloridzin (PZ) and docosahexaenoic acid (DHA). PZ-DHA is more stable than DHA and exhibits higher cellular uptake and bioavailability than PZ.OBJECTIVE: To investigate the effects of PZ-DHA on insulin resistance in the skeletal muscle and the related mechanisms, we used palmitic acid (PA)-treated C2C12 myotubes as an insulin resistance model.
RESULTS: We found that PZ-DHA increased the activity of AMP-activated protein kinase (AMPK) and improved glucose uptake and mitochondrial function in an AMPK-dependent manner in untreated C2C12 myotubes. PZ-DHA treatment of the myotubes reversed PA-induced insulin resistance; this was indicated by increases in glucose uptake and the expression of membrane glucose transporter 4 (Glut4) and phosphorylated Akt. Moreover, PZ-DHA treatment reversed PA-induced inflammation and oxidative stress. These effects of PZ-DHA were mediated by AMPK. Furthermore, the increase in AMPK activity, improvement in insulin resistance, and decrease in inflammatory and oxidative responses after PZ-DHA treatment diminished upon co-treatment with a liver kinase B1 (LKB1) inhibitor, suggesting that PZ-DHA improved AMPK activity by regulating its upstream kinase, LKB1.
CONCLUSION: The effects of PZ-DHA on insulin resistance in C2C12 myotubes may be mediated by the LKB1-AMPK signaling pathway. Hence, PZ-DHA is a promising therapeutic agent for insulin resistance in type 2 diabetes.

Keywords:  Docosahexaenoic acid; glucose uptake; insulin resistance; myotubes; phloridzin

DOI:  https://doi.org/10.2174/1381612828666220518102440
Physiol Rep. 2022 May;10(10): e15300

Aerobic exercise training-induced follistatin-like 1 secretion in the skeletal muscle is related to arterial stiffness via arterial NO production in obese rats.

Kenichiro Inoue, Shumpei Fujie, Naoki Horii, Henry Yamazaki, Masataka Uchida, Motoyuki Iemitsu.

  Follistatin-like 1 (FSTL1), which is mainly secreted from skeletal muscle and myocardium, upregulates protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS) phosphorylation in vascular endothelial cells. It is unclear whether skeletal muscle- and myocardium-derived FSTL1 secretion induced by aerobic exercise training is involved in the reduction of arterial stiffness via arterial NO production in obese rats. This study aimed to clarify whether aerobic exercise training-induced FSTL1 secretion in myocardium and skeletal muscle is associated with a reduction in arterial stiffness via arterial Akt-eNOS signaling pathway in obese rats. Sixteen Otsuka Long-Evans Tokushima Fatty (OLETF) obese rats were randomly divided into two groups: sedentary control (OLETF-CON) and eight-week aerobic exercise training (treadmill for 60min at 25m/min, 5days/week, OLETF-AT). Eight Long-Evans Tokushima Otsuka (LETO) rats were used as a healthy sedentary control group. In OLETF-CON, serum FSTL1, arterial Akt and eNOS phosphorylation, and arterial nitrite/nitrate (NOx) levels were significantly lower, and carotid-femoral pulse wave velocity (cfPWV) was significantly greater than those in LETO. These parameters were improved in the OLETF-AT compared to the OLETF-CON. In the OLETF-AT, FSTL1 levels in slow-twitch fiber-rich soleus muscle were significantly greater than those in the OLETF-CON, but not in myocardium, fast-twitch fiber-rich tibialis anterior muscle, and adipose tissue. Serum FSTL1 levels were positively correlated with soleus FSTL1, arterial eNOS phosphorylation, and NOx levels and negatively correlated with cfPWV. Thus, aerobic exercise training-induced FSTL1 secretion in slow-twitch fiber-rich muscles may be associated with a reduction in arterial stiffness via arterial NO production in obese rats.

Keywords:  aerobic exercise training; follistatin-like 1; nitric oxide; obesity; skeletal muscle fiber type

DOI:  https://doi.org/10.14814/phy2.15300
Front Physiol. 2022 ;13 837611

What Can We Learn from Single Sarcomere and Myofibril Preparations?

Walter Herzog.

  Sarcomeres are the smallest functional contractile unit of muscle, and myofibrils are striated muscle organelles that are comprised of sarcomeres that are strictly aligned in series. Furthermore, passive forces in sarcomeres and myofibrils are almost exclusively produced by the structural protein titin, and all contractile, regulatory, and structural proteins are in their natural configuration. For these mechanical and structural reasons single sarcomere and myofibril preparations are arguably the most powerful to answer questions on the mechanisms of striated muscle contraction. We developed and optimized single myofibril research over the past 20 years and were the first to mechanically isolate and test single sarcomeres. The results from this research led to the uncovering of the crucial role of titin in muscle contraction, first molecular explanations for the origins of the passive and the residual force enhancement properties of skeletal and cardiac muscles, the discovery of sarcomere length stability on the descending limb of the force-length relationship, and culminating in the formulation of the three-filament theory of muscle contraction that, aside from actin and myosin, proposes a crucial role of titin in active force production. Aside from all the advantages and possibilities that single sarcomere and myofibril preparations offer, there are also disadvantages. These include the fragility of the preparation, the time-consuming training to master these preparations, the limited spatial resolution for length and force measurements, and the unavailability of commercial systems for single sarcomere/myofibril research. Ignoring the mechanics that govern serially linked systems, not considering the spatial resolution and associated accuracies of myofibril systems, and neglecting the fragility of myofibril preparations, has led to erroneous interpretations of results and misleading conclusions. Here, we will attempt to describe the methods and possible applications of single sarcomere/myofibril research and discuss the advantages and disadvantages by focusing on specific applications. It is hoped that this discussion may contribute to identifying the enormous potential of single sarcomere/myofibril research in discovering the secrets of muscle contraction.

Keywords:  actin; mechanics of contraction; myosin; passive force enhancement; residual force enhancement; serial sarcomeres; three-filament theory; titin

DOI:  https://doi.org/10.3389/fphys.2022.837611
J Exp Biol. 2022 May 19. pii: jeb.243730. [Epub ahead of print]

Specialized androgen synthesis in skeletal muscles that actuate elaborate social displays.

Eric R Schuppe, Daniel Tobiansky, Franz Goller, Matthew J Fuxjager.

  Androgens mediate the expression of many reproductive behaviors, including the elaborate displays used to navigate courtship and territorial interactions. In some vertebrates, males can produce androgen-dependent sexual behavior even when levels of testosterone (T) is low in the bloodstream. One idea is that select tissues make their own androgens from scratch to support behavioral performance. We first study this phenomenon in the skeletal muscles that actuate elaborate sociosexual displays in downy woodpeckers and two songbirds. We show that the woodpecker display muscle maintains elevated T when the testes are regressed in the non-breeding season. Both the display muscles of woodpeckers, as well as the display muscles in the avian vocal organ (syrinx or SYR) of songbirds, express all transporters and enzymes necessary to convert cholesterol into bioactive androgens locally. In a final analysis, we broaden our study by looking for these same transporters and enzymes in mammalian muscles that operate at different speeds. Using RNA-seq data, we find that the capacity for de novo synthesis is only present in "superfast" extraocular muscle. Together, our results suggest that skeletal muscle specialized to generate extraordinary twitch-times and/or extremely rapid contractile speeds may depend on androgenic hormones produced locally within the muscle itself. Our study therefore uncovers an important new dimension of androgenic regulation of behavior.

Keywords:  Androgens; Display; Sexual selection; Steroid synthesis

DOI:  https://doi.org/10.1242/jeb.243730
Acta Neuropathol Commun. 2022 May 14. 10(1): 75

Slow motor neurons resist pathological TDP-43 and mediate motor recovery in the rNLS8 model of amyotrophic lateral sclerosis.

Seong Kwon Hur, Mandana Hunter, Myrna A Dominique, Madona Farag, Dejania Cotton-Samuel, Tahiyana Khan, John Q Trojanowski, Krista J Spiller, Virginia M-Y Lee.

  In the intermediate stages of amyotrophic lateral sclerosis (ALS), surviving motor neurons (MNs) that show intrinsic resistance to TDP-43 proteinopathy can partially compensate for the loss of their more disease-susceptible counterparts. Elucidating the mechanisms of this compensation may reveal approaches for attenuating motor impairment in ALS patients. In the rNLS8 mouse model of ALS-like pathology driven by doxycycline-regulated neuronal expression of human TDP-43 lacking a nuclear localization signal (hTDP-43ΔNLS), slow MNs are more resistant to disease than fast-fatigable (FF) MNs and can mediate recovery following transgene suppression. In the present study, we used a viral tracing strategy to show that these disease-resistant slow MNs sprout to reinnervate motor endplates of adjacent muscle fibers vacated by degenerated FF MNs. Moreover, we found that neuromuscular junctions within fast-twitch skeletal muscle (tibialis anterior, TA) reinnervated by SK3-positive slow MNs acquire resistance to axonal dieback when challenged with a second course of hTDP-43ΔNLS pathology. The selective resistance of reinnervated neuromuscular junctions was specifically induced by the unique pattern of reinnervation following TDP-43-induced neurodegeneration, as recovery from unilateral sciatic nerve crush did not produce motor units resistant to subsequent hTDP-43ΔNLS. Using cross-reinnervation and self-reinnervation surgery in which motor axons are disconnected from their target muscle and reconnected to a new muscle, we show that FF MNs remain hTDP-43ΔNLS-susceptible and slow MNs remain resistant, regardless of which muscle fibers they control. Collectively, these findings demonstrate that MN identity dictates the susceptibility of neuromuscular junctions to TDP-43 pathology and slow MNs can drive recovery of motor systems due to their remarkable resilience to TDP-43-driven degeneration. This study highlights a potential pathway for regaining motor function with ALS pathology in the advent of therapies that halt the underlying neurodegenerative process.

Keywords:  Amyotrophic lateral sclerosis; Cross-reinnervation surgery; Neurodegeneration; Neuropathology; TDP-43; rNLS8

DOI:  https://doi.org/10.1186/s40478-022-01373-0
J Physiol. 2022 May 16.

Rho-kinase inhibition improves haemodynamic responses and circulating ATP during hypoxia and moderate intensity handgrip exercise in healthy older adults.

Matthew L Racine, Janée D Terwoord, Nathaniel B Ketelhut, Nate P Bachman, Jennifer C Richards, Gary J Luckasen, Frank A Dinenno.

  KEY POINTS: Skeletal muscle blood flow responses to hypoxia and exercise are impaired with age. Blunted increases in circulating ATP, a vasodilator, in older adults may contribute to age-related impairments in haemodynamics. Red blood cells (RBCs) are a primary source of circulating ATP, and treating isolated RBCs with a Rho-kinase inhibitor improves age-related impairments in deoxygenation-induced RBC ATP release. In this study, treating healthy older adults systemically with the Rho-kinase inhibitor fasudil improved blood flow and circulating ATP responses during hypoxia and moderate intensity handgrip exercise compared to young adults, and also tended to improve isolated RBC ATP release. Improved blood flow regulation with fasudil was also associated with increased skeletal muscle oxygen delivery during hypoxia and exercise in older adults. This is the first study to demonstrate that Rho-kinase inhibition can significantly improve age-related impairments in haemodynamic and circulating ATP responses to physiological stimuli, which may have therapeutic implications.ABSTRACT: Skeletal muscle haemodynamics and circulating adenosine triphosphate (ATP) responses during hypoxia and exercise are blunted in older (OA) vs young (YA) adults, which may be associated with impaired red blood cell (RBC) ATP release. Rho-kinase inhibition improves deoxygenation-induced ATP release from OA isolated RBCs. We tested the hypothesis that Rho-kinase inhibition (via fasudil) in vivo would improve local haemodynamic and ATP responses during hypoxia and exercise in OA. Healthy YA (25±3yr; n = 12) and OA (65±5yr; n = 13) participated in a randomized, double-blind, placebo-controlled, crossover study on 2 days (≥5d between visits). A forearm deep venous catheter was used to administer saline/fasudil and sample venous plasma ATP ([ATP]V ). Forearm vascular conductance (FVC) and [ATP]V were measured at rest, during isocapnic hypoxia (80% SpO2 ), and during graded rhythmic handgrip exercise that was similar between groups (5, 15, and 25% maximum voluntary contraction (MVC)). Isolated RBC ATP release was measured during normoxia/hypoxia. With saline, ΔFVC was lower (P<0.05) in OA vs YA during hypoxia (∼60%) and during 15 and 25% MVC (∼25-30%), and these impairments were abolished with fasudil. Similarly, [ATP]V and ATP effluent responses from normoxia to hypoxia and rest to 25% MVC were lower in OA vs YA and improved with fasudil (P<0.05). Isolated RBC ATP release during hypoxia was impaired in OA vs YA (∼75%; P<0.05), which tended to improve with fasudil in OA (P = 0.082). These data suggest Rho-kinase inhibition improves haemodynamic responses to hypoxia and moderate intensity exercise in OA, which may be due in part to improved circulating ATP. Abstract figure legend This article is protected by copyright. All rights reserved.

Keywords:  ATP; aging; blood flow; exercise; fasudil; hypoxia

DOI:  https://doi.org/10.1113/JP282730
Mol Ther Methods Clin Dev. 2022 Jun 09. 25 333-343

Combined overexpression of ATXN1L and mutant ATXN1 knockdown by AAV rescue motor phenotypes and gene signatures in SCA1 mice.

Ellie M Carrell, Megan S Keiser, Ashley B Robbins, Beverly L Davidson.

  Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disease caused by a (CAG) repeat expansion in the coding sequence of ATXN1. The primary mechanism of disease in SCA1 is toxic gain of function by polyglutamine-expanded mutant ATXN1 and is compounded by partial loss of wild-type function. Addressing both disease mechanisms, we have shown that virally expressed RNA interference targeting ATXN1 can both prevent and reverse disease phenotypes in SCA1 mice, and that overexpression of the ATXN1 homolog, ataxin 1-like (ATXN1L), improves disease readouts when delivered pre-symptomatically. Here, we combined these therapeutic approaches into two, dual component recombinant adeno-associated virus (rAAV) vectors and tested their ability to reverse disease in symptomatic SCA1 mice using behavior, pathological, and next-generation sequencing assays. Mice treated with vectors expressing human ATXN1L (hATXN1L) alone showed motor improvements and changes in gene expression that reflected increases in pro-development pathways. When hATN1L was combined with miS1, a previously validated microRNA targeting h ATXN1, there was added normalization of disease allele-induced changes in gene expression along with motor improvements. Our data show the additive nature of this two-component approach for a more effective SCA1 therapy.

Keywords:  AAV; RNAi; ataxin-1; ataxin-1-like; cerebellum; miRNA; neurodegeneration; spinocerebellar ataxia type 1

DOI:  https://doi.org/10.1016/j.omtm.2022.04.004
Physiol Rep. 2022 May;10(10): e15285

Heterozygous SOD2 deletion selectively impairs SERCA function in the soleus of female mice.

Jessica L Braun, Holt N Messner, Riley E G Cleverdon, Ryan W Baranowski, Sophie I Hamstra, Mia S Geromella, Jeffrey A Stuart, Val A Fajardo.

The sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA) restores intracellular Ca2+ ([Ca2+ ]i ) to resting levels after muscle contraction, ultimately eliciting relaxation. SERCA pumps are highly susceptible to tyrosine (T)-nitration, impairing their ability to take up Ca2+ resulting in reduced muscle function and increased [Ca2+ ]i and cellular damage. The mitochondrial antioxidant enzyme, superoxide dismutase 2 (SOD2), converts superoxide radicals into less reactive H2 O2 . Heterozygous deletion of SOD2 (Sod2+/- ) in mice increases mitochondrial oxidative stress; however, the consequences of reduced SOD2 expression in skeletal and cardiac muscle, specifically the effect on SERCA pumps, has yet to be investigated. We obtained soleus, extensor digitorum longus (EDL), and left ventricle (LV) muscles from 6 to 7 month-old wild-type (WT) and Sod2+/- female C57BL/6J mice. Ca2+ -dependent SERCA activity assays were performed to assess SERCA function. Western blotting was conducted to examine the protein content of SERCA, phospholamban, and sarcolipin; and immunoprecipitation experiments were done to assess SERCA2a- and SERCA1a-specific T-nitration. Heterozygous SOD2 deletion did not alter SERCA1a or SERCA2a expression in the soleus or LV but reduced SERCA2a in the EDL compared with WT, though this was not statistically significant. Soleus muscles from Sod2+/- mice showed a significant reduction in SERCA's apparent affinity for Ca2+ when compared to WT, corresponding with significantly elevated SERCA2a T-nitration in the soleus. No effect was seen in the EDL or the LV. This is the first study to investigate the effects of SOD2 deficiency on muscle SERCA function and shows that it selectively impairs SERCA function in the soleus.

DOI: https://doi.org/10.14814/phy2.15285
Cell Rep Med. 2022 May 17. pii: S2666-3791(22)00158-6. [Epub ahead of print]3(5): 100633

Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults.

Anurag Singh, Davide D'Amico, Pénélope A Andreux, Andréane M Fouassier, William Blanco-Bose, Mal Evans, Patrick Aebischer, Johan Auwerx, Chris Rinsch.

  Targeting mitophagy to activate the recycling of faulty mitochondria during aging is a strategy to mitigate muscle decline. We present results from a randomized, placebo-controlled trial in middle-aged adults where we administer a postbiotic compound Urolithin A (Mitopure), a known mitophagy activator, at two doses for 4 months (NCT03464500). The data show significant improvements in muscle strength (∼12%) with intake of Urolithin A. We observe clinically meaningful improvements with Urolithin A on aerobic endurance (peak oxygen oxygen consumption [VO2]) and physical performance (6 min walk test) but do not notice a significant improvement on peak power output (primary endpoint). Levels of plasma acylcarnitines and C-reactive proteins are significantly lower with Urolithin A, indicating higher mitochondrial efficiency and reduced inflammation. We also examine expression of proteins linked to mitophagy and mitochondrial metabolism in skeletal muscle and find a significant increase with Urolithin A administration. This study highlights the benefit of Urolithin A to improve muscle performance.

Keywords:  Mitopure; Urolithin A; aging; clinical trial; exercise performance; mitochondria; mitophagy; muscle strength

DOI:  https://doi.org/10.1016/j.xcrm.2022.100633
Elife. 2022 May 17. pii: e77553. [Epub ahead of print]11

Residual force enhancement is affected more by quadriceps muscle length than stretch amplitude.

Patrick Bakenecker, Tobias Weingarten, Daniel Hahn, Brent James Raiteri.

  Little is known about how muscle length affects residual force enhancement (rFE) in humans. We therefore investigated rFE at short, long, and very long muscle lengths within the human quadriceps and patellar tendon (PT) using conventional dynamometry with motion capture (rFETQ) and a new, non-invasive shear-wave tensiometry technique (rFEWS). Eleven healthy male participants performed submaximal (50% max.) EMG-matched fixed-end reference and stretch-hold contractions across these muscle lengths while muscle fascicle length changes of the vastus lateralis (VL) were captured using B-mode ultrasound. We found significant rFETQ at long (7±5%) and very long (12±8%), but not short (2±5%) muscle lengths, whereas rFEWS was only significant at the very long (38±27%), but not short (8±12%) or long (6±10%) muscle lengths. We also found significant relationships between VL fascicle length and rFETQ (r=0.63, p=.001) and rFEWS (r=0.52, p=.017), but relationships were not significant between VL fascicle stretch amplitude and rFETQ (r=0.33, p=.126) or rFEWS (r=0.29, p=.201). PT shear-wave speed-angle relationships did not agree with estimated PT force-angle relationships, which indicates that estimating PT loads from shear-wave tensiometry might be inaccurate. We conclude that increasing muscle length rather than stretch amplitude contributes more to rFE during submaximal voluntary contractions of the human quadriceps.

Keywords:  human; physics of living systems

DOI:  https://doi.org/10.7554/eLife.77553