bims-muscge 2023-11-05 papers

bims-muscge

Biomed News

on Muscle stem cells and gene therapy

Issue of 2023–11–05
fiveteen papers selected by
Chance Bowman, Dartmouth College

Loss of adenosine A3 receptors accelerates skeletal muscle regeneration in mice following cardiotoxin-induced injury.
ALDH1A1 and ALDH1A3 paralogues of aldehyde dehydrogenase 1 control myogenic differentiation of skeletal muscle satellite cells by retinoic acid-dependent and -independent mechanisms.
Regenerating human skeletal muscle forms an emerging niche in vivo to support PAX7 cells.
Gli1 marks a sentinel muscle stem cell population for muscle regeneration.
Efficient exon skipping by base-editor-mediated abrogation of exonic splicing enhancers.
Dystrophin- and Utrophin-Based Therapeutic Approaches for Treatment of Duchenne Muscular Dystrophy: A Comparative Review.
Efficient Myogenic Activities Achieved through Blade-Casting-Assisted Bioprinting of Aligned Myoblasts Laden in Collagen Bioink.
T cell biology in neuromuscular disorders: a focus on Duchenne Muscular Dystrophy and Amyotrophic Lateral Sclerosis.
Metabolomic and Lipidomic Signature of Skeletal Muscle with Constitutively Active mTORC1.
Muscle Bmal1 is necessary for normal transcriptomic and metabolomic adaptation to endurance exercise training.
Receptor interacting protein kinase-3 mediates both myopathy and cardiomyopathy in preclinical animal models of Duchenne muscular dystrophy.
Lactate promotes myogenesis via activating H3K9 lactylation-dependent up-regulation of Neu2 expression.
Muscle-targeted nanoparticles strengthen the effects of small-molecule inhibitors in ameliorating sarcopenia.
Protein-coding potential of non-canonical open reading frames in human transcriptome.
Ivabradine ameliorates cardiomyopathy progression in a Duchenne muscular dystrophy model rat.

Cell Death Dis. 2023 Oct 28. 14(10): 706

Loss of adenosine A3 receptors accelerates skeletal muscle regeneration in mice following cardiotoxin-induced injury.

Nastaran Tarban, Albert Bálint Papp, Dávid Deák, Péter Szentesi, Hajnalka Halász, Andreas Patsalos, László Csernoch, Zsolt Sarang, Zsuzsa Szondy.

Skeletal muscle regeneration is a complex process orchestrated by multiple interacting steps. An increasing number of reports indicate that inflammatory responses play a central role in linking initial muscle injury responses to timely muscle regeneration following injury. The nucleoside adenosine has been known for a long time as an endogenously produced anti-inflammatory molecule that is generated in high amounts during tissue injury. It mediates its physiological effects via four types of adenosine receptors. From these, adenosine A3 receptors (A3Rs) are not expressed by the skeletal muscle but are present on the surface of various inflammatory cells. In the present paper, the effect of the loss of A3Rs was investigated on the regeneration of the tibialis anterior (TA) muscle in mice following cardiotoxin-induced injury. Here we report that regeneration of the skeletal muscle from A3R-/- mice is characterized by a stronger initial inflammatory response resulting in a larger number of transmigrating inflammatory cells to the injury site, faster clearance of cell debris, enhanced proliferation and faster differentiation of the satellite cells (the muscle stem cells), and increased fusion of the generated myoblasts. This leads to accelerated skeletal muscle tissue repair and the formation of larger myofibers. Though the infiltrating immune cells expressed A3Rs and showed an increased inflammatory profile in the injured A3R-/- muscles, bone marrow transplantation experiments revealed that the increased response of the tissue-resident cells to tissue injury is responsible for the observed phenomenon. Altogether our data indicate that A3Rs are negative regulators of injury-related regenerative inflammation and consequently also that of the muscle fiber growth in the TA muscle. Thus, inhibiting A3Rs might have a therapeutic value during skeletal muscle regeneration following injury.

DOI: https://doi.org/10.1038/s41419-023-06228-7
Cell Tissue Res. 2023 Oct 31.

ALDH1A1 and ALDH1A3 paralogues of aldehyde dehydrogenase 1 control myogenic differentiation of skeletal muscle satellite cells by retinoic acid-dependent and -independent mechanisms.

Laura Steingruber, Florian Krabichler, Sophie Franzmeier, Wei Wu, Jürgen Schlegel, Marco Koch.

  ALDH1A1 and ALDH1A3 paralogues of aldehyde dehydrogenase 1 (ALDH1) control myogenic differentiation of skeletal muscle satellite cells (SC) by formation of retinoic acid (RA) and subsequent cell cycle adjustments. The respective relevance of each paralogue for myogenic differentiation and the mechanistic interaction of each paralogue within RA-dependent and RA-independent pathways remain elusive.We analysed the impact of ALDH1A1 and ALDH1A3 activity on myogenesis of murine C2C12 myoblasts. Both paralogues are pivotal factors in myogenic differentiation, since CRISPR/Cas9-edited single paralogue knock-out impaired serum withdrawal-induced myogenic differentiation, while successive recombinant re-expression of ALDH1A1 or ALDH1A3, respectively, in the corresponding ALDH1 paralogue single knock-out cell lines, recovered the differentiation potential. Loss of differentiation in single knock-out cell lines was restored by treatment with RA-analogue TTNPB, while RA-receptor antagonization by AGN 193109 inhibited differentiation of wildtype cell lines, supporting the idea that RA-dependent pathway is pivotal for myogenic differentiation which is accomplished by both paralogues.However, overexpression of ALDH1-paralogues or disulfiram-mediated inhibition of ALDH1 enzymatic activity not only increased ALDH1A1 and ALDH1A3 protein levels but also induced subsequent differentiation of C2C12 myoblasts independently from serum withdrawal, indicating that ALDH1-dependent myogenic differentiation relies on different cellular conditions. Remarkably, ALDH1-paralogue knock-out impaired the autophagic flux, namely autophagosome cargo protein p62 formation and LC3B-I to LC3B-II conversion, demonstrating that ALDH1-paralogues interact with autophagy in myogenesis. Together, ALDH1 paralogues play a crucial role in myogenesis by orchestration of complex RA-dependent and RA-independent pathways.

Keywords:  ALDH1A1/ALDH1A3 paralogue; Autophagy; CRISPR/Cas9 knock-out; Myogenic differentiation; Retinoic acid

DOI:  https://doi.org/10.1007/s00441-023-03838-7
Nat Cell Biol. 2023 Nov 02.

Regenerating human skeletal muscle forms an emerging niche in vivo to support PAX7 cells.

Michael R Hicks, Kholoud K Saleh, Ben Clock, Devin E Gibbs, Mandee Yang, Shahab Younesi, Lily Gane, Victor Gutierrez-Garcia, Haibin Xi, April D Pyle.

Skeletal muscle stem and progenitor cells including those derived from human pluripotent stem cells (hPSCs) offer an avenue towards personalized therapies and readily fuse to form human-mouse myofibres in vivo. However, skeletal muscle progenitor cells (SMPCs) inefficiently colonize chimeric stem cell niches and instead associate with human myofibres resembling foetal niches. We hypothesized competition with mouse satellite cells (SCs) prevented SMPC engraftment into the SC niche and thus generated an SC ablation mouse compatible with human engraftment. Single-nucleus RNA sequencing of SC-ablated mice identified the absence of a transient myofibre subtype during regeneration expressing Actc1. Similarly, ACTC1+ human myofibres supporting PAX7+ SMPCs increased in SC-ablated mice, and after re-injury we found SMPCs could now repopulate into chimeric niches. To demonstrate ACTC1+ myofibres are essential to supporting PAX7 SMPCs, we generated caspase-inducible ACTC1 depletion human pluripotent stem cells, and upon SMPC engraftment we found a 90% reduction in ACTC1+ myofibres and a 100-fold decrease in PAX7 cell numbers compared with non-induced controls. We used spatial RNA sequencing to identify key factors driving emerging human niche formation between ACTC1+ myofibres and PAX7+ SMPCs in vivo. This revealed that transient regenerating human myofibres are essential for emerging niche formation in vivo to support PAX7 SMPCs.

DOI: https://doi.org/10.1038/s41556-023-01271-0
Nat Commun. 2023 Nov 01. 14(1): 6993

Gli1 marks a sentinel muscle stem cell population for muscle regeneration.

Jiayin Peng, Lili Han, Biao Liu, Jiawen Song, Yuang Wang, Kunpeng Wang, Qian Guo, XinYan Liu, Yu Li, Jujin Zhang, Wenqing Wu, Sheng Li, Xin Fu, Cheng-le Zhuang, Weikang Zhang, Shengbao Suo, Ping Hu, Yun Zhao.

Adult skeletal muscle regeneration is mainly driven by muscle stem cells (MuSCs), which are highly heterogeneous. Although recent studies have started to characterize the heterogeneity of MuSCs, whether a subset of cells with distinct exists within MuSCs remains unanswered. Here, we find that a population of MuSCs, marked by Gli1 expression, is required for muscle regeneration. The Gli1+ MuSC population displays advantages in proliferation and differentiation both in vitro and in vivo. Depletion of this population leads to delayed muscle regeneration, while transplanted Gli1+ MuSCs support muscle regeneration more effectively than Gli1- MuSCs. Further analysis reveals that even in the uninjured muscle, Gli1+ MuSCs have elevated mTOR signaling activity, increased cell size and mitochondrial numbers compared to Gli1- MuSCs, indicating Gli1+ MuSCs are displaying the features of primed MuSCs. Moreover, Gli1+ MuSCs greatly contribute to the formation of GAlert cells after muscle injury. Collectively, our findings demonstrate that Gli1+ MuSCs represents a distinct MuSC population which is more active in the homeostatic muscle and enters the cell cycle shortly after injury. This population functions as the tissue-resident sentinel that rapidly responds to injury and initiates muscle regeneration.

DOI: https://doi.org/10.1038/s41467-023-42837-8
Cell Rep. 2023 Oct 30. pii: S2211-1247(23)01352-9. [Epub ahead of print]42(11): 113340

Efficient exon skipping by base-editor-mediated abrogation of exonic splicing enhancers.

Han Qiu, Geng Li, Juanjuan Yuan, Dian Yang, Yunqing Ma, Feng Wang, Yi Dai, Xing Chang.

  Duchenne muscular dystrophy (DMD) is a severe genetic disease caused by the loss of the dystrophin protein. Exon skipping is a promising strategy to treat DMD by restoring truncated dystrophin. Here, we demonstrate that base editors (e.g., targeted AID-mediated mutagenesis [TAM]) are able to efficiently induce exon skipping by disrupting functional redundant exonic splicing enhancers (ESEs). By developing an unbiased and high-throughput screening to interrogate exonic sequences, we successfully identify novel ESEs in DMD exons 51 and 53. TAM-CBE (cytidine base editor) induces near-complete skipping of the respective exons by targeting these ESEs in patients' induced pluripotent stem cell (iPSC)-derived cardiomyocytes. Combined with strategies to disrupt splice sites, we identify suitable single guide RNAs (sgRNAs) with TAM-CBE to efficiently skip most DMD hotspot exons without substantial double-stranded breaks. Our study thus expands the repertoire of potential targets for CBE-mediated exon skipping in treating DMD and other RNA mis-splicing diseases.

Keywords:  CP: Genomics; CRISPR; DMD; base editor; exon skipping; exonic splicing enhancer; gene therapy; genome editing; muscular dystrophy

DOI:  https://doi.org/10.1016/j.celrep.2023.113340
BioDrugs. 2023 Nov 02.

Dystrophin- and Utrophin-Based Therapeutic Approaches for Treatment of Duchenne Muscular Dystrophy: A Comparative Review.

Sylwia Szwec, Zuzanna Kapłucha, Jeffrey S Chamberlain, Patryk Konieczny.

Duchenne muscular dystrophy is a devastating disease that leads to progressive muscle loss and premature death. While medical management focuses mostly on symptomatic treatment, decades of research have resulted in first therapeutics able to restore the affected reading frame of dystrophin transcripts or induce synthesis of a truncated dystrophin protein from a vector, with other strategies based on gene therapy and cell signaling in preclinical or clinical development. Nevertheless, recent reports show that potentially therapeutic dystrophins can be immunogenic in patients. This raises the question of whether a dystrophin paralog, utrophin, could be a more suitable therapeutic protein. Here, we compare dystrophin and utrophin amino acid sequences and structures, combining published data with our extended in silico analyses. We then discuss these results in the context of therapeutic approaches for Duchenne muscular dystrophy. Specifically, we focus on strategies based on delivery of micro-dystrophin and micro-utrophin genes with recombinant adeno-associated viral vectors, exon skipping of the mutated dystrophin pre-mRNAs, reading through termination codons with small molecules that mask premature stop codons, dystrophin gene repair by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9)-mediated genetic engineering, and increasing utrophin levels. Our analyses highlight the importance of various dystrophin and utrophin domains in Duchenne muscular dystrophy treatment, providing insights into designing novel therapeutic compounds with improved efficacy and decreased immunoreactivity. While the necessary actin and β-dystroglycan binding sites are present in both proteins, important functional distinctions can be identified in these domains and some other parts of truncated dystrophins might need redesigning due to their potentially immunogenic qualities. Alternatively, therapies based on utrophins might provide a safer and more effective approach.

DOI: https://doi.org/10.1007/s40259-023-00632-3
Biomacromolecules. 2023 Nov 02.

Efficient Myogenic Activities Achieved through Blade-Casting-Assisted Bioprinting of Aligned Myoblasts Laden in Collagen Bioink.

SuHyeok Lee, WonJin Kim, GeunHyung Kim.

This study investigated mechanical stimulation combined with three-dimensional (3D) bioprinting as a new approach for introducing biophysical and biological cues for tissue regeneration. A blade-casting method in conjunction with bioprinting was employed to fabricate bioengineered skeletal muscle constructs using a bioink composed of C2C12 myoblasts and collagen type-I. Various printing process parameters were selected and optimized to achieve a highly organized cell alignment within the constructs. The resulting cell-aligned constructs demonstrated remarkable improvement in actin filament alignment and cell proliferation compared with conventionally printed cell-laden constructs. This improvement can be attributed to the synergistic effects of mechanotransduction, facilitating the cellular response to mechanical cues and the alignment of fibrillated collagen, which plays a significant role in modulating cellular functions and promoting muscle tissue regeneration. Furthermore, we assessed the impact of blade casting combined with 3D bioprinting on gene expression. The expression levels of myogenesis-related genes were substantially upregulated, with an approximately 1.6-fold increase compared to the constructs fabricated without the blade-casting technique. The results demonstrated the effectiveness of combining mechanical stimulation through blade casting with 3D bioprinting in promoting aligned cell structures, enhancing cellular functions, and driving muscle tissue regeneration.

DOI: https://doi.org/10.1021/acs.biomac.3c00749
Front Immunol. 2023 ;14 1202834

T cell biology in neuromuscular disorders: a focus on Duchenne Muscular Dystrophy and Amyotrophic Lateral Sclerosis.

Julia Pereira Lemos, Liliane Patrícia Gonçalves Tenório, Vincent Mouly, Gillian Butler-Browne, Daniella Arêas Mendes-da-Cruz, Wilson Savino, Piera Smeriglio.

  Growing evidence demonstrates a continuous interaction between the immune system, the nerve and the muscle in neuromuscular disorders of different pathogenetic origins, such as Duchenne Muscular Dystrophy (DMD) and Amyotrophic Lateral Sclerosis (ALS), the focus of this review. Herein we highlight the complexity of the cellular and molecular interactions involving the immune system in neuromuscular disorders, as exemplified by DMD and ALS. We describe the distinct types of cell-mediated interactions, such as cytokine/chemokine production as well as cell-matrix and cell-cell interactions between T lymphocytes and other immune cells, which target cells of the muscular or nervous tissues. Most of these interactions occur independently of exogenous pathogens, through ligand-receptor binding and subsequent signal transduction cascades, at distinct levels of specificity. Although this issue reveals the complexity of the system, it can also be envisioned as a window of opportunity to design therapeutic strategies (including synthetic moieties, cell and gene therapy, as well as immunotherapy) by acting upon one or more targets. In this respect, we discuss ongoing clinical trials using VLA-4 inhibition in DMD, and in ALS, with a focus on regulatory T cells, both revealing promising results.

Keywords:  Amyotrophic Lateral Sclerosis; Duchenne Muscular Dystrophy; effector T cells; immunotherapy; regulatory T cells

DOI:  https://doi.org/10.3389/fimmu.2023.1202834
J Nutr. 2023 Oct 26. pii: S0022-3166(23)72670-9. [Epub ahead of print]

Metabolomic and Lipidomic Signature of Skeletal Muscle with Constitutively Active mTORC1.

Rosario Maroto, Ted G Graber, Trevor B Romsdahl, Andrzej Kudlicki, William K Russell, Blake B Rasmussen.

   BACKGROUND: Regulation of mTORC1 plays an important role in aging and nutrition. For example, caloric restriction reduces mTORC1 signaling and extends lifespan, whereas nutrient abundance and obesity increase mTORC1 signaling and reduce lifespan. Skeletal muscle specific knockout of DEP domain-containing 5 protein (DEPDC5) results in constitutively active mTORC1 signaling, muscle hypertrophy and an increase in mitochondrial respiratory capacity. The metabolic profile of skeletal muscle, in the setting of hyperactive mTORC1 signaling, is not well known.
OBJECTIVES: To determine the metabolomic and lipidomic signature in skeletal muscle from female and male wildtype and DEPDC5 knockout mice.
METHODS: Tibialis anterior (TA) muscles from wildtype (WT) and transgenic (conditional skeletal muscle specific DEPDC5 knockout, KO) were obtained from female (F) and male (M) adult mice. Polar metabolites and lipids were extracted using a Bligh-Dyer extraction from 5 samples per group and identified and quantified by LC-MS/MS. Resulting analyte peak areas were analyzed with t-test, ANOVA, and volcano plots for group comparisons (e.g., WT vs KO) and multivariate statistical analysis for genotype and sex comparisons.
RESULTS: A total of 162 polar metabolites (organic acids, amino acids and amines and acyl carnitines) and 1141 lipid metabolites were detected in TA samples by LC-MS/MS. Few polar metabolites showed significant differences in KO muscles compared to WT within the same sex group. P-aminobenzoic acid, β-alanine and dopamine were significantly higher in KO male muscle whereas erythrose- 4-phosphate and oxoglutaric acid were significantly reduced in KO females. The lipidomic profile of the KO groups revealed an increase of muscle phospholipids and reduced triacylglycerol and diacylglycerol compared to the WT groups.
CONCLUSIONS: Sex differences were detected in polar metabolome and lipids were dependent on genotype. Metabolomic profile of mice with hyperactive skeletal muscle mTORC1 is consistent with an upregulation of mitochondrial function and amino acid utilization for protein synthesis.

Keywords:  DEPDC5; metabolites; sexual dimorphism; skeletal muscle

DOI:  https://doi.org/10.1016/j.tjnut.2023.10.016
bioRxiv. 2023 Oct 17. pii: 2023.10.13.562100. [Epub ahead of print]

Muscle Bmal1 is necessary for normal transcriptomic and metabolomic adaptation to endurance exercise training.

Mark R Viggars, Hannah E Berko, Miguel A Gutierrez-Monreal, Ryan A Martin, Karyn A Esser.

Objective: The skeletal muscle circadian clock plays a pivotal role in muscle homeostasis and metabolic flexibility. Recently, this clock mechanism has been linked to both transcriptional and metabolic responses to acute exercise. However, the contribution of the circadian clock mechanism to the molecular and phenotypic adaptations to exercise training have not been defined.
Methods: Inducible skeletal muscle-specific Bmal1-floxed mice were treated with tamoxifen to induce skeletal muscle specific deletion of Bmal1 (iMSBmal1KO) or given a vehicle. Mice were assigned to normal cage conditions, or 6-weeks of progressive treadmill training. Exercise performance, body composition, and tissue/serum indices of metabolic health were assessed over the timecourse of training. Gastrocnemius muscles were collected 48-hours after their last exercise bout for histological, biochemical, and molecular analyses including RNA-sequencing and untargeted metabolomics.
Results: Improvements in exercise workload and maximal performance were comparable between iMSBmal1KO mice and vehicle treated controls after 6-weeks of exercise training. However, exercise training in the absence of Bmal1 was not able to rescue the metabolic phenotype and hyperinsulinemia of the iMSBmal1KO mice, attributed to the continued dysregulation of core clock components and gene expression relating to glucose metabolism. Importantly, a much larger and divergent transcriptional reprogramming occurred in the muscle of iMSBmal1KO mice in comparison to their vehicle treated counterparts. This response included a large compensatory upregulation of genes associated with fatty acid β-oxidation, pyruvate metabolism, citric acid cycle components and oxidative phosphorylation components, including mitochondrial subunits and mitoribosome units.
Conclusions: Collectively, we propose that endurance training requires muscle Bmal1 , and the core clock network, to elicit well recognized molecular adaptations. In the absence of Bmal1 , exercise training results in a much larger and divergent re-networking of the basal skeletal muscle transcriptome and metabolome. We also demonstrate that skeletal muscle Bmal1 is indispensable for the transcriptional regulation of glucose homeostasis, even after a 6-weeks exercise training programme.

DOI: https://doi.org/10.1101/2023.10.13.562100
J Cachexia Sarcopenia Muscle. 2023 Nov 01.

Receptor interacting protein kinase-3 mediates both myopathy and cardiomyopathy in preclinical animal models of Duchenne muscular dystrophy.

Maximilien Bencze, Baptiste Periou, Isabel Punzón, Inès Barthélémy, Valentina Taglietti, Cyrielle Hou, Louai Zaidan, Kaouthar Kefi, Stéphane Blot, Onnik Agbulut, Marianne Gervais, Geneviève Derumeaux, François-Jérôme Authier, Laurent Tiret, Fréderic Relaix.

   BACKGROUND: Duchenne muscular dystrophy (DMD) is a progressive muscle degenerative disorder, culminating in a complete loss of ambulation, hypertrophic cardiomyopathy and a fatal cardiorespiratory failure. Necroptosis is the form of necrosis that is dependent upon the receptor-interacting protein kinase (RIPK) 3; it is involved in several inflammatory and neurodegenerative conditions. We previously identified RIPK3 as a key player in the acute myonecrosis affecting the hindlimb muscles of the mdx dystrophic mouse model. Whether necroptosis also mediates respiratory and heart disorders in DMD is currently unknown.
METHODS: Evidence of activation of the necroptotic axis was examined in dystrophic tissues from Golden retriever muscular dystrophy (GRMD) dogs and R-DMDdel52 rats. A functional assessment of the involvement of necroptosis in dystrophic animals was performed on mdx mice that were genetically depleted for RIPK3. Dystrophic mice aged from 12 to 18 months were analysed by histology and molecular biology to compare the phenotype of muscles from mdxRipk3+/+ and mdxRipk3-/- mice. Heart function was also examined by echocardiography in 40-week-old mice.
RESULTS: RIPK3 expression in sartorius and biceps femoris muscles from GRMD dogs positively correlated to myonecrosis levels (r = 0.81; P = 0.0076). RIPK3 was also found elevated in the diaphragm (P ≤ 0.05). In the slow-progressing heart phenotype of GRMD dogs, the phosphorylated form of RIPK1 at the Serine 161 site was dramatically increased in cardiomyocytes. A similar p-RIPK1 upregulation characterized the cardiomyocytes of the severe DMDdel52 rat model, associated with a marked overexpression of Ripk1 (P = 0.007) and Ripk3 (P = 0.008), indicating primed activation of the necroptotic pathway in the dystrophic heart. MdxRipk3-/- mice displayed decreased compensatory hypertrophy of the heart (P = 0.014), and echocardiography showed a 19% increase in the relative wall thickness (P < 0.05) and 29% reduction in the left ventricle mass (P = 0.0144). Besides, mdxRipk3-/- mice presented no evidence of a regenerative default or sarcopenia in skeletal muscles, moreover around 50% less affected by fibrosis (P < 0.05).
CONCLUSIONS: Our data highlight molecular and histological evidence that the necroptotic pathway is activated in degenerative tissues from dystrophic animal models, including the diaphragm and the heart. We also provide the genetic proof of concept that selective inhibition of necroptosis in dystrophic condition improves both histological features of muscles and cardiac function, suggesting that prevention of necroptosis is susceptible to providing multiorgan beneficial effects for DMD.

Keywords:  Animal model; Cardiac failure; Duchenne muscular dystrophy; Fibrosis; Myogenesis; Myonecrosis; Necroptosis; Programmed cell death

DOI:  https://doi.org/10.1002/jcsm.13265
J Cachexia Sarcopenia Muscle. 2023 Nov 02.

Lactate promotes myogenesis via activating H3K9 lactylation-dependent up-regulation of Neu2 expression.

Weilong Dai, Gang Wu, Ke Liu, Qianqian Chen, Jingli Tao, Honglin Liu, Ming Shen.

   BACKGROUND: Lactate, a glycolytic metabolite mainly produced in muscles, has been suggested to regulate myoblast differentiation, although the underlying mechanism remains elusive. Recently, lactate-mediated histone lactylation is identified as a novel epigenetic modification that promotes gene transcription.
METHODS: We used mouse C2C12 cell line and 2-month-old male mice as in vitro and in vivo models, respectively. These models were treated with lactate to explore the biological function and latent mechanism of lactate-derived histone lactylation on myogenic differentiation by quantitative real-time PCR, western blotting, immunofluorescence staining, chromatin immunoprecipitation, cleavage under targets and tagmentation assay and RNA sequencing.
RESULTS: Using immunofluorescence staining and western blotting, we proposed that lactylation might occur in the histones. Inhibition of lactate production or intake both impaired myoblast differentiation, accompanied by diminished lactylation in the histones. Using lactylation site-specific antibodies, we demonstrated that lactate preferentially increased H3K9 lactylation (H3K9la) during myoblast differentiation (CT VS 5, 10, 15, 20, 25 mM lactate treatment, P = 0.0012, P = 0.0007, and the rest of all P < 0.0001). Notably, inhibiting H3K9la using P300 antagonist could block lactate-induced myogenesis. Through combined omics analysis using cleavage under targets and tagmentation assay and RNA sequencing, we further identified Neu2 as a potential target gene of H3K9la. IGV software analysis (P = 0.0013) and chromatin immunoprecipitation-qPCR assay (H3K9la %Input, LA group = 9.0076, control group = 2.7184, IgG = 0.3209) confirmed that H3K9la is enriched in the promoter region of Neu2. Moreover, siRNAs or inhibitors against Neu2 both abrogated myoblast differentiation despite lactate treatment, suggesting that Neu2 is required for lactate-mediated myoblast differentiation.
CONCLUSIONS: Our findings provide novel understanding of histone lysine lactylation, suggesting its role in myogenesis, and as potential therapeutic targets for muscle diseases.

Keywords:  Histone lactylation; Lactate; Myoblast differentiation

DOI:  https://doi.org/10.1002/jcsm.13363
Nanomedicine (Lond). 2023 Nov 01.

Muscle-targeted nanoparticles strengthen the effects of small-molecule inhibitors in ameliorating sarcopenia.

Jinyu Wang, Yikang He, Baoyue Wang, Ruian Yin, Biao Chen, Hongxing Wang.

  Background: Sarcopenia is an aging-related degeneration of muscle mass and strength. Small-molecule inhibitor SW033291 has been shown to attenuate muscle atrophy. Targeted nanodrug-delivery systems can improve the efficacy of small-molecule inhibitors. Methods: The skeletal muscle cell-targeted nanoparticle was called AP@SW033291, which consisted of SW033291, modular peptide ASSLNIAGGRRRRRG and PEG-DSPE. Nanoparticles were featured with particle size, fluorescence emission spectra and targeting ability. We also investigated their effects on muscle mass and function. Results: The size of AP@SW033291 was 125.7 nm and it demonstrated targeting effects on skeletal muscle; thus, it could improve muscle mass and muscle function. Conclusion: Nanoparticle AP@SW033291 could become a potential strategy to strengthen the treatment effects of small-molecule inhibitors in sarcopenia.

Keywords:  SAMP8; nanoparticle; sarcopenia; small-molecule inhibitor; targeted therapy

DOI:  https://doi.org/10.2217/nnm-2023-0201
Biochem Biophys Res Commun. 2023 Oct 13. pii: S0006-291X(23)01104-X. [Epub ahead of print]684 149040

Protein-coding potential of non-canonical open reading frames in human transcriptome.

Hitesh Kore, Keshava K Datta, Shivashankar H Nagaraj, Harsha Gowda.

  In recent years, proteogenomics and ribosome profiling studies have identified a large number of proteins encoded by noncoding regions in the human genome. They are encoded by small open reading frames (sORFs) in the untranslated regions (UTRs) of mRNAs and long non-coding RNAs (lncRNAs). These sORF encoded proteins (SEPs) are often <150AA and show poor evolutionary conservation. A subset of them have been functionally characterized and shown to play an important role in fundamental biological processes including cardiac and muscle function, DNA repair, embryonic development and various human diseases. How many novel protein-coding regions exist in the human genome and what fraction of them are functionally important remains a mystery. In this review, we discuss current progress in unraveling SEPs, approaches used for their identification, their limitations and reliability of these identifications. We also discuss functionally characterized SEPs and their involvement in various biological processes and diseases. Lastly, we provide insights into their distinctive features compared to canonical proteins and challenges associated with annotating these in protein reference databases.

Keywords:  Non-coding RNAs; Novel proteins; Protein-coding potential; SEPs

DOI:  https://doi.org/10.1016/j.bbrc.2023.09.068
Exp Anim. 2023 Nov 01.

Ivabradine ameliorates cardiomyopathy progression in a Duchenne muscular dystrophy model rat.

Ryota Tochinai, Koichi Kimura, Takeru Saika, Wataru Fujii, Hiroyuki Morita, Koki Nakanishi, Yoshiharu Tsuru, Shin-Ichi Sekizawa, Keitaro Yamanouchi, Masayoshi Kuwahara.

  Duchenne muscular dystrophy (DMD) is an X-linked recessive myopathy caused by dystrophin mutations. Inevitable progressive cardiomyopathy is a current leading cause of premature death although respiratory management has improved the prognosis of patients with DMD. Recent evidence shows that reducing the heart rate is expected as one of the promising strategies for heart failure treatment, but administering a sufficient dose of β-blocker for patients with DMD with tachycardia is difficult because of their low blood pressure (BP). Thus, this study aimed to clarify the role of ivabradine, which suppresses cardiac sinus node pacemakers without decreasing BP, in ameliorating cardiomyopathy progression in a rat model with DMD. A trans-oral single ivabradine administration demonstrated a declined dose-dependent heart rate without any significant BP reduction. Trans-gastric repeated administrations of 5 mg/kg of ivabradine twice a day for 3 months showed ameliorated cardiomyopathy in DMD rats based on echocardiography and histopathological observations (left ventricular dysfunction, right ventricular dysfunction, and myocardial fibrosis) as compared with vehicle administration. Our finding indicates that ivabradine is expected as another treatment choice for patients with DMD having tachycardia.

Keywords:  HCN channel; dystrophin deficiency; echocardiography; heart; tachycardia

DOI:  https://doi.org/10.1538/expanim.23-0087