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1.
Nat Commun ; 12(1): 89, 2021 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-33397958

RESUMO

The RNA-binding protein QKI belongs to the hnRNP K-homology domain protein family, a well-known regulator of pre-mRNA alternative splicing and is associated with several neurodevelopmental disorders. Qki is found highly expressed in developing and adult hearts. By employing the human embryonic stem cell (hESC) to cardiomyocyte differentiation system and generating QKI-deficient hESCs (hESCs-QKIdel) using CRISPR/Cas9 gene editing technology, we analyze the physiological role of QKI in cardiomyocyte differentiation, maturation, and contractile function. hESCs-QKIdel largely maintain normal pluripotency and normal differentiation potential for the generation of early cardiogenic progenitors, but they fail to transition into functional cardiomyocytes. In this work, by using a series of transcriptomic, cell and biochemical analyses, and the Qki-deficient mouse model, we demonstrate that QKI is indispensable to cardiac sarcomerogenesis and cardiac function through its regulation of alternative splicing in genes involved in Z-disc formation and contractile physiology, suggesting that QKI is associated with the pathogenesis of certain forms of cardiomyopathies.


Assuntos
Processamento Alternativo/genética , Desenvolvimento Muscular/genética , Contração Miocárdica/genética , Precursores de RNA/metabolismo , Proteínas de Ligação a RNA/metabolismo , Actinina/genética , Animais , Diferenciação Celular/genética , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Camundongos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Miócitos Cardíacos/ultraestrutura , Precursores de RNA/genética , Proteínas de Ligação a RNA/genética , Transcriptoma/genética
2.
J Surg Res ; 257: 56-68, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32818785

RESUMO

BACKGROUND: Burn injuries (BIs) due to scalding are one of the most common accidents among children. BIs greater than 40% of total body surface area are considered extensive and result in local and systemic response. We sought to assess morphological and myogenic mechanisms through both short- and long-term intensive insulin therapies that affect the skeletal muscle after extensive skin BI in young rats. MATERIALS AND METHODS: Wistar rats aged 21 d were distributed into four groups: control (C), control with insulin (C + I), scald burn injury (SI), and SI with insulin (SI + I). The SI groups were submitted to a 45% total body surface area burn, and the C + I and SI + I groups received insulin (5 UI/Kg/d) for 4 or 14 d. Glucose tolerance and the homeostatic model assessment of insulin resistance index were determined. Gastrocnemius muscles were analyzed for histopathological, morphometric, and immunohistochemical myogenic parameters (Pax7, MyoD, and MyoG); in addition, the expression of genes related to muscle atrophy (MuRF1 and MAFbx) and its regulation (IGF-1) were also assessed. RESULTS: Short-term treatment with insulin favored muscle regeneration by primary myogenesis and decreased muscle atrophy in animals with BIs, whereas the long-term treatment modulated myogenesis by increasing the MyoD protein. Both treatments improved histopathological parameters and secondary myogenesis by increasing the MyoG protein. CONCLUSIONS: Treatment with insulin benefits myogenic parameters during regeneration and modulates MuRF1, an important mediator of muscle atrophy.


Assuntos
Queimaduras/complicações , Insulina/administração & dosagem , Desenvolvimento Muscular/efeitos dos fármacos , Atrofia Muscular/prevenção & controle , Animais , Glicemia/análise , Superfície Corporal , Queimaduras/patologia , Queimaduras/fisiopatologia , Expressão Gênica/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/genética , Masculino , Proteínas Musculares/genética , Músculo Esquelético/química , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Atrofia Muscular/etiologia , Atrofia Muscular/genética , Proteína MyoD/análise , Miogenina/análise , Fatores de Transcrição Box Pareados/análise , Ratos , Ratos Wistar , Proteínas Ligases SKP Culina F-Box/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética
3.
Gene ; 768: 145319, 2021 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-33246031

RESUMO

Myogenesis is a complex, regulated process that involves myoblast proliferation, migration, adhesion, and fusion into myotubes. To investigate early development of embryonic muscles and the expression of regulatory genes during myogenesis in chicken, quail and their hybrids, meat-breeding cocks and egg-breeding cocks were selected as male parents, quails were used as female parents. Their offspring were meat and egg hybrids via Artificial insemination. We measured expression of MUSTN1, IGF-1, and PDK4 using qRT-PCR. We examined muscle fiber diameter using scanning electron microscopy. The results showed that muscle development was two days slower in chicken, egg hybrid, and meat hybrid than in quail. Muscle fiber spacing was the largest in chicken, followed by meat hybrid, egg hybrid, and quail. A similar trend was obtained for muscle fiber diameter. Additionally, muscle fiber diameter increased with embryogenesis. The sarcomere was present on day 17 of incubation in quail, but not in the other species. MUSTN1 could up-regulated IGF-1 by activating PI3K/Akt. IGF-1 expression was consistent with myoblast proliferation and myotube fusion. PDK4 was expressed from E7 to E17. The first peak was reached on E10, egg hybrid and meat hybrid reached their peak at E15. PDK4 is involved in the early proliferation and differentiation of muscle, thereby affecting muscle growth and development. Our findings demonstrated that MUSTN1, IGF-1 and PDK4 genes are expressed to varying levels in breast muscle of chicken, quail, egg hybrid and meat hybrid during the embryonic period. Interestingly, with increasing embryonic age, muscle development was approximately 48 h faster in quail than in other species. We speculated that MUSTN1, IGF-1 and PDK4 genes may be the main candidate genes that cause differences in poultry muscle traits, but the molecular regulation mechanisms need to be further studied. Our findings shed some light on the avian embryo muscle formation and molecular breeding of poultry muscle traits, which provide theoretical basis for poultry breeding.


Assuntos
Quimera/embriologia , Fator de Crescimento Insulin-Like I/genética , Músculo Esquelético/crescimento & desenvolvimento , Proteínas Nucleares/genética , /genética , Animais , Cruzamento , Galinhas , Quimera/genética , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Microscopia de Força Atômica , Desenvolvimento Muscular , Músculo Esquelético/metabolismo , Codorniz
4.
Nucleic Acids Res ; 48(22): 12943-12956, 2020 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-33270893

RESUMO

Long noncoding (lnc)RNAs potently regulate gene expression programs in physiology and disease. Here, we describe a key function for lncRNA OIP5-AS1 in myogenesis, the process whereby myoblasts differentiate into myotubes during muscle development and muscle regeneration after injury. In human myoblasts, OIP5-AS1 levels increased robustly early in myogenesis, and its loss attenuated myogenic differentiation and potently reduced the levels of the myogenic transcription factor MEF2C. This effect relied upon the partial complementarity of OIP5-AS1 with MEF2C mRNA and the presence of HuR, an RNA-binding protein (RBP) with affinity for both transcripts. Remarkably, HuR binding to MEF2C mRNA, which stabilized MEF2C mRNA and increased MEF2C abundance, was lost after OIP5-AS1 silencing, suggesting that OIP5-AS1 might serve as a scaffold to enhance HuR binding to MEF2C mRNA, in turn increasing MEF2C production. These results highlight a mechanism whereby a lncRNA promotes myogenesis by enhancing the interaction of an RBP and a myogenic mRNA.


Assuntos
Desenvolvimento Muscular/genética , RNA Longo não Codificante/genética , Regeneração/genética , Diferenciação Celular/genética , Linhagem Celular , Proliferação de Células/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Fatores de Transcrição MEF2/genética , Mioblastos/metabolismo , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Transdução de Sinais/genética
5.
Nat Commun ; 11(1): 6222, 2020 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-33277476

RESUMO

Using Hi-C, promoter-capture Hi-C (pCHi-C), and other genome-wide approaches in skeletal muscle progenitors that inducibly express a master transcription factor, Pax7, we systematically characterize at high-resolution the spatio-temporal re-organization of compartments and promoter-anchored interactions as a consequence of myogenic commitment and differentiation. We identify key promoter-enhancer interaction motifs, namely, cliques and networks, and interactions that are dependent on Pax7 binding. Remarkably, Pax7 binds to a majority of super-enhancers, and together with a cadre of interacting transcription factors, assembles feed-forward regulatory loops. During differentiation, epigenetic memory and persistent looping are maintained at a subset of Pax7 enhancers in the absence of Pax7. We also identify and functionally validate a previously uncharacterized Pax7-bound enhancer hub that regulates the essential myosin heavy chain cluster during skeletal muscle cell differentiation. Our studies lay the groundwork for understanding the role of Pax7 in orchestrating changes in the three-dimensional chromatin conformation in muscle progenitors.


Assuntos
Diferenciação Celular/genética , Cromatina/genética , Células-Tronco Embrionárias Murinas/metabolismo , Desenvolvimento Muscular/genética , Músculo Esquelético/metabolismo , Células 3T3-L1 , Animais , Células Cultivadas , Cromatina/metabolismo , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Redes Reguladoras de Genes , Humanos , Camundongos , Músculo Esquelético/citologia , Fator de Transcrição PAX7/genética , Fator de Transcrição PAX7/metabolismo
6.
Rev Saude Publica ; 54: 113, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33237171

RESUMO

OBJECTIVE: To analyze the effects of early determinants on adolescent fat-free mass. METHODS: A cohort study with 579 adolescents evaluated at birth and adolescence in a birth cohort in São Luís, Maranhão. In the proposed model, estimated by structural equation modeling, socioeconomic status (SES) at birth, maternal age, pregestational body mass index (BMI), gestational smoking, gestational weight gain, type of delivery, gestational age, sex of the newborn, length and weight at birth, adolescent socioeconomic status, "neither study/nor work" generation, adolescent physical activity level and alcohol consumption were tested as early determinants of adolescent fat-free mass (FFM). RESULTS: A higher pregestational BMI resulted in higher FFM in adolescence (Standardized Coefficient, SC = 0.152; p < 0.001). Being female implied a lower FFM in adolescence (SC = -0.633; p < 0.001). The negative effect of gender on FFM was direct (SC = -0.523; p < 0.001), but there was an indirect negative effect via physical activity level (SC = -0.085; p < 0.001). Women were less active (p < 0.001). An increase of 0.5 kg (1 Standard Deviation, SD) in birth weight led to a gain of 0.25 kg/m2 (0.106 SD) in adolescent FFM index (p = 0.034). Not studying or working had a negative effect on the adolescent's FFM (SC = -0.106; p = 0.015). Elevation of 1 SD in the adolescent's physical activity level represented an increase of 0.5 kg/m2 (0.207 SD) in FFM index (p < 0.001). CONCLUSIONS: The early determinants with the greatest effects on adolescent FFM are gender, adolescent physical activity level, pregestational BMI, birth weight and belonging to the "neither-nor" generation.


Assuntos
Adiposidade , Desenvolvimento do Adolescente/fisiologia , Peso ao Nascer , Composição Corporal , Gordura Subcutânea/crescimento & desenvolvimento , Adolescente , Saúde do Adolescente , Índice de Massa Corporal , Brasil , Estudos de Coortes , Feminino , Idade Gestacional , Humanos , Recém-Nascido , Análise de Classes Latentes , Masculino , Desenvolvimento Muscular , Determinantes Sociais da Saúde , Fatores Socioeconômicos
8.
Am J Physiol Cell Physiol ; 319(6): C1020-C1028, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32936696

RESUMO

Myoblast differentiation is a crucial process for myogenesis. Mitochondria function as an energy-providing machine that is critical to this process, and mitochondrial dysfunction can prevent myoblasts from fusing into myotubes. However, the molecular mechanisms underlying the dynamic regulation of mitochondrial networks remain poorly understood. In the present study, we found that the PTEN induced kinase 1 (PINK1)/Parkin (an E3 ubiquitin-protein ligase) pathway is activated at the early stage of myoblast differentiation. Moreover, downregulation of mitofusin 2 (Mfn2) and increased dynamin-related protein 1 (Drp1) resulted in loosely formed mitochondria during this period. Furthermore, selective knockdown of the mitochondrial matrix protein Lon peptidase-1 (LonP1) at the early stage of myoblast differentiation induced mitochondrial depolarization and suppressed the PINK1/Parkin pathway and reduced Mfn2 and Drp1 levels, which blocked mitochondrial remodeling and myoblast differentiation. Overall, these data demonstrate that LonP1 promotes myoblast differentiation by regulating PINK1/Parkin-mediated mitochondrial remodeling.


Assuntos
Proteases Dependentes de ATP/metabolismo , Diferenciação Celular/fisiologia , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Mioblastos/metabolismo , Proteínas Quinases/metabolismo , Transdução de Sinais/fisiologia , Ubiquitina-Proteína Ligases/metabolismo , Animais , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Desenvolvimento Muscular/fisiologia
9.
PLoS Genet ; 16(9): e1009044, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32991581

RESUMO

The transcription factor nuclear factor I-A (NFIA) is a regulator of brown adipocyte differentiation. Here we show that the C-terminal 17 amino acid residues of NFIA (which we call pro#3 domain) are required for the transcriptional activity of NFIA. Full-length NFIA-but not deletion mutant lacking pro#3 domain-rescued impaired expression of PPARγ, the master transcriptional regulator of adipogenesis and impaired adipocyte differentiation in NFIA-knockout cells. Mechanistically, the ability of NFIA to penetrate chromatin and bind to the crucial Pparg enhancer is mediated through pro#3 domain. However, the deletion mutant still binds to Myod1 enhancer to repress expression of MyoD, the master transcriptional regulator of myogenesis as well as proximally transcribed non-coding RNA called DRReRNA, via competition with KLF5 in terms of enhancer binding, leading to suppression of myogenic gene program. Therefore, the negative effect of NFIA on the myogenic gene program is, at least partly, independent of the positive effect on PPARγ expression and its downstream adipogenic gene program. These results uncover multiple ways of action of NFIA to ensure optimal regulation of brown and beige adipocyte differentiation.


Assuntos
Adipócitos Bege/citologia , Adipócitos Marrons/citologia , Adipogenia/fisiologia , Desenvolvimento Muscular/fisiologia , Fatores de Transcrição NFI/metabolismo , Adipócitos Bege/fisiologia , Adipócitos Marrons/fisiologia , Adipogenia/genética , Animais , Diferenciação Celular/genética , Cromatina/genética , Cromatina/metabolismo , Regulação da Expressão Gênica , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Desenvolvimento Muscular/genética , Proteína MyoD/genética , Miogenina/genética , Fatores de Transcrição NFI/genética , PPAR gama/genética , PPAR gama/metabolismo , Prolina , Domínios Proteicos
10.
PLoS One ; 15(9): e0238441, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32881965

RESUMO

Limb-girdle muscular dystrophy type 2B (LGMD2B) is caused by mutations in the dysferlin gene, resulting in non-functional dysferlin, a key protein found in muscle membrane. Treatment options available for patients are chiefly palliative in nature and focus on maintaining ambulation. Our hypothesis is that galectin-1 (Gal-1), a soluble carbohydrate binding protein, increases membrane repair capacity and myogenic potential of dysferlin-deficient muscle cells and muscle fibers. To test this hypothesis, we used recombinant human galectin-1 (rHsGal-1) to treat dysferlin-deficient models. We show that rHsGal-1 treatments of 48 h-72 h promotes myogenic maturation as indicated through improvements in size, myotube alignment, myoblast migration, and membrane repair capacity in dysferlin-deficient myotubes and myofibers. Furthermore, increased membrane repair capacity of dysferlin-deficient myotubes, independent of increased myogenic maturation is apparent and co-localizes on the membrane of myotubes after a brief 10min treatment with labeled rHsGal-1. We show the carbohydrate recognition domain of Gal-1 is necessary for observed membrane repair. Improvements in membrane repair after only a 10 min rHsGal-1treatment suggest mechanical stabilization of the membrane due to interaction with glycosylated membrane bound, ECM or yet to be identified ligands through the CDR domain of Gal-1. rHsGal-1 shows calcium-independent membrane repair in dysferlin-deficient and wild-type myotubes and myofibers. Together our novel results reveal Gal-1 mediates disease pathologies through both changes in integral myogenic protein expression and mechanical membrane stabilization.


Assuntos
Disferlina/genética , Galectina 1/farmacologia , Distrofia Muscular do Cíngulo dos Membros/terapia , Animais , Linhagem Celular , Modelos Animais de Doenças , Disferlina/metabolismo , Galectina 1/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Membranas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Desenvolvimento Muscular/genética , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Distrofia Muscular do Cíngulo dos Membros/metabolismo , Miofibrilas/metabolismo
11.
PLoS One ; 15(9): e0239047, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32941492

RESUMO

Muscle aging is accompanied by blunted muscle regeneration in response to injury and disuse. Oxidative stress likely underlies this diminished response, but muscle redox sensors that act in regeneration have not yet been characterized. Calmodulin contains multiple redox sensitive methionines whose oxidation alters the regulation of numerous cellular targets. We have used the CRISPR-Cas9 system to introduce a single amino acid substitution M109Q that mimics oxidation of methionine to methionine sulfoxide in one or both alleles of the CALM1 gene, one of three genes encoding the muscle regulatory protein calmodulin, in C2C12 mouse myoblasts. When signaled to undergo myogenesis, mutated myoblasts failed to differentiate into myotubes. Although early myogenic regulatory factors were present, cells with the CALM1 M109Q mutation in one or both alleles were unable to withdraw from the cell cycle and failed to express late myogenic factors. We have shown that a single oxidative modification to a redox-sensitive muscle regulatory protein can halt myogenesis, suggesting a molecular target for mitigating the impact of oxidative stress in age-related muscle degeneration.


Assuntos
Calmodulina/metabolismo , Desenvolvimento Muscular/fisiologia , Animais , Calmodulina/genética , Calmodulina/fisiologia , Diferenciação Celular/fisiologia , Camundongos , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Mioblastos/metabolismo , Oxirredução , Estresse Oxidativo/fisiologia
12.
Am J Physiol Endocrinol Metab ; 319(6): E1008-E1018, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32954829

RESUMO

Skeletal muscle is sensitive to environmental cues that are first present in utero. Maternal overnutrition is a model of impaired muscle development leading to structural and metabolic dysfunction in adult life. In this study, we investigated the effect of an obesogenic maternal environment on growth and postnatal myogenesis in the offspring. Male C57BL/6J mice born to chow- or high-fat-diet-fed mothers were allocated to four different groups at the end of weaning. For the following 10 wk, half of the pups were maintained on the same diet as their mother and half of the pups were switched to the other diet (chow or high-fat). At 12 wk of age, muscle injury was induced using an intramuscular injection of barium chloride. Seven days later, mice were humanely killed and muscle tissue was harvested. A high-fat maternal diet impaired offspring growth patterns and downregulated satellite cell activation and markers of postnatal myogenesis 7 days after injury without altering the number of newly synthetized fibers over the whole 7-day period. Importantly, a healthy postnatal diet could not reverse any of these effects. In addition, we demonstrated that postnatal myogenesis was associated with a diet-independent upregulation of three miRNAs, mmu-miR-31-5p, mmu-miR-136-5p, and mmu-miR-296-5p. Furthermore, in vitro analysis confirmed the role of these miRNAs in myocyte proliferation. Our findings are the first to demonstrate that maternal overnutrition impairs markers of postnatal myogenesis in the offspring and are particularly relevant to today's society where the incidence of overweight/obesity in women of childbearing age is increasing.


Assuntos
Dieta Hiperlipídica , Crescimento e Desenvolvimento/fisiologia , Desenvolvimento Muscular/fisiologia , Efeitos Tardios da Exposição Pré-Natal , Células Satélites de Músculo Esquelético/fisiologia , Animais , Biomarcadores/análise , Biomarcadores/metabolismo , Dieta Hiperlipídica/efeitos adversos , Feminino , Masculino , Fenômenos Fisiológicos da Nutrição Materna , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Obesidade/etiologia , Obesidade/fisiopatologia , Hipernutrição/complicações , Hipernutrição/fisiopatologia , Gravidez , Complicações na Gravidez/etiologia , Complicações na Gravidez/fisiopatologia , Efeitos Tardios da Exposição Pré-Natal/etiologia , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia
13.
Sci Rep ; 10(1): 15086, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32934320

RESUMO

The myogenic factor MyoD regulates skeletal muscle differentiation by interacting with a variety of chromatin-modifying complexes. Although MyoD can induce and maintain chromatin accessibility at its target genes, its binding and trans-activation ability can be limited by some types of not fully characterized epigenetic constraints. In this work we analysed the role of PARP1 in regulating MyoD-dependent gene expression. PARP1 is a chromatin-associated enzyme, playing a well recognized role in DNA repair and that is implicated in transcriptional regulation. PARP1 affects gene expression through multiple mechanisms, often involving the Poly(ADP-ribosyl)ation of chromatin proteins. In line with PARP1 down-regulation during differentiation, we observed that PARP1 depletion boosts the up-regulation of MyoD targets, such as p57, myogenin, Mef2C and p21, while its re-expression reverts this effect. We also found that PARP1 interacts with some MyoD-binding regions and that its presence, independently of the enzymatic activity, interferes with MyoD recruitment and gene induction. We finally suggest a relationship between the binding of PARP1 and the loss of the activating histone modification H3K4me3 at MyoD-binding regions. This work highlights not only a novel player in the epigenetic control of myogenesis, but also a repressive and catalytic-independent mechanisms by which PARP1 regulates transcription.


Assuntos
Diferenciação Celular/genética , Regulação da Expressão Gênica/genética , Músculos/fisiologia , Proteína MyoD/genética , Poli(ADP-Ribose) Polimerase-1/genética , Animais , Linhagem Celular , Cromatina/genética , Reparo do DNA/genética , Histonas/genética , Camundongos , Desenvolvimento Muscular/genética , Transcrição Genética/genética , Regulação para Cima/genética
14.
Toxicology ; 443: 152561, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32800841

RESUMO

Methylmercury (MeHg) is a ubiquitous environmental contaminant and developmental toxicant known to cause a variety of persistent motor and cognitive deficits. While previous research has focused predominantly on neurotoxic MeHg effects, emerging evidence points to a myotoxic role whereby MeHg induces defects in muscle development and maintenance. A genome wide association study for developmental sensitivity to MeHg in Drosophila has revealed several conserved muscle morphogenesis candidate genes that function in an array of processes from myoblast migration and fusion to myotendinous junction (MTJ) formation and myofibrillogenesis. Here, we investigated candidates for a role in mediating MeHg disruption of muscle development by evaluating morphological and functional phenotypes of the indirect flight muscles (IFMs) in pupal and adult flies following 0, 5, 10, and 15 µM MeHg exposure via feeding at the larval stage. Developmental MeHg exposure induced a dose-dependent increase in muscle detachments (myospheres) within dorsal bundles of the IFMs, which paralleled reductions eclosion and adult flight behaviors. These effects were selectively phenocopied by altered expression of kon-tiki (kon), a chondroitin sulfate proteoglycan 4/NG2 homologue and a central component of MTJ formation. MeHg elevated kon transcript expression at a crucial window of IFM development and transgene overexpression of kon could also phenocopy myosphere phenotypes and eclosion and flight deficits. Finally, the myosphere phenotype resulting from 10 µM MeHg was partially rescued in a background of reduced kon expression using a targeted RNAi approach. Our findings implicate a component of the MTJ as a MeHg toxicity target which broaden the understanding of how motor deficits can emerge from early life MeHg exposure.


Assuntos
Drosophila/efeitos dos fármacos , Compostos de Metilmercúrio/toxicidade , Desenvolvimento Muscular/efeitos dos fármacos , Miotoxicidade , Animais , Comportamento Animal/efeitos dos fármacos , Drosophila/genética , Drosophila/crescimento & desenvolvimento , Proteínas de Drosophila/genética , Feminino , Voo Animal/efeitos dos fármacos , Larva/efeitos dos fármacos , Larva/genética , Larva/crescimento & desenvolvimento , Masculino , Desenvolvimento Muscular/genética , Proteínas do Tecido Nervoso/genética , Pupa/efeitos dos fármacos , Pupa/genética , Pupa/crescimento & desenvolvimento
16.
Cell Prolif ; 53(9): e12879, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32770602

RESUMO

OBJECTIVE: This study aims to clarify the mechanisms underlying transcriptional regulation and regulatory roles of lncRNAs in skeletal muscle cell differentiation. METHODS: We analysed the expression patterns of lncRNAs via time-course RNA-seq. Then, we further combined the ATAC-seq and ChIP-seq to investigate the governing mechanisms of transcriptional regulation of differentially expressed (DE) lncRNAs. Weighted correlation network analysis and GO analysis were conducted to identify the transcription factor (TF)-lncRNA pairs related to skeletal muscle cell differentiation. RESULTS: We identified 385 DE lncRNAs during C2C12 differentiation, the transcription of which is determined by chromatin states around their transcriptional start sites. The TF-lncRNA correlation network showed substantially concordant changes in DE lncRNAs between C2C12 differentiation and satellite cell rapid growth stages. Moreover, the up-regulated lncRNAs showed a significant decrease following the differentiation capacity of satellite cells, which gradually declines during skeletal muscle development. Notably, inhibition of the lncRNA Atcayos and Trp53cor1 led to the delayed differentiation of satellite cells. Those lncRNAs were significantly up-regulated during the rapid growth stage of satellite cells (4-6 weeks) and down-regulated with reduced differentiation capacity (8-12 weeks). It confirms that these lncRNAs are positively associated with myogenic differentiation of satellite cells during skeletal muscle development. CONCLUSIONS: This study extends the understanding of mechanisms governing transcriptional regulation of lncRNAs and provides a foundation for exploring their functions in skeletal muscle cell differentiation.


Assuntos
Diferenciação Celular , Cromatina/genética , Desenvolvimento Muscular , Músculo Esquelético/citologia , RNA Longo não Codificante/genética , Animais , Linhagem Celular , Proliferação de Células , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Músculo Esquelético/metabolismo , Células Satélites de Músculo Esquelético/citologia , Células Satélites de Músculo Esquelético/metabolismo , Ativação Transcricional
17.
Biochim Biophys Acta Gen Subj ; 1864(12): 129707, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32810562

RESUMO

BACKGROUND: Heparan sulfate (HS) is a sulfated linear polysaccharide on cell surfaces that plays an important role in physiological processes. HS is present in skeletal muscles but its detailed role in this tissue remains unclear. METHODS: We examined the role of HS in the differentiation of C2C12 cells, a mouse myoblast cell line. We also phenotyped the impact of HS deletion in mouse skeletal muscles on their functions by using Cre-loxP system. RESULTS: CRISPR-Cas9-dependent HS deletion or pharmacological removal of HS dramatically impaired myoblast differentiation of C2C12 cells. To confirm the importance of HS in vivo, we deleted Ext1, which encodes an enzyme essential for HS biosynthesis, specifically in the mouse skeletal muscles (referred to as mExt1CKO mice). Treadmill and wire hang tests demonstrated that mExt1CKO mice exhibited muscle weakness. The contraction of isolated soleus muscles from mExt1CKO mice was also impaired. Morphological examination of mExt1CKO muscle tissue under light and electron microscopes revealed smaller cross sectional areas and thinner myofibrils. Finally, a model of muscle regeneration following BaCl2 injection into the tibialis anterior muscle of mice demonstrated that mExt1CKO mice had reduced expression of myosin heavy chain and an increased number of centronucleated cells. This indicates that muscle regeneration after injury was attenuated in the absence of HS expression in muscle cells. SIGNIFICANCE: These results demonstrate that HS plays an important role in skeletal muscle function by promoting differentiation.


Assuntos
Heparitina Sulfato/metabolismo , Desenvolvimento Muscular , Músculo Esquelético/fisiologia , Mioblastos/citologia , Animais , Sistemas CRISPR-Cas , Diferenciação Celular , Linhagem Celular , Heparitina Sulfato/antagonistas & inibidores , Heparitina Sulfato/genética , Camundongos , Atividade Motora , Músculo Esquelético/citologia , Mioblastos/metabolismo
18.
Am J Physiol Renal Physiol ; 319(4): F624-F635, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32830539

RESUMO

Recently, we reported a mutation in γ-adducin (ADD3) was associated with an impaired myogenic response of the afferent arteriole and hypertension-induced chronic kidney disease (CKD) in fawn hooded hypertensive (FHH) rats. However, the mechanisms by which altered renal blood flow (RBF) autoregulation promotes hypertension-induced renal injury remain to be determined. The present study compared the time course of changes in renal hemodynamics and the progression of CKD during the development of DOCA-salt hypertension in FHH 1BN congenic rats [wild-type (WT)] with an intact myogenic response versus FHH 1BN Add3KO (Add3KO) rats, which have impaired myogenic response. RBF was well autoregulated in WT rats but not in Add3KO rats. Glomerular capillary pressure rose by 6 versus 14 mmHg in WT versus Add3KO rats when blood pressure increased from 100 to 150 mmHg. After 1 wk of hypertension, glomerular filtration rate increased by 38% and glomerular nephrin expression decreased by 20% in Add3KO rats. Neither were altered in WT rats. Proteinuria doubled in WT rats versus a sixfold increase in Add3KO rats. The degree of renal injury was greater in Add3KO than WT rats after 3 wk of hypertension. RBF, glomerular filtration rate, and glomerular capillary pressure were lower by 20%, 28%, and 19% in Add3KO rats than in WT rats, which was associated with glomerular matrix expansion and loss of capillary filtration area. The results indicated that impaired RBF autoregulation and eutrophic remodeling of preglomerular arterioles increase the transmission of pressure to glomeruli, which induces podocyte loss and accelerates the progression of CKD in hypertensive Add3KO rats.


Assuntos
Pressão Sanguínea , Taxa de Filtração Glomerular , Hipertensão/complicações , Glomérulos Renais/irrigação sanguínea , Proteinúria/etiologia , Circulação Renal , Insuficiência Renal Crônica/etiologia , Animais , Arteríolas/metabolismo , Arteríolas/fisiopatologia , Proteínas de Ligação a Calmodulina/genética , Proteínas de Ligação a Calmodulina/metabolismo , Acetato de Desoxicorticosterona , Modelos Animais de Doenças , Progressão da Doença , Homeostase , Hipertensão/genética , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Masculino , Desenvolvimento Muscular , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiopatologia , Proteinúria/genética , Proteinúria/metabolismo , Proteinúria/fisiopatologia , Ratos Transgênicos , Insuficiência Renal Crônica/genética , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/fisiopatologia , Cloreto de Sódio na Dieta , Remodelação Vascular
19.
Arterioscler Thromb Vasc Biol ; 40(10): 2454-2467, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32787524

RESUMO

OBJECTIVE: There has been little success in translating preclinical studies of mouse hind limb ischemia into benefit for patients with peripheral artery disease. Using systematic strategies, we sought to define the injury and angiogenesis landscapes in mice subjected to hind limb ischemia and ascertain whether published studies to date have used an analysis strategy concordant with these data. Approach and Results: Maps of ischemic injury were generated from 22 different hind limb muscles and 33 muscle territories in 12-week-old C57BL/6 mice, based on loss or centralization of myofiber nuclei. Angiogenesis was similarly mapped based on CD (cluster of differentiation) 31-positive capillary content. Only 10 of 33 muscle territories displayed consistent muscle injury, with the distal anterior hind limb muscles most reliably injured. Angiogenesis was patchy and exclusively associated with zones of regenerated muscle (central nuclei). Angiogenesis was not observed in normal appearing muscle, necrotic muscle, or injury border zones. Systematic review of mouse hind limb angiogenesis studies identified 5147 unique publications, of which 509 met eligibility criteria for analysis. Only 7% of these analyzed manuscripts evaluated angiogenesis in distal anterior hind limb muscles and only 15% consistently examined for angiogenesis in zones of muscle regeneration. CONCLUSIONS: In 12-week C57BL/6 mice, angiogenesis postfemoral artery excision proceeds exclusively in zones of muscle regeneration. Only a minority of studies to date have analyzed angiogenesis in regions of demonstrably regenerating muscle or in high-likelihood territories. Quality assurance standards, informed by the atlas and mapping data herein, could augment data reliability and potentially help translate mouse hind limb ischemia studies to patient care.


Assuntos
Isquemia/fisiopatologia , Músculo Esquelético/irrigação sanguínea , Neovascularização Fisiológica , Projetos de Pesquisa/normas , Animais , Confiabilidade dos Dados , Modelos Animais de Doenças , Membro Posterior , Isquemia/patologia , Masculino , Camundongos Endogâmicos C57BL , Desenvolvimento Muscular , Músculo Esquelético/patologia , Necrose , Regeneração , Fluxo Sanguíneo Regional , Fatores de Tempo
20.
J Anim Sci ; 98(9)2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32841352

RESUMO

The study was conducted to determine the effects of mineral methionine hydroxy analog chelate (MMHAC) partially replacing inorganic trace minerals in sow diets on epigenetic and transcriptional changes in the muscle and jejunum of progeny. The MMHAC is zinc (Zn), manganese (Mn), and copper (Cu) chelated with methionine hydroxy analog (Zn-, Mn-, and Cu-methionine hydroxy analog chelate [MHAC]). On day 35 of gestation, 60 pregnant sows were allotted to two dietary treatments in a randomized completed block design using parity as a block: 1) ITM: inorganic trace minerals with zinc sulfate (ZnSO4), manganese oxide (MnO), and copper sulfate (CuSO4) and 2) CTM: 50% of ITM was replaced with MMHAC (MINTREX trace minerals, Novus International Inc., St Charles, MO). Gestation and lactation diets were formulated to meet or exceed NRC requirements. On days 1 and 18 of lactation, milk samples from 16 sows per treatment were collected to measure immunoglobulins (immunoglobulin G, immunoglobulin A, and immunoglobulin M) and micromineral concentrations. Two pigs per litter were selected to collect blood to measure the concentration of immunoglobulins in the serum, and then euthanized to collect jejunal mucosa, jejunum tissues, and longissimus muscle to measure global deoxyribonucleic acid methylation, histone acetylation, cytokines, and jejunal histomorphology at birth and day 18 of lactation. Data were analyzed using Proc MIXED of SAS. Supplementation of MMHAC tended to decrease (P = 0.059) body weight (BW) loss of sows during lactation and tended to increase (P = 0.098) piglet BW on day 18 of lactation. Supplementation of MMHAC increased (P < 0.05) global histone acetylation and tended to decrease myogenic regulatory factor 4 messenger ribonucleic acid (mRNA; P = 0.068) and delta 4-desaturase sphingolipid1 (DEGS1) mRNA (P = 0.086) in longissimus muscle of piglets at birth. Supplementation of MMHAC decreased (P < 0.05) nuclear factor kappa B mRNA in the jejunum and DEGS1 mRNA in longissimus muscle and tended to decrease mucin-2 (MUC2) mRNA (P = 0.057) and transforming growth factor-beta 1 (TGF-ß1) mRNA (P = 0.057) in the jejunum of piglets on day 18 of lactation. There were, however, no changes in the amounts of tumor necrosis factor-alpha, interleukin-8, TGF-ß, MUC2, and myogenic factor 6 in the tissues by MMHAC. In conclusion, maternal supplementation of MMHAC could contribute to histone acetylation and programming in the fetus, which potentially regulates intestinal health and skeletal muscle development of piglets at birth and weaning, possibly leading to enhanced growth of their piglets.


Assuntos
Imunoglobulinas/sangue , Metionina/análogos & derivados , Minerais/metabolismo , Suínos/fisiologia , Ração Animal/análise , Animais , Dieta/veterinária , Feminino , Lactação , Metionina/farmacologia , Desenvolvimento Muscular/efeitos dos fármacos , Paridade , Gravidez , Suínos/genética , Suínos/crescimento & desenvolvimento , Oligoelementos/farmacologia , Desmame
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