scholarly journals Streptozotocin induces G2 arrest in skeletal muscle myoblasts and impairs muscle growth in vivo

2007 ◽  
Vol 292 (3) ◽  
pp. C1033-C1040 ◽  
Author(s):  
Adam P. W. Johnston ◽  
Jonathan E. Campbell ◽  
Jeremy G. Found ◽  
Michael C. Riddell ◽  
Thomas J. Hawke
Keyword(s):  
Skeletal Muscle ◽  
Muscle Growth ◽  
Phase Cell ◽  
Saline Control ◽  
Dependent Manner ◽  
Β Cell ◽  
Cross Sectional ◽  
M Phase

Streptozotocin (STZ) is used extensively to induce pancreatic β-cell death and ultimately diabetes mellitus in animal models. However, the direct effects of STZ on muscle are largely unknown. To delineate the effects of STZ from the effects of hypoinsulinemia/hyperglycemia, we injected young rats with 1) saline (control), 2) STZ (120 mg/kg) or 3) STZ and insulin (STZ-INS; to maintain euglycemia). STZ rats demonstrated significantly elevated blood glucose throughout the 48-h protocol, while control and STZ-INS rats were euglycemic. Body mass increased in control (13 ± 4 g), decreased by 19 ± 2 g in STZ and remained unchanged in STZ-INS rats (−0.3 ± 2 g). Cross-sectional areas of gastrocnemius muscle fibers were smaller in STZ vs. control (1,480 ± 149 vs. 1,870 ± 40 μm2, respectively; P < 0.05) and insulin treatment did not rescue this defect (STZ-INS: 1,476 ± 143 μm2). Western blot analysis revealed a detectable increase in ubiquitinated proteins in the STZ skeletal muscles compared with control and STZ-INS. To further define the effects of STZ on skeletal muscle, independent of hyperglycemia, myoblasts were exposed to varying doses of STZ (0.25–3.0mg/ml) in vitro. Both acute and chronic exposures of STZ significantly impaired proliferative capacity in a dose-dependent manner. Within STZ-treated myoblasts, increased reactive oxygen species was associated with significant G2/M phase cell-cycle arrest. Taken together, our findings show that the effects of STZ are not β-cell specific and reveal that STZ should not be used for studies examining diabetic myopathy.

scholarly journals Exogenous Expression of an Alternative Splicing Variant of Myostatin Prompts Leg Muscle Fiber Hyperplasia in Japanese Quail

2019 ◽  
Vol 20 (18) ◽  
pp. 4617 ◽  
Author(s):  
Paula Renee Chen ◽  
Yeunsu Suh ◽  
Sangsu Shin ◽  
Rachel Marie Woodfint ◽  
Seongsoo Hwang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Alternative Splicing ◽  
Japanese Quail ◽  
Muscle Development ◽  
Pectoralis Major Muscle ◽  
Muscle Growth ◽  
Cross Sectional ◽  
Proliferation And Differentiation

Myostatin (MSTN) negatively regulates muscle growth and development through inhibiting myoblast proliferation and differentiation. Five alternative splicing isoforms of MSTN (MSTN-A to MSTN-E) have been discovered in domestic avian species. MSTN-A has high expression in skeletal muscle and encodes the full-length peptide with anti-myogenic activity. Another isoform, MSTN-B, is also highly expressed in skeletal muscle and encodes a truncated peptide that has pro-myogenic capabilities in vitro, which include promoting the proliferation and differentiation of quail muscle precursor cells. The objective of this study was to investigate overexpression of MSTN-B in vivo by using two independent lines of transgenic Japanese quail with expression directed in the skeletal muscle. Unexpectedly, the chicken skeletal muscle alpha actin 1 (cACTA1) promoter resulted in restricted exogenous MSTN-B protein expression to certain skeletal muscles, such as the gastrocnemius and tibialis anterior, but not the pectoralis major muscle. Gastrocnemius weight as a percentage of body weight in transgenic quail was increased compared to non-transgenic quail at posthatch day 21 (D21) and posthatch D42. An increase in the size of the gastrocnemius in transgenic quail was attributed to an increase in fiber number but not fiber cross-sectional area (CSA). During embryonic development, paired box 7 (PAX7) expression was prolonged in the transgenic embryos, but other myogenic regulatory factors (MRFs) were unchanged after MSTN-B overexpression. Taken together, these data provide novel insights into the regulation of skeletal muscle development by alternative splicing mechanisms in avians.

scholarly journals Androgens induce growth of the limb skeletal muscles in a rapamycin-insensitive manner

2018 ◽  
Vol 315 (4) ◽  
pp. R721-R729 ◽  
Author(s):  
Michael L. Rossetti ◽  
David H. Fukuda ◽  
Bradley S. Gordon
Keyword(s):  
Skeletal Muscle ◽  
Fiber Type ◽  
Muscle Growth ◽  
Total Protein Content ◽  
Cross Sectional ◽  
S6 Kinase ◽  
Mechanistic Target Of Rapamycin ◽  
Ribosomal S6 Kinase

Signaling through the mechanistic target of rapamycin complex 1 (mTORC1) has been well defined as an androgen-sensitive transducer mediating skeletal muscle growth in vitro; however, this has yet to be tested in vivo. As such, male mice were subjected to either sham or castration surgery and allowed to recover for 7 wk to induce atrophy of skeletal muscle. Then, castrated mice were implanted with either a control pellet or a pellet that administered rapamycin (~2.5 mg·kg−1·day−1). Seven days postimplant, a subset of castrated mice with control pellets and all castrated mice with rapamycin pellets were given once weekly injections of nandrolone decanoate (ND) to induce muscle growth over a six-week period. Effective blockade of mTORC1 by rapamycin was noted in the skeletal muscle by the inability of insulin to induce phosphorylation of ribosomal S6 kinase 1 70 kDa (Thr389) and uncoordinated-like kinase 1 (Ser757). While castration reduced tibialis anterior (TA) mass, muscle fiber cross-sectional area, and total protein content, ND administration restored these measures to sham levels in a rapamycin-insensitive manner. Similar findings were also observed in the plantaris and soleus, suggesting this rapamycin-insensitive effect was not specific to the TA or fiber type. Androgen-mediated growth was not due to changes in translational capacity. Despite these findings in the limb skeletal muscle, rapamycin completely prevented the ND-mediated growth of the heart. In all, these data indicate that mTORC1 has a limited role in the androgen-mediated growth of the limb skeletal muscle; however, mTORC1 was necessary for androgen-mediated growth of heart muscle.

scholarly journals FGF-2–dependent signaling activated in aged human skeletal muscle promotes intramuscular adipogenesis

2021 ◽  
Vol 118 (37) ◽  
pp. e2021013118 ◽  
Author(s):  
Sebastian Mathes ◽  
Alexandra Fahrner ◽  
Umesh Ghoshdastider ◽  
Hannes A. Rüdiger ◽  
Michael Leunig ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Transcriptional Activation ◽  
Human Skeletal Muscle ◽  
Muscle Growth ◽  
Muscle Cells ◽  
Adeno Associated Virus ◽  
Adult Skeletal Muscle

Aged skeletal muscle is markedly affected by fatty muscle infiltration, and strategies to reduce the occurrence of intramuscular adipocytes are urgently needed. Here, we show that fibroblast growth factor-2 (FGF-2) not only stimulates muscle growth but also promotes intramuscular adipogenesis. Using multiple screening assays upstream and downstream of microRNA (miR)-29a signaling, we located the secreted protein and adipogenic inhibitor SPARC to an FGF-2 signaling pathway that is conserved between skeletal muscle cells from mice and humans and that is activated in skeletal muscle of aged mice and humans. FGF-2 induces the miR-29a/SPARC axis through transcriptional activation of FRA-1, which binds and activates an evolutionary conserved AP-1 site element proximal in the miR-29a promoter. Genetic deletions in muscle cells and adeno-associated virus–mediated overexpression of FGF-2 or SPARC in mouse skeletal muscle revealed that this axis regulates differentiation of fibro/adipogenic progenitors in vitro and intramuscular adipose tissue (IMAT) formation in vivo. Skeletal muscle from human donors aged >75 y versus <55 y showed activation of FGF-2–dependent signaling and increased IMAT. Thus, our data highlights a disparate role of FGF-2 in adult skeletal muscle and reveals a pathway to combat fat accumulation in aged human skeletal muscle.

Wogonoside Inhibits Prostate Cancer Cell Growth and Metastasis via Regulating Wnt/β-Catenin Pathway and Epithelial-Mesenchymal Transition

Pharmacology ◽  
2019 ◽  
Vol 104 (5-6) ◽  
pp. 312-319 ◽  
Author(s):  
Can Wei ◽  
Junfeng Jing ◽  
Yanbin Zhang ◽  
Ling Fang
Keyword(s):  
Prostate Cancer ◽  
Cell Viability ◽  
Epithelial Mesenchymal Transition ◽  
Phase Cell ◽  
Dependent Manner ◽  
Mesenchymal Transition ◽  
Pc3 Cells ◽  
Gsk 3Β

Background: Wogonoside, an effective component of Scutellaria baicalensis extract, has recently become a hot topic for its newly discovered anticancer efficacy, but the underlying pharmacological mechanism is still unclear. In this study, we tested the inhibitory effects of wogonoside in human prostate cancer PC3 cells in vitro and vivo. Methods: The effects of wogonoside on cell viability, cycle progression, invasion, migration, and apoptosis were assessed in vitro. The levels of proteins in related signaling pathways were detected by western blotting assay. Finally, nude mouse tumorigenicity assay was conducted to detect the anticancer effect of wogonoside in vivo. Results: Wogonoside inhibited cell viability, invasive and migratory ability in a time- and dose-dependent manner. Flow cytometry indicated that wogonoside could induce cell apoptosis and S phase cell-cycle arrest. Mechanically, wogonoside suppressed the Wnt/β-catenin signaling pathway, and the level of p-glycogen synthase kinase-3β (GSK-3β; Ser9) was inhibited by wogonoside. The epithelial-mesenchymal transition (EMT) process was also reversed in PC3 cell line after wogonoside treatment. In vivo experiments showed that wogonoside inhibited tumor growth in xenograft mouse models. Conclusion: These findings revealed that wogonoside could suppress Wnt/β-catenin pathway and reversing the EMT process in PC3 cells. GSK-3β acts as a tumor suppressor in prostate cancer. Wogonoside may serve as an effective agent for treating prostate cancer.

Expression of steroidogenic enzymes and synthesis of sex steroid hormones from DHEA in skeletal muscle of rats

2007 ◽  
Vol 292 (2) ◽  
pp. E577-E584 ◽  
Author(s):  
Katsuji Aizawa ◽  
Motoyuki Iemitsu ◽  
Seiji Maeda ◽  
Subrina Jesmin ◽  
Takeshi Otsuki ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Steroid Hormones ◽  
Skeletal Muscles ◽  
Muscle Cells ◽  
Sex Steroid Hormones ◽  
Sex Steroid ◽  
Dependent Manner ◽  
Steroidogenic Enzymes

The functional importance of sex steroid hormones (testosterone and estrogens), derived from extragonadal tissues, has recently gained significant appreciation. Circulating dehydroepiandrosterone (DHEA) is peripherally taken up and converted to testosterone by 3β-hydroxysteroid dehydrogenase (HSD) and 17β-HSD, and testosterone in turn is irreversibly converted to estrogens by aromatase cytochrome P-450 (P450arom). Although sex steroid hormones have been implicated in skeletal muscle regulation and adaptation, it is unclear whether skeletal muscles have a local steroidogenic enzymatic machinery capable of metabolizing circulating DHEA. Thus, here, we investigate whether the three key steroidogenic enzymes (3β-HSD, 17β-HSD, and P450arom) are present in the skeletal muscle and are capable of generating sex steroid hormones. Consistent with our hypothesis, the present study demonstrates mRNA and protein expression of these enzymes in the skeletal muscle cells of rats both in vivo and in culture (in vitro). Importantly, we also show an intracellular formation of testosterone and estradiol from DHEA or testosterone in cultured muscle cells in a dose-dependent manner. These findings are novel and important in that they provide the first evidence showing that skeletal muscles are capable of locally synthesizing sex steroid hormones from circulating DHEA or testosterone.

scholarly journals PL - 033 A translational model of muscle protein synthetic bioactivity in vitro, ex vivo and in vivo

2018 ◽  
Vol 1 (1) ◽  
Author(s):  
Brian Carson ◽  
Robert Davies ◽  
Joseph Bass ◽  
Catherine Norton ◽  
Bijal Patel ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stable Isotope ◽  
Resistance Exercise ◽  
Ex Vivo ◽  
Muscle Protein ◽  
Muscle Growth ◽  
Translational Model ◽  
Nutrient Feeding

Objective The aim of this research was the development and validation of a translational model for the evaluation of exercise and nutrient stimulated muscle protein synthesis (MPS). To achieve this overall aim, three primary objectives had to be realised: (i) Development of an in vitro skeletal muscle cell bioassay to measure muscle growth and MPS; (ii) Development of an ex vivo model to evaluate the humoral effect on MPS in response to nutrient feeding and exercise; (iii) Use of a stable isotope technique to evaluate MPS in response to nutrient feeding and exercise in vivo. Methods To develop a novel in vitro skeletal muscle cell bioassay to measure muscle growth and MPS, C2C12 myoblasts were proliferated and subsequently differentiated to myotubes over 8 days in DMEM (2% HS). Changes in cell behavior and adhesion properties were monitored by measuring impedance via interdigitated microelectrodes using the xCELLigence system. MPS was measured by puromycin incorporation using the SUnSET technique, intracellular signalling measured by western blot, and myotube thickness by microscopy. To demonstrate the capability to monitor nutrient regulation of muscle growth, media was conditioned with a known potent regulator of MPS (leucine) in a dose response experiment (0.20 - 2.0 mM). To establish the ability of the bioassay to measure the humoral effect of MPS in response to feeding and exercise, media was conditioned by ex vivo human serum from fasted, rested, fed (protein and isonitrogenous non-essential amino acid (NEAA) control)  and post-exercise conditions. To evaluate MPS in response to nutrient feeding and exercise in vivo, acute MPS (5 h) was assessed by measuring stable isotope deuterium oxide (D2O) incorporation into m. vastus lateralis skeletal muscle following consumption of either a Whey Protein (WP) or an isonitrogenous NEAA control combined with resistance exercise in resistance trained males. Results In vitro experiments observed a dose-response effect with a 32 % increase in cell index and a 27 % increase in cell thickness after 2 h in the presence of 2.0 mM leucine when compared with control myotubes. Ex vivo serum following ingestion of NEAA had no effect on protein signalling or MPS whereas WP fed serum significantly increased mTOR, P70S6K and 4E-BP1 phosphorylation (p<0.01, p<0.05) compared to fasted serum. Furthermore, the effect of WP fed serum on protein signalling and MPS was significantly increased (p<0.01, p<0.05) compared to NEAA fed serum.  Ex vivo human serum following resistance exercise was also increased MPS (29 %) and phosphorylation of mTOR (6 %), p70S6K (12 %) and 4EBP1 (7 %), compared with resting serum. These ex vivo/in vitro findings translated to the in vivo model as myofibrillar fractional synthetic rates (myoFSR) (Basal 0.068±0.002%h-1 vs. WP 0.084±0.006 %h-1, p=0.033) and absolute synthetic rates (ASR) (Basal 10.34±1.01 vs. WP 13.18±0.71 g.day-1, p=0.026) were increased from basal levels only when resistance exercise was combined with WP ingestion and not the NEAA control (NEAA MPS 0.072±0.004%h-1, NEAA ASR 10.23±0.80 g.day-1).  Thus, ingestion of WP in combination with resistance training augments acute MPS responses in resistance trained young men. Conclusions We have developed a translational model of muscle protein synthetic bioactivity using in vitro, ex vivo and in vivo methodologies. We have shown that we can impact MPS in vitro using ex vivo human serum to condition media, that MPS in vitro is differentially regulated by ex vivo serum containing bioactive WP compared to a non-bioactive NEAA control, and that this tranlates for resistance exercise combined with WP in humans when MyoFSR is measured using stable isotope technology.  These experiments demonstrate that ex vivo/in vitro experiments translate to the in vivo model and these methods can be used to inform both exercise and nutrient human interventions. 

scholarly journals Preptin, another peptide product of the pancreatic β-cell, is osteogenic in vitro and in vivo

2007 ◽  
Vol 292 (1) ◽  
pp. E117-E122 ◽  
Author(s):  
J. Cornish ◽  
K. E. Callon ◽  
U. Bava ◽  
M. Watson ◽  
X. Xu ◽  
...  
Keyword(s):  
Bone Mass ◽  
Map Kinases ◽  
Cell Number ◽  
Dependent Manner ◽  
Pancreatic Β Cell ◽  
Hepatitis C Infection ◽  
Β Cell ◽  
High Bone Mass

Several hormones that regulate nutritional status also impact on bone metabolism. Preptin is a recently isolated 34-amino acid peptide hormone that is cosecreted with insulin and amylin from the pancreatic β-cells. Preptin corresponds to Asp69-Leu102 of pro-IGF-II. Increased circulating levels of a pro-IGF-II peptide complexed with IGF-binding protein-2 have been implicated in the high bone mass phenotype observed in patients with chronic hepatitis C infection. We have assessed preptin's activities on bone. Preptin dose-dependently stimulated the proliferation (cell number and DNA synthesis) of primary fetal rat osteoblasts and osteoblast-like cell lines at periphysiological concentrations (>10−11 M). In addition, thymidine incorporation was stimulated in murine neonatal calvarial organ culture, likely reflecting the proliferation of cells from the osteoblast lineage. Preptin did not affect bone resorption in this model. Preptin induced phosphorylation of p42/p44 MAP kinases in osteoblastic cells in a dose-dependent manner (10−8-10−10 M), and its proliferative effects on primary osteoblasts were blocked by MAP kinase kinase inhibitors. Preptin also reduced osteoblast apoptosis induced by serum deprivation, reducing the number of apoptotic cells by >20%. In vivo administration of preptin increased bone area and mineralizing surface in adult mice. These data demonstrate that preptin, which is cosecreted from the pancreatic β-cell with amylin and insulin, is anabolic to bone and may contribute to the preservation of bone mass observed in hyperinsulinemic states such as obesity.

The effect of caffeine on skeletal muscle anabolic signaling and hypertrophy

2017 ◽  
Vol 42 (6) ◽  
pp. 621-629 ◽  
Author(s):  
Timothy M. Moore ◽  
Xavier M. Mortensen ◽  
Conrad K. Ashby ◽  
Alexander M. Harris ◽  
Karson J. Kump ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Muscle Growth ◽  
Mtor Signaling ◽  
Signaling Proteins ◽  
Signaling Protein ◽  
Before And After ◽  
Amp Activated Protein Kinase

Caffeine is a widely consumed stimulant with the potential to enhance physical performance through multiple mechanisms. However, recent in vitro findings have suggested that caffeine may block skeletal muscle anabolic signaling through AMP-activated protein kinase (AMPK)-mediated inhibition of mechanistic target of rapamycin (mTOR) signaling pathway. This could negatively affect protein synthesis and the capacity for muscle growth. The primary purpose of this study was to assess the effect of caffeine on in vivo AMPK and mTOR pathway signaling, protein synthesis, and muscle growth. In cultured C2C12 muscle cells, physiological levels of caffeine failed to impact mTOR activation or myoblast proliferation or differentiation. We found that caffeine administration to mice did not significantly enhance the phosphorylation of AMPK or inhibit signaling proteins downstream of mTOR (p70S6k, S6, or 4EBP1) or protein synthesis after a bout of electrically stimulated contractions. Skeletal muscle-specific knockout of LKB1, the primary AMPK activator in skeletal muscle, on the other hand, eliminated AMPK activation by contractions and enhanced S6k, S6, and 4EBP1 activation before and after contractions. In rats, the addition of caffeine did not affect plantaris hypertrophy induced by the tenotomy of the gastrocnemius and soleus muscles. In conclusion, caffeine administration does not impair skeletal muscle load-induced mTOR signaling, protein synthesis, or muscle hypertrophy.

scholarly journals Identification of PCIF1, a POZ Domain Protein That Inhibits PDX-1 (MODY4) Transcriptional Activity

2004 ◽  
Vol 24 (10) ◽  
pp. 4372-4383 ◽  
Author(s):  
Aihua Liu ◽  
Biva M. Desai ◽  
Doris A. Stoffers
Keyword(s):  
Late Onset ◽  
Dependent Manner ◽  
Gene Promoters ◽  
Transactivation Domain ◽  
Β Cells ◽  
Β Cell ◽  
C Terminus ◽  
Domain Protein

ABSTRACT Hox factors are evolutionarily conserved homeodomain-containing transcription factors that activate and repress gene expression in a precise temporally and spatially regulated manner during development and differentiation. Pancreatic-duodenal homeobox 1 (PDX-1) is a Hox-type protein that is a critical requirement for normal pancreas development and for proper differentiation of the endocrine pancreas. In humans, PDX-1 gene mutation causes pancreatic agenesis and early- and late-onset type 2 diabetes. PDX-1 consists of an N-terminal transactivation domain, a homeodomain responsible for DNA binding and nuclear localization, and a conserved C terminus that is mutated in human diabetes but whose function is poorly understood. We have identified a novel POZ domain protein, PDX-1 C terminus-interacting factor 1 (PCIF1)/SPOP, that interacts with PDX-1 both in vitro and in vivo. PCIF1 is localized to the nucleus in a speckled pattern, and coexpression of PDX-1 alters the subnuclear distribution of PCIF1. Functionally, PCIF1 inhibits PDX-1 transactivation of established target gene promoters in a specific and dose-dependent manner that requires critical amino acids in the PDX-1 C terminus. PCIF1 is expressed in adult pancreatic insulin-producing β cells, and overexpression of PCIF1 inhibits the rat insulin 1 and rat insulin 2 promoters in the MIN6 insulinoma β cell line. The coexpression of PCIF1 with PDX-1 in β cells and the ability of PCIF1 to repress PDX-1 transactivation suggest that modulation of PDX-1 function by PCIF1 may regulate normal β cell differentiation.

scholarly journals Alcohol impairs skeletal muscle protein synthesis and mTOR signaling in a time-dependent manner following electrically stimulated muscle contraction

2014 ◽  
Vol 117 (10) ◽  
pp. 1170-1179 ◽  
Author(s):  
Jennifer L. Steiner ◽  
Charles H. Lang
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Mammalian Target Of Rapamycin ◽  
Mtor Signaling ◽  
Saline Control ◽  
Dependent Manner ◽  
Skeletal Muscle Protein

Alcohol (EtOH) decreases protein synthesis and mammalian target of rapamycin (mTOR)-mediated signaling and blunts the anabolic response to growth factors in skeletal muscle. The purpose of the current investigation was to determine whether acute EtOH intoxication antagonizes the contraction-induced increase in protein synthesis and mTOR signaling in skeletal muscle. Fasted male mice were injected intraperitoneally with 3 g/kg EtOH or saline (control), and the right hindlimb was electrically stimulated (10 sets of 6 contractions). The gastrocnemius muscle complex was collected 30 min, 4 h, or 12 h after stimulation. EtOH decreased in vivo basal protein synthesis (PS) in the nonstimulated muscle compared with time-matched Controls at 30 min, 4 h, and 12 h. In Control, but not EtOH, PS was decreased 15% after 30 min. In contrast, PS was increased in Control 4 h poststimulation but remained unchanged in EtOH. Last, stimulation increased PS 10% in Control and EtOH at 12 h, even though the absolute rate remained reduced by EtOH. The stimulation-induced increase in the phosphorylation of S6K1 Thr421/Ser424 (20–52%), S6K1 Thr389 (45–57%), and its substrate rpS6 Ser240/244 (37–72%) was blunted by EtOH at 30 min, 4 h, and 12 h. Phosphorylation of 4E-BP1 Ser65 was also attenuated by EtOH (61%) at 4 h. Conversely, phosphorylation of extracellular signal-regulated kinase Thr202/Tyr204 was increased by stimulation in Control and EtOH mice at 30 min but only in Control at 4 h. Our data indicate that acute EtOH intoxication suppresses muscle protein synthesis for at least 12 h and greatly impairs contraction-induced changes in synthesis and mTOR signaling.

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