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Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro.

Publication ,  Journal Article
Cheng, CS; El-Abd, Y; Bui, K; Hyun, Y-E; Hughes, RH; Kraus, WE; Truskey, GA
Published in: Am J Physiol Cell Physiol
February 15, 2014

Conditions under which skeletal myoblasts are cultured in vitro are critical to growth and differentiation of these cells into mature skeletal myofibers. We examined several culture conditions that promoted human skeletal myoblast (HSkM) culture and examined the effect of microRNAs and mechanical stimulation on differentiation. Culture conditions for HSkM are different from those that enable rapid C2C12 myoblast differentiation. Culture on a growth factor-reduced Matrigel (GFR-MG)-coated surface in 2% equine serum-supplemented differentiation medium to promote HSkM differentiation under static conditions was compared with culture conditions used for C2C12 cell differentiation. Such conditions led to a >20-fold increase in myogenic miR-1, miR-133a, and miR-206 expression, a >2-fold increase in myogenic transcription factor Mef-2C expression, and an increase in sarcomeric α-actinin protein. Imposing ±10% cyclic stretch at 0.5 Hz for 1 h followed by 5 h of rest over 2 wk produced a >20% increase in miR-1, miR-133a, and miR-206 expression in 8% equine serum and a >35% decrease in 2% equine serum relative to static conditions. HSkM differentiation was accelerated in vitro by inhibition of proliferation-promoting miR-133a: immunofluorescence for sarcomeric α-actinin exhibited accelerated development of striations compared with the corresponding negative control, and Western blotting showed 30% more α-actinin at day 6 postdifferentiation. This study showed that 100 μg/ml GFR-MG coating and 2% equine serum-supplemented differentiation medium enhanced HSkM differentiation and myogenic miR expression and that addition of antisense miR-133a alone can accelerate primary human skeletal muscle differentiation in vitro.

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Published In

Am J Physiol Cell Physiol

DOI

EISSN

1522-1563

Publication Date

February 15, 2014

Volume

306

Issue

4

Start / End Page

C385 / C395

Location

United States

Related Subject Headings

  • Transfection
  • Time Factors
  • Proteoglycans
  • Primary Cell Culture
  • Physiology
  • Myoblasts, Skeletal
  • Muscle Development
  • MicroRNAs
  • Mice
  • Male
 

Citation

APA
Chicago
ICMJE
MLA
NLM
Cheng, C. S., El-Abd, Y., Bui, K., Hyun, Y.-E., Hughes, R. H., Kraus, W. E., & Truskey, G. A. (2014). Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro. Am J Physiol Cell Physiol, 306(4), C385–C395. https://doi.org/10.1152/ajpcell.00179.2013
Cheng, Cindy S., Yasser El-Abd, Khanh Bui, Young-Eun Hyun, Rebecca Harbuck Hughes, William E. Kraus, and George A. Truskey. “Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro.Am J Physiol Cell Physiol 306, no. 4 (February 15, 2014): C385–95. https://doi.org/10.1152/ajpcell.00179.2013.
Cheng CS, El-Abd Y, Bui K, Hyun Y-E, Hughes RH, Kraus WE, et al. Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro. Am J Physiol Cell Physiol. 2014 Feb 15;306(4):C385–95.
Cheng, Cindy S., et al. “Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro.Am J Physiol Cell Physiol, vol. 306, no. 4, Feb. 2014, pp. C385–95. Pubmed, doi:10.1152/ajpcell.00179.2013.
Cheng CS, El-Abd Y, Bui K, Hyun Y-E, Hughes RH, Kraus WE, Truskey GA. Conditions that promote primary human skeletal myoblast culture and muscle differentiation in vitro. Am J Physiol Cell Physiol. 2014 Feb 15;306(4):C385–C395.

Published In

Am J Physiol Cell Physiol

DOI

EISSN

1522-1563

Publication Date

February 15, 2014

Volume

306

Issue

4

Start / End Page

C385 / C395

Location

United States

Related Subject Headings

  • Transfection
  • Time Factors
  • Proteoglycans
  • Primary Cell Culture
  • Physiology
  • Myoblasts, Skeletal
  • Muscle Development
  • MicroRNAs
  • Mice
  • Male