Skip to main content

Cell Density and Joint microRNA-133a and microRNA-696 Inhibition Enhance Differentiation and Contractile Function of Engineered Human Skeletal Muscle Tissues.

Publication ,  Journal Article
Cheng, CS; Ran, L; Bursac, N; Kraus, WE; Truskey, GA
Published in: Tissue Eng Part A
April 2016

To utilize three-dimensional (3D) engineered human skeletal muscle tissue for translational studies and in vitro studies of drug toxicity, there is a need to promote differentiation and functional behavior. In this study, we identified conditions to promote contraction of engineered human skeletal muscle bundles and examined the effects of transient inhibition of microRNAs (miRs) on myogenic differentiation and function of two-dimensional (2D) and 3D cultures of human myotubes. In 2D cultures, simultaneously inhibiting both miR-133a, which promotes myoblast proliferation, and miR-696, which represses oxidative metabolism, resulted in an increase in sarcomeric α-actinin protein and the metabolic coactivator PGC-1α protein compared to transfection with a scrambled miR sequence (negative control). Although PGC-1α was elevated following joint inhibition of miRs 133a and 696, there was no difference in myosin heavy chain (MHC) protein isoforms. 3D engineered human skeletal muscle myobundles seeded with 5 × 10(6) human skeletal myoblasts (HSkM)/mL and cultured for 2 weeks after onset of differentiation consistently did not contract when stimulated electrically, whereas those seeded with myoblasts at 10 × 10(6) HSkM/mL or higher did contract. When HSkM were transfected with both anti-miRs and seeded into fibrin hydrogels and cultured for 2 weeks under static conditions, twitch and tetanic specific forces after electrical stimulation were greater than for myobundles prepared with HSkM transfected with scrambled sequences. Immunofluorescence and Western blots of 3D myobundles indicate that anti-miR-133a or anti-miR-696 treatment led to modest increases in slow MHC, but no consistent increase in fast MHC. Similar to results in 2D, only myobundles prepared with myoblasts treated with anti-miR-133a and anti-miR-696 produced an increase in PGC-1α mRNA. PGC-1α targets were differentially affected by the treatment. HIF-2α mRNA showed an expression pattern similar to that of PGC-1α mRNA, but COXII mRNA levels were not affected by the anti-miRs. Overall, joint inhibition of miR-133a and miR-696 accelerated differentiation, elevated the metabolic coactivator PGC-1α, and increased the contractile force in 3D engineered human skeletal muscle bundles.

Duke Scholars

Altmetric Attention Stats
Dimensions Citation Stats

Published In

Tissue Eng Part A

DOI

EISSN

1937-335X

Publication Date

April 2016

Volume

22

Issue

7-8

Start / End Page

573 / 583

Location

United States

Related Subject Headings

  • Transfection
  • Tissue Engineering
  • Sarcomeres
  • RNA, Messenger
  • Protein Isoforms
  • Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
  • Myosin Heavy Chains
  • Myoblasts
  • Muscle, Skeletal
  • Muscle Development
 

Citation

APA
Chicago
ICMJE
MLA
NLM
Cheng, C. S., Ran, L., Bursac, N., Kraus, W. E., & Truskey, G. A. (2016). Cell Density and Joint microRNA-133a and microRNA-696 Inhibition Enhance Differentiation and Contractile Function of Engineered Human Skeletal Muscle Tissues. Tissue Eng Part A, 22(7–8), 573–583. https://doi.org/10.1089/ten.TEA.2015.0359
Cheng, Cindy S., Lydia Ran, Nenad Bursac, William E. Kraus, and George A. Truskey. “Cell Density and Joint microRNA-133a and microRNA-696 Inhibition Enhance Differentiation and Contractile Function of Engineered Human Skeletal Muscle Tissues.Tissue Eng Part A 22, no. 7–8 (April 2016): 573–83. https://doi.org/10.1089/ten.TEA.2015.0359.
Cheng, Cindy S., et al. “Cell Density and Joint microRNA-133a and microRNA-696 Inhibition Enhance Differentiation and Contractile Function of Engineered Human Skeletal Muscle Tissues.Tissue Eng Part A, vol. 22, no. 7–8, Apr. 2016, pp. 573–83. Pubmed, doi:10.1089/ten.TEA.2015.0359.

Published In

Tissue Eng Part A

DOI

EISSN

1937-335X

Publication Date

April 2016

Volume

22

Issue

7-8

Start / End Page

573 / 583

Location

United States

Related Subject Headings

  • Transfection
  • Tissue Engineering
  • Sarcomeres
  • RNA, Messenger
  • Protein Isoforms
  • Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
  • Myosin Heavy Chains
  • Myoblasts
  • Muscle, Skeletal
  • Muscle Development