Small activating RNA induces myogenic differentiation of rat adipose-derived stem cells by upregulating MyoD

Int Braz J Urol. 2015 Jul-Aug;41(4):764-72. doi: 10.1590/S1677-5538.IBJU.2014.0400.

Abstract

Purpose: RNA activation (RNAa) is a mechanism of gene activation triggered by promoter-targeted small double stranded RNAs (dsRNAs), also known as small activating RNAs (saRNAs). Myogenic regulatory factor MyoD is regarded as the master activator of myogenic differentiation cascade by binding to enhancer of muscle specific genes. Stress urinary incontinence (SUI) is a condition primarily resulted from urethral sphincter deficiency. It is thus expected that by promoting differentiation of adipose-derived stem cells (ADSCs) into myoblasts by activating MyoD gene through RNAa may offer benefits to SUI.

Materials and methods: Rats ADSCs were isolated, proliferated in vitro, and identified by flow cytometry. Purified ADSCs were then transfected with a MyoD saRNA or control transfected. Real-time polymerase chain reaction (RT-PCR) and western blotting were used to detect MyoD mRNA and protein expression, respectively. Immunocytochemical staining was applied to determine the expression of desmin protein in transfected cells. Cell viability was measured by using CellTiter 96R AQueous One Solution Cell Proliferation Assay kit.

Results: Transfection of a MyoD saRNA (dsMyoD) into ADSCs significantly induced the expression of MyoD at both the mRNA and protein levels, and inhibited cell proliferation. Desmin protein expression was detected in dsMyoD treated ADSCs 2 weeks later.

Conclusion: Our findings show that RNAa mediated overexpression of MyoD can promote transdifferentiation of ADSCs into myoblasts and may help treat stress urinary incontinence (SUI)-a condition primarily resulted from urethral sphincter deficiency.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adipose Tissue / cytology*
  • Animals
  • Blotting, Western
  • Cell Differentiation / genetics*
  • Cell Survival
  • Desmin / metabolism*
  • Flow Cytometry
  • Gene Expression
  • Immunohistochemistry
  • MyoD Protein / genetics*
  • MyoD Protein / metabolism
  • Myoblasts / cytology*
  • Myoblasts / metabolism
  • Primary Cell Culture
  • Promoter Regions, Genetic / physiology
  • RNA, Double-Stranded*
  • Rats
  • Real-Time Polymerase Chain Reaction
  • Stem Cells / cytology*
  • Stem Cells / metabolism
  • Transcriptional Activation / physiology
  • Transfection
  • Urethra / pathology
  • Urinary Incontinence, Stress / genetics
  • Urinary Incontinence, Stress / metabolism

Substances

  • Desmin
  • MyoD Protein
  • RNA, Double-Stranded

Grants and funding

This study was supported by the National Natural Science Foundation of PR China (grant number 30873018).