Abstract
ABSTRACTPurpose: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 96® 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.
Highlights
Small double stranded RNAs are known to be able to induce sequence-specific genes expression by targeting gene promoter regions, a phenomenon known as RNA activation (RNAa) and named as small activating RNA [1]
It is well known that RNA interference (RNAi) is being actively pursued as therapeutics for many ibju | double stranded RNAs (dsRNAs) induces myogenic differentiation of rat adipose-derived stem cells (ADSCs)
Our results show that a candidate dsMyoD-373 potently upregulated MyoD expression and induced rat ADSCs differentiated into myoblasts
Summary
Small double stranded RNAs (dsRNAs) are known to be able to induce sequence-specific genes expression by targeting gene promoter regions, a phenomenon known as RNA activation (RNAa) and named as small activating RNA (saRNAs) [1]. Li and colleagues have demonstrated that saRNAs could activate genes E-cadherin, p21WAF1/CIP1 and VEGF expression in human cell lines [1]. Our previous study confirmed that saRNA could elicit antitumor activity by triggering the expression of p21WAF1/CIP1 in human bladder cancer cell lines [2]. Some diseases, especially related to tissue degeneration or damage, are often caused by decreased expression of certain gene products involved in crucial physiological function [6]. These defects are difficult to be cured by RNAi and may be restored by RNAa
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