Abstract 15867: Myogenic Target of Mir-290 Cluster Promotes Reprogramming of Skeletal Myoblast to Produce Induced Pluripotent Stem Cells
Background: Skeletal myoblasts (SkMs) are an ideal candidate for reprogramming due to inherent myogenic properties. MicroRNAs (miRNAs) play an important role in cell reprogramming by producing induced pluripotent stem (iPS) cells. Pax7 is a powerful myogenic inducer and plays a critical role in muscle development and regeneration. This study investigates the potential mechanisms of the miR-290s cluster for promoting reprogramming efficiency of SkMs.
Methods and Results: The SkMs cells were isolated from the hind limb skeletal muscle of 2 to 4-week-old mice. RT-PCR and immunocytochemical analysis showed that SkMs endogenously expressed not only myogenic genes but also some of pluripotency genes Nanog, Oct4, Sox2, Klf4 and cMyc. SkMs were then infected to produce iPS cells, and embryonic stem (ES) cell-like colonies appeared at 15 days after transfection. SkM-derived iPS (SiPS) cells were profiled by microarray to determine miRNAs responsible for reprogramming of SkMs. Expression of each member of the miR-290 cluster was found including miR-290a, miR-291a, miR-292, miR-293, miR-294, and miR-295. These were significantly higher in SiPS cells as compared to SkMs, a result validated by RT-PCR. In addition, expression of Pax7 was silenced in SiPS cells as compared to SkMs. Computational analysis was then performed to determine the target genes of miR-290 cluster to gain insight into the molecular mechanisms. The seed sequence of miR-291a-5p was found to bind to the 3’ UTR of Pax7 gene and reprogramming efficiency of SiPS cells was successfully increased using transfection with lentiviral vector containing miR-291a-5p.
Conclusions: Reprogramming of SkMs by introduction of miR-290 clusters is an efficient way to generate iPS cells, and Pax7 is one of the potential targets of miR-290s cluster.
Author Disclosures: L. Wang: None. H. Kondo: None. R. Ma: None. L. Guo: None. H. Kim: None. Y. Wang: None.
- © 2014 by American Heart Association, Inc.