Dystrophin Dp71ab is monoclonally expressed in human satellite cells and enhances proliferation of myoblast cells

Manal Farea,Hisahide Nishio,Kazuhiro Maeta,Abdul Qawee Mahyoob Rani,Masafumi Matsuo

doi:10.1038/s41598-020-74157-y

Manal Farea, Hisahide Nishio + Show 3 more

Open Access

https://doi.org/10.1038/s41598-020-74157-y

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Journal: Scientific Reports	Publication Date: Oct 13, 2020
Citations: 17	License type: open-access

Affiliation: Kobe Gakuin University

Abstract

Dystrophin Dp71 is the smallest isoform of the DMD gene, mutations in which cause Duchenne muscular dystrophy (DMD). Dp71 has also been shown to have roles in various cellular processes. Stem cell-based therapy may be effective in treating DMD, but the inability to generate a sufficient number of stem cells remains a significant obstacle. Although Dp71 is comprised of many variants, Dp71 in satellite cells has not yet been studied. Here, the full-length Dp71 consisting of 18 exons from exons G1 to 79 was amplified by reverse transcription-PCR from total RNA of human satellite cells. The amplified product showed deletion of both exons 71 and 78 in all sequenced clones, indicating monoclonal expression of Dp71ab. Western blotting of the satellite cell lysate showed a band corresponding to over-expressed Dp71ab. Transfection of a plasmid expressing Dp71ab into human myoblasts significantly enhanced cell proliferation when compared to the cells transfected with the mock plasmid. However, transfection of the Dp71 expression plasmid encoding all 18 exons did not enhance myoblast proliferation. These findings indicated that Dp71ab, but not Dp71, is a molecular enhancer of myoblast proliferation and that transfection with Dp71ab may generate a high yield of stem cells for DMD treatment.

Highlights

Dystrophin Dp71 is the smallest isoform of the DMD gene, mutations in which cause Duchenne muscular dystrophy (DMD)
The present study assessed the expression of Dp71 in human satellite cells by reverse transcription (RT)-PCR amplification and Western blotting
Over-expression of Dp71ab enhanced myoblast proliferation, suggesting that expression of Dp71ab may generate a high yield of stem cells for DMD treatment

Summary

Introduction

Dystrophin Dp71 is the smallest isoform of the DMD gene, mutations in which cause Duchenne muscular dystrophy (DMD). Transfection of the Dp71 expression plasmid encoding all 18 exons did not enhance myoblast proliferation These findings indicated that Dp71ab, but not Dp71, is a molecular enhancer of myoblast proliferation and that transfection with Dp71ab may generate a high yield of stem cells for DMD treatment. One alternative downstream promoter located in intron 62 produces a transcript consisting of a unique exon, G1 and DMD exons 63–79 shared with other dystrophin isoforms[4,5] This transcript encodes a nearly 71-kDa protein, dystrophin Dp71, an isoform that is expressed u biquitously[6]. Duchenne muscular dystrophy (DMD) is a fatal progressive muscle wasting disease, caused by mutations in the DMD gene[1,22] Many strategies, such as stem cell-based therapy have been proposed to treat DMD23. Over-expression of Dp71ab enhanced myoblast proliferation, suggesting that expression of Dp71ab may generate a high yield of stem cells for DMD treatment

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