PMID- 16810717 OWN - NLM STAT- MEDLINE DCOM- 20061108 LR - 20220310 IS - 1099-498X (Print) IS - 1099-498X (Linking) VI - 8 IP - 9 DP - 2006 Sep TI - Myostatin short interfering hairpin RNA gene transfer increases skeletal muscle mass. PG - 1171-81 AB - BACKGROUND: Myostatin negatively regulates skeletal muscle growth. Myostatin knockout mice exhibit muscle hypertrophy and decreased interstitial fibrosis. We investigated whether a plasmid expressing a short hairpin interfering RNA (shRNA) against myostatin and transduced using electroporation would increase local skeletal muscle mass. METHODS: Short interfering RNAs (siRNAs) targeting myostatin were co-transfected with a myostatin-expressing plasmid into HEK293 cells and identified for myostatin silencing by Western blot. Corresponding shRNAs were cloned into plasmid shRNA expression vectors. Myostatin or a randomer negative control shRNA plasmid was injected and electroporated into the tibialis anterior or its contralateral muscle, respectively, of nine rats that were sacrificed after 2 weeks. Six other rats received a beta-galactosidase reporter plasmid and were sacrificed at 1, 2, and 4 weeks. Uptake of plasmid was examined by beta-galactosidase expression, whereas myostatin expression was determined by real-time polymerase chain reaction (PCR) and Western blotting. Muscle fiber size was determined by histochemistry. Satellite cell proliferation was determined by PAX7 immunohistochemistry. Myosin heavy chain type II (MHCII) expression was determined by Western blot. RESULTS: beta-Galactosidase reporter plasmid was expressed at 1 and 2 weeks but diminished by 4 weeks in tibialis anterior skeletal muscle. Myostatin shRNA reduced myostatin mRNA and protein expression by 27 and 48%, respectively. Tibialis anterior weight, fiber size, and MHCII increased by 10, 34, and 38%, respectively. Satellite cell number was increased by over 2-fold. CONCLUSIONS: This is the first demonstration that myostatin shRNA gene transfer is a potential strategy to increase muscle mass. CI - Copyright (c) 2006 John Wiley & Sons, Ltd. FAU - Magee, Thomas R AU - Magee TR AD - Department of Urology, David Geffen School of Medicine, UCLA, Los Angeles, CA 90024, USA. tmagee@labiomed.org FAU - Artaza, Jorge N AU - Artaza JN FAU - Ferrini, Monica G AU - Ferrini MG FAU - Vernet, Dolores AU - Vernet D FAU - Zuniga, Freddi I AU - Zuniga FI FAU - Cantini, Liliana AU - Cantini L FAU - Reisz-Porszasz, Suzanne AU - Reisz-Porszasz S FAU - Rajfer, Jacob AU - Rajfer J FAU - Gonzalez-Cadavid, Nestor F AU - Gonzalez-Cadavid NF LA - eng GR - 3S06GM068510-02S21/GM/NIGMS NIH HHS/United States GR - 5P20MD000545/MD/NIMHD NIH HHS/United States GR - G12RR-03026/RR/NCRR NIH HHS/United States GR - R01DK-52069/DK/NIDDK NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PL - England TA - J Gene Med JT - The journal of gene medicine JID - 9815764 RN - 0 (MSTN protein, human) RN - 0 (Mstn protein, mouse) RN - 0 (Mstn protein, rat) RN - 0 (Myostatin) RN - 0 (RNA, Small Interfering) RN - 0 (Transforming Growth Factor beta) SB - IM MH - Animals MH - Base Sequence MH - Cell Line MH - Electroporation MH - Gene Expression MH - Gene Silencing MH - *Gene Transfer Techniques MH - Genes, Reporter MH - Genetic Vectors MH - Humans MH - Lac Operon MH - Male MH - Mice MH - Muscle, Skeletal/*growth & development/*metabolism MH - Myostatin MH - Plasmids/genetics MH - RNA, Small Interfering/*genetics MH - Rats MH - Rats, Inbred F344 MH - Transfection MH - Transforming Growth Factor beta/*antagonists & inhibitors/*genetics EDAT- 2006/07/01 09:00 MHDA- 2006/11/10 09:00 CRDT- 2006/07/01 09:00 PHST- 2006/07/01 09:00 [pubmed] PHST- 2006/11/10 09:00 [medline] PHST- 2006/07/01 09:00 [entrez] AID - 10.1002/jgm.946 [doi] PST - ppublish SO - J Gene Med. 2006 Sep;8(9):1171-81. doi: 10.1002/jgm.946.