Adenoviral mediated MyoD gene transfer into fibroblasts: myogenic disease diagnosis.
MyoD, a master regulatory gene for myogenesis, converts mesoderm derived cells to the skeletal muscle phenotype MyoD gene transfer into skin fibroblasts has been attempted in an effort to diagnose genetic muscle diseases. Although the gene transduction efficiency of adenoviral gene delivery systems is higher than that of various other systems, the rate of myo-conversion is insignificant. Since high adenovirus doses are cytotoxic and exogenous MyoD expression is insufficient for skin fibroblasts to re-differentiate into muscle cells, we constructed the novel adeno-MyoD vector, Ad.CAGMyoD using the recombinant CAG promoter. Even at a lower multiplicities of infection most skin fibroblasts infected with Ad.CAGMyoD could convert into myotubes without vector-induced cytotoxicity. The converted cells expressed muscle-specific desmin and full-length dystrophin, both of which were detected by Western blotting. Genetic and immunohistochemical analyses using skin fibroblasts and our vector system are reliable and useful for the clinical diagnosis of genetic muscle diseases.
Laboratory of Clinical Pharmacology and Therapeutics, Department of Pharmaceutical Sciences, Division of Clinical Pharmacy, Faculty of Pharmaceutical Sciences, Sojo University, 4-22-1 Ikeda, Kumamoto-city, Kumamoto 860-0082, Japan. isa-f@ph.sojo-u.ac.jp
Isao
I
Fujii
Makoto
M
Matsukura
Makoto
M
Ikezawa
Satoru
S
Suzuki
Takashi
T
Shimada
Teruhisa
T
Miike
D000256:Adenoviridae / Q000502:physiology*; D015153:Blotting, Western / Q000379:methods; D002478:Cells, Cultured; D002675:Child, Preschool; D003893:Desmin / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics* / Q000473:pathology*; D017570:MyoD Protein / Q000235:genetics* / Q000378:metabolism; D013997:Time Factors
D003893:Desmin; D017570:MyoD Protein; D049452:Green Fluorescent Proteins