Bacterial beta-galactosidase and human dystrophin genes are expressed in mouse skeletal muscle fibers after ballistic transfection

FEBS Lett. 1997 Sep 8;414(2):319-22. doi: 10.1016/s0014-5793(97)01019-3.

Abstract

Ballistic transfection, based on cell and tissue bombardment by the tungsten and gold microparticles covered with the gene DNA, was used for the delivery of a bacterial beta-galactosidase and a full-length cDNA copy of the human dystrophin genes into mouse skeletal muscles. CMV-lacZ, SV40-lacZ, LTR-lacZneo and full-length cDNA dystrophin (pDMD-1, approximately 16 kb) in eukaryotic expression vector pJ OMEGA driven by mouse leukaemia virus promotor (pMLVDy) were used throughout the studies. Musculus glutaeus superficialis of C57BL/6J and quadriceps femoris of mdx male mice were opened surgically under anesthesia and bombarded by means of the gene-gun technique originally developed by us. Different mixtures of gold and tungsten particles at ratios of 4:1, 1:1, 1:4 were applied. X-gal assay revealed marked beta-gal activity, both in total muscles and whole muscle fibers on histological sections, up to three months after transfection. The most intensive staining was observed after SV40-lacZ delivery. No staining was detected with LTR-lacZneo DNA as well as in untreated muscles. The higher tungsten particle concentration in the bombardment mixture correlated with more intense X-gal staining. At the gold/tungsten ratio of 1:4 the microparticles penetrated the musculus glutaeus superficialis and transfected the underlying musculus glutaeus medius as well. Immuno-cytochemical assay for human dystrophin revealed dystrophin positive myofibers (DPM) in the bombarded area up to two months after transfection. The proportion of DMP varied from 2.5% on day 17 up two 5% on day 60 after bombardment compared to only 0.5% in the control mdx mice. These results suggest the applicability of particle bombardment for gene delivery into muscle fibers.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biolistics / methods
  • DNA, Complementary
  • Dystrophin / biosynthesis*
  • Dystrophin / genetics
  • Genetic Vectors
  • Humans
  • Leukemia Virus, Murine
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Inbred mdx
  • Muscle Fibers, Skeletal / metabolism*
  • Muscle, Skeletal / metabolism*
  • Promoter Regions, Genetic
  • Transfection / methods
  • beta-Galactosidase / biosynthesis*
  • beta-Galactosidase / genetics

Substances

  • DNA, Complementary
  • Dystrophin
  • beta-Galactosidase