EFFICIENT AND STABLE ADENOASSOCIATED VIRUS-MEDIATED TRANSDUCTION IN THE SKELETAL-MUSCLE OF ADULT IMMUNOCOMPETENT MICE

Recombinant adeno-associated virus (rAAV) vectors were evaluated for g ene transfer into the skeletal muscle of adult immunocompetent mice, A study using a vector encoding nuclear localized beta-galactosidase (r AAV-nls-lacZ) examined: (i) the efficiency and duration of transgene e xpression; (ii) the status of the AAV genome in the transduced fibers; and (iii) the possibility of improving gene transfer by inducing musc le regeneration, In the absence of regeneration, the injection of 1.7 x 10(7) particles in the quadriceps resulted in gene transfer to 10-70 % of myofibers, Histological analysis indicated that the vector was ab le to reach myofiber nuclei distant from the injection point, Cellular infiltrates were absent at early time points but became conspicuous i n the vicinity of some positive fibers at 4-8 weeks and subsided by 26 weeks, Southern analysis indicated that one to three copies of the ve ctor genome were present per cell genome equivalent, They were associa ted with high-molecular-weight DNA in the form of tandem oligomers or interlocked circles, Gene transfer was not facilitated in the regenera ting muscle, Rather, an early inflammatory response resulted in the el imination of most positive fibers after 8 weeks, The presence of regen erated fibers with beta-galactosidase-positive nuclei suggested that m yoblasts had been transduced and were able to fuse to form new fibers, Gene transfer in the absence of immune reactions against the transgen e product was studied by injecting mice with a rAAV carrying the murin e erythropoietin (mEpo) cDNA, Dose-dependent elevation in the hematocr it was measured for over 200 days and corresponded to 5- to 20-fold in creases in plasma Epo levels, We conclude that AAV vectors efficiently and stably transduce post-mitotic muscle fibers and myoblasts in vivo .