Efficient gene transfer into normal human skeletal cells using recombinantadenovirus and conjugated adenovirus-DNA complexes

In order to assess efficient DNA gene transfer into human primary cell cult ures derived from the skeleton we tested two viral-based procedures. First, replication-deficient recombinant adenoviruses (ADV) were used to infect p ost-confluent human marrow stromal fibroblasts (HMSF) and human trabecular bone (HTB) cells. Both cell types were readily infected by modified adenovi ral vectors carrying a reporter gene making this virus an attractive candid ate to facilitate DNA gene transfer. In a second approach we coincubated DN A with ADV that had polylysine (PLL) covalently attached. With this ADV/PLL /DNA complex, very efficient gene transfer into multilayered HMSF and HTB c ell cultures was observed, and DNA coincubated with unmodified ADV failed t o be effectively transferred. These data imply that the covalently bound PL L more effectively binds exogenous DNA, resulting in a highly efficient int ernalization event in both cell types. Thus, this latter method has many ad vantages over conventional ADV gene transfer procedures. It is simple, rapi d, and it does not require engineering of DNA into the viral genome, thereb y allowing transfer of large fragments of DNA.