Human bone marrow stromal cells are efficiently transduced by vesicular stomatitis virus-pseudotyped retrovectors without affecting subsequent osteoblastic differentiation

This study tested the transduction efficiency of human bone marrow stromal cells (hBMSCs) with vesicular stomatitis virus (VSV)-pseudotyped retrovecto rs and their subsequent osteogenic differentiation in vitro. Two different retrovectors encoding beta -galactosidase (beta -gal) or enhanced green flu orescent protein (eGFP) as marker genes were examined for transduction of h BMSCs. hBMSCs were obtained from bone marrow filtrates of normal donors (ag ed 5-35 years), cultured in alpha -minimal essential medium (alpha -MEM) co ntaining 10% fetal calf serum and infected with retrovectors soon after the adherent cells started to form individual colonies. Transduced hBMSCs were observed to express eGFP protein 4-7 days after infection in primary cultu res, and the majority of hBMSCs were eGFP-positive. hBMSCs were also staine d for beta -gal in the secondary cultures and virtually all hBMSCs expresse d beta -gal activity. Transduced hBMSCs were examined for their osteogenic potential. These cells were found to express markers of osteogenic differen tiation, including alkaline phosphatase, type I collagen, bone sialoprotein , decorin, and osteocalcin, as strongly as uninfected control cells. Minera lization was also induced by dexamethasone in transduced cells as well as c ontrol cells. These results demonstrate that hBMSCs are highly susceptible to infection with VSV-pseudotyped retrovectors with the majority. p of cult ured cells expressing the viral transgenes without antibiotic selection. Tr ansduced cells retain their osteogenic potential in vitro. hBMSCs are a pro mising cellular vehicle for systemic human gene therapy and VSV-pseudotyped retrovectors should be effective for their in vitro transduction prior to cellular engraftment. (C) 2001 by Elsevier Science Inc. All rights reserved .