OBJECTIVES: Recombinant viral vectors based on the nonpathogenic parvovirus, adeno-associated virus (AAV), have a number of attractive features for gene therapy, including the ability to transduce non-dividing cells and its long-term transgene expression. In this study, an AAV vector containing bacterial beta-galactosidase gene (lacZ) was used to transduce cultured rat vascular smooth muscle cells (VSMC) in vitro and rat thoracic aortas ex vivo. METHODS: VSMC were transduced with AAV-lacZ at multiplicities of infection (MOI) ranging from 5.0 x 10(5) to 1.0 x 10(7). Expression of beta-galactosidase (beta-gal) in VSMC was evaluated by X-gal staining and a beta-gal ELISA method. Excised rat aortas were incubated with medium containing AAV-lacZ. Expression of beta-gal in the aortic segments was evaluated by X-gal staining. RESULTS: With increasing MOI, up to 50% of cultured VSMC were positive by X-gal staining and the beta-gal expression increased up to 15 ng/mg protein. The expression gradually decreased during the culture but was detectable for at least 1 month. In the ex vivo study, AAV vectors transduced endothelial and adventitial cells in rat aortic segments, while no expression was seen in medial VSMC. CONCLUSIONS: AAV vectors can efficiently transduce rat VSMC in vitro. AAV-mediated ex vivo gene transfer into the normal aorta resulted in efficient gene transfer into endothelial and adventitial cells but not into medial VSMC. These findings suggest that AAV-based vectors are promising for use in cardiovascular gene therapy.
OBJECTIVES: Recombinant viral vectors based on the nonpathogenic parvovirus, adeno-associated virus (AAV), have a number of attractive features for gene therapy, including the ability to transduce non-dividing cells and its long-term transgene expression. In this study, an AAV vector containing bacterial beta-galactosidase gene (lacZ) was used to transduce cultured rat vascular smooth muscle cells (VSMC) in vitro and rat thoracic aortas ex vivo. METHODS: VSMC were transduced with AAV-lacZ at multiplicities of infection (MOI) ranging from 5.0 x 10(5) to 1.0 x 10(7). Expression of beta-galactosidase (beta-gal) in VSMC was evaluated by X-gal staining and a beta-gal ELISA method. Excised rat aortas were incubated with medium containing AAV-lacZ. Expression of beta-gal in the aortic segments was evaluated by X-gal staining. RESULTS: With increasing MOI, up to 50% of cultured VSMC were positive by X-gal staining and the beta-gal expression increased up to 15 ng/mg protein. The expression gradually decreased during the culture but was detectable for at least 1 month. In the ex vivo study, AAV vectors transduced endothelial and adventitial cells in rat aortic segments, while no expression was seen in medial VSMC. CONCLUSIONS:AAV vectors can efficiently transduce rat VSMC in vitro. AAV-mediated ex vivo gene transfer into the normal aorta resulted in efficient gene transfer into endothelial and adventitial cells but not into medial VSMC. These findings suggest that AAV-based vectors are promising for use in cardiovascular gene therapy.
Authors: Sifeng Chen; Matthias Kapturczak; Scott A Loiler; Sergei Zolotukhin; Olena Y Glushakova; Kirsten M Madsen; Richard J Samulski; William W Hauswirth; Martha Campbell-Thompson; Kenneth I Berns; Terence R Flotte; Mark A Atkinson; C Craig Tisher; Anupam Agarwal Journal: Hum Gene Ther Date: 2005-02 Impact factor: 5.695
Authors: Susan T Laing; Hyunggun Kim; Jonathan A Kopechek; Devang Parikh; Shaoling Huang; Melvin E Klegerman; Christy K Holland; David D McPherson Journal: J Liposome Res Date: 2010-06 Impact factor: 3.648
Authors: Divya Pankajakshan; Toluwalope O Makinde; Rohit Gaurav; Michael Del Core; George Hatzoudis; Iraklis Pipinos; Devendra K Agrawal Journal: J Surg Res Date: 2011-03-21 Impact factor: 2.192
Authors: A-M Lompré; L Hadri; E Merlet; Z Keuylian; N Mougenot; I Karakikes; J Chen; F Atassi; A Marchand; R Blaise; I Limon; S W J McPhee; R J Samulski; R J Hajjar; L Lipskaia Journal: Gene Ther Date: 2013-03-28 Impact factor: 5.250