Adenoviral gene transfer of eNOS: high-level expression in ex vivo expanded marrow stromal cells.
Endothelial nitric oxide synthase (eNOS) is an attractive target for cardiovascular gene therapy. Marrow stromal cells (MSCs), also known as mesenchymal stem cells, hold great promise for use in adult stem cell-based cell and gene therapy. To determine the feasibility of adenoviral-mediated eNOS gene transfer into ex vivo expanded MSCs, rat MSCs (rMSCs) were isolated, expanded ex vivo, and transduced with Ad5RSVeNOS, an adenoviral vector containing the eNOS gene under the control of the Rous sarcoma virus promoter. The presence of eNOS protein in Ad5RSVeNOS-transduced rMSCs was confirmed by immunohistochemical and Western blot analysis. Transduction efficiency was dose dependent, and eNOS transgene expression in rMSCs persisted for > or =21 days in culture. The rMSCs retained multipotential differentiation capability after adenoviral-mediated eNOS gene transfer. Furthermore, intracavernosal injection of Ad5RSVeNOS-transduced rMSCs increased the expression of eNOS in the corpus cavernosum, and stem cells were identified within corporal sinusoids. These findings demonstrate that replication-deficient recombinant adenovirus can be used to engineer ex vivo expanded rMSCs and that high-level eNOS transgene expression can be achieved, pointing out the clinical potential of using this novel adult stem cell-based gene therapy method for the treatment of cardiovascular diseases.
Deng
Weiwen
W
Department of Pharmacology, SL83, Tulane University Health Sciences Center, 1430 Tulane Avenue, New Orleans, LA 70112, USA.
Bivalacqua
Trinity J
TJ
Chattergoon
Natasha N
NN
Hyman
Albert L
AL
Jeter
James R
JR
Kadowitz
Philip J
PJ
D000256:Adenoviridae / Q000201:enzymology* / Q000235:genetics*; D000818:Animals; D015153:Blotting, Western; D001854:Bone Marrow Cells / Q000166:cytology / Q000201:enzymology*; D002417:Cattle; D005786:Gene Expression Regulation / Q000502:physiology*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D007276:Injections, Intraventricular; D008297:Male; D019001:Nitric Oxide Synthase / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics*; D052250:Nitric Oxide Synthase Type III; D015202:Protein Engineering / Q000379:methods; D051381:Rats; D011914:Rats, Inbred BN; D017154:Stromal Cells / Q000166:cytology / Q000201:enzymology; D014161:Transduction, Genetic; D019076:Transgenes
D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C496316:Nos3 protein, rat