In-vivo vs. ex-vivo adenoviral gene transfer to rabbit carotid artery: A comparative study of transgene expression
Youngblood SA., Channon KM., Shetty GA., Blazing MA., George SE.
Recombinant adenovtruses have emerged as a promising vehicle for in-vlvo gene therapy. One major limitation of this method Is decreased recombinant gene expression due to host Immunity. To assess the relative Importance of host factors In reducing transgene expression In-vlvo, we performed adenoviral gene transfer of B-Galactosldase (B-Qal) to rabbit carotid arteries (CA), Gene transfer was carried out to both CA using passive dwell for 15 min using liters of 1x10", 2x10'°, or 8x10 pfu/ml. One artery was then left in place while the contralateral artery was removed and incubated ex-vivo. At three days, we compared 6-Gal expression In the ex-vivo arteries with the In-vlvo arteries using histologie sections of X-Gal stained vessels and direct measurement of B-Qal protein in vessel Ivsates with an ELISA. Results: Vessels from 6 rabbits were studied lor B-Gal expression 3 days after gene transler. AI gene-transferred vessels stained blue with x-gal. B-Gal protein levels per mg of vessel protein are shown (Table). The highest levels of transgene expression were seen at 2x10'° ofu/ml. with reduced expression at the higher liter of 1x10" pfu/ml. β-Gal levels In ex-vrvo vessels were significantly greater than in ln-vlvo vessels (p < 0.05). Conclusions: Adenoviral gene transfer to rabbit CA was highly efficient, with only minor differences in expression across the three tilers. The reduced expression at the highest liter may represent direct viral cytotoxidty. More significantly however, 0-Gal levels in ex-vivo arteries were approximately 6-fold higher relative to paired in-vtvo arteries for each liter. Therefore, host responses to adenoviral-medlated arterial gene transfer markedly reduce transgene expression in-vivo. Titer B-Gal (noAng prolelnlhnean ±SD) rtu/ml In-VIvo Ex-Vivo 1x1011 172 881 (±14.8) 2x1010 213(151.5) 1260(±171.6) 8x109 137 (±64.8) 954(1189.4).