Analysis of tissue-specific gene delivery by recombinant adenoviruses containing cardiac-specific promoters.

Abstract

To approach heart muscle diseases by gene transfer, an adenoviral vector system was intended to be established suitable for gene expression in ventricular and/or atrial myocardium. Two adenoviral vectors (Ad-mhcLuc, Ad-mlcLuc) were constructed, in which the luciferase reporter gene is under control of either the ventricle-specific myosin light chain-2 (mlc-2v) or the atrial- and ventricular-specific alpha-myosin heavy chain (alpha-mhc) promoter. For controls, a recombinant adenovirus without promoter (Ad-Luc) and one with the Rous sarcoma virus (rsv) promoter (Ad-rsvLuc) were generated. A volume of 20 microliters containing 2 x 10(9) plaque forming units (pfu) of the recombinant adenoviruses Ad-mhcLuc, Ad-mlcLuc, Ad-rsvLuc or Ad-Luc was injected into the cardiac cavity or the quadriceps femoris muscle of neonatal rats. After five days animals were sacrificed and nine different tissues were analyzed for reporter gene expression by detection of light activity relative to mg of tissue. Injections of recombinant adenoviruses into the cardiac cavity of neonatal rats resulted in heart-specific gene expression of Ad-mlcLuc (20 fold of Ad-Luc; 11% of Ad-rsvLuc), whereas Ad-mhcLuc gave mainly luciferase activity in the heart (6.5-fold of Ad-Luc; 3% of Ad-rsvLuc) with additional activity in lung and liver (2-4 fold of Ad-Luc). In the ventricular tissue Ad-mlcLuc revealed a 35-fold higher luciferase activity, whereas Ad-mhcLuc, Ad-rsvLuc and Ad-Luc showed only 2-fold higher luciferase activities compared to the atrium. Viral DNA in atrial and ventricular tissue was detected by PCR at approximately the same abundance independent of the injected type of adenovirus. Direct injection of Ad-mhcLuc and Ad-mlcLuc into the thigh muscle revealed only background luciferase activities. In the adenoviral system only the mlc-2v promoter may fulfil the safety requirements for a myocardial specific gene expression with a high selectivity for the ventricular myocardium, thus providing a promising tool for future gene therapy of cardiomyopathies.