ABSTRACT
BACKGROUND: Cardiac gene therapy using the adeno-associated virus serotype 9 vector is widely used because of its efficient transduction. However, the promoters used to drive expression often cause off-target localization. To overcome this, studies have applied cardiac-specific promoters, although expression is debilitated compared to that of ubiquitous promoters. To address these issues in the context of atrial-specific gene expression, an enhancer calsequestrin cis-regulatory module 4 (CRM4) and the highly atrial-specific promoter sarcolipin were combined to enhance expression and minimize off tissue expression. METHODS: To observe expression and bio-distribution, constructs were generated using two different reporter genes: luciferase and enhanced green fluorescent protein (EGFP). The ubiquitous cytomegalovirus (CMV), sarcolipin (SLN) and CRM4 combined with sarcolipin (CRM4.SLN) were compared and analyzed using the luciferase assay, western blotting, a quantitative polymerase chain reaction and fluorescence imaging. RESULTS: The CMV promoter containing vectors showed the strongest expression in vitro and in vivo. However, the module SLN combination showed enhanced atrial expression and a minimized off-target effect even when compared with the individual SLN promoter. CONCLUSIONS: For gene therapy involving atrial gene transfer, the CRM4.SLN combination is a promising alternative to the use of the CMV promoter. CRM4.SLN had significant atrial expression and minimized extra-atrial expression.
