Ultrasound-targeted antisense oligonucleotide attenuates ischemia/reperfusion-induced myocardial tumor necrosis factor-alpha

Ultrasound contrast agents are now emerging as effective vehicles for delivering therapeutic agents to target tissues. In the present study, we used ultrasound-targeted, contrast-bound antisense oligonucleotides to inhibit the expression of tumor necrosis factor-alpha (TNF-α), a proinflammatory cytokine with negative inotropic effects. We compared the efficacy of left ventricular vs. intravenous administration and determined the optimal time for delivery. WKY rats were treated with perfluorocarbon-exposed sonicated dextrose albumin (PESDA) microspheres incubated with 100 μg of antisense oligonucleotide directed against TNF-α. Contrast was infused into either the superior vena cava or the left ventricular cavity along with simultaneous application of ultrasound. Twenty-four hours later, the animals underwent 15 min of ischemia and 2 h reperfusion. Control animals underwent sham operation only, ischemia/reperfusion only, or received PESDA only. A second group received treatment just prior to, or immediately after the onset of ischemia. At the end of the experimental period, hearts were removed and analyzed for TNF-α by northern and western blotting. While no TNF-α expression was detected in sham-operated animals, robust expression of TNF-α mRNA and protein was seen in controls treated with ultrasound and PESDA alone. In contrast, intravenous or left ventricular administration of antisense oligonucleotides significantly inhibited ischemia/reperfusion-induced TNF-α expression. Direct delivery into the left ventricular cavity was more effective than intravenous administration, and delivery just prior to ischemia was most effective in attenuating TNF-α expression. Furthermore, attenuation of TNF-α expression also significantly inhibited other post-ischemic inflammatory mediators including IL-1β and intercellular adhesion molecule-1 (ICAM-1). Thus, ultrasound-targeted antisense oligonucleotides can effectively attenuate post-ischemic cytokine expression when delivered in a clinically relevant time frame, obviating the need for pretreatment.