Critical Role of AT₁ Receptor Expression After Ischemia/Reperfusion in Isolated Rat Hearts

Beneficial Effect of Antisense Oligodeoxynucleotides Directed at AT₁ Receptor mRNA

From the Departments of Medicine, Pediatrics, and Physiology, University of Florida, and the VA Medical Center, Gainesville, Fla.

Correspondence to J.L. Mehta, MD, PhD, Department of Medicine, University of Florida College of Medicine, 1600 Archer Rd, PO Box 100277 JHMHC, Gainesville, FL 32610. E-mail mehta{at}medmac.ufl.edu

Abstract—To examine the relevance of angiotensin II type 1 receptor (AT₁R) expression in the determination of myocardial function after ischemia/reperfusion, Sprague-Dawley rats were treated intravenously with antisense oligodeoxynucleotides (AS-ODNs) directed at AT₁R mRNA (100 µg/rat, n=9) or scrambled antisense oligodeoxynucleotides (Scr-ODNs, 100 µg/rat, n=6). Both AS-ODNs and Scr-ODNs were given along with 300 µg/rat of liposome DOTAP/DOPE, a positive electron carrier (wt:wt=1:1). The hearts from AS-ODN– or Scr-ODN–treated rats were excised 24 hours later, perfused in vitro, and subjected to 25 minutes of global ischemia followed by 30 minutes of reperfusion. Parallel groups of rats were given the specific AT₁R antagonist losartan (10 mg/kg IV, n=6) or saline (n=7) 4 to 6 hours before excising the hearts. Ischemia/reperfusion resulted in a significant increase in myocardial AT₁R expression (autoradiography and binding assay) and myocardial dysfunction, indicated by increases in coronary perfusion pressure and left ventricular end-diastolic pressure and a decrease in developed left ventricular pressure (all P<0.01 versus baseline) in the saline-treated group. AT₁R protein and mRNA levels also increased in ischemic/reperfused myocardial tissues. Administration of AS-ODNs or losartan, but not Scr-ODNs, preserved myocardial function and blocked the increased AT₁R binding after ischemia/reperfusion (both P<0.01). Myocardial AT₁R mRNA levels were not affected by either AS-ODNs or losartan, and the AT₁R protein levels were significantly reduced by AS-ODN, but not losartan, treatment. Plasma angiotensin II levels increased after administration of losartan but not after administration of AS-ODNs. These observations imply a critical role of AT₁R upregulation in determining myocardial function immediately after ischemia/reperfusion. AS-ODNs to AT₁R mRNA may be more beneficial than losartan, because losartan does not affect the plasma angiotensin II level. The sustained increase in AT₁R mRNA, but diminished protein expression, in rat hearts treated with AS-ODNs suggests that AS-ODNs block AT₁R at the translational level.

Key Words: angiotensin II • antisense • AT₁ receptor • ischemia/reperfusion • losartan • mRNA

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