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(Circulation Research. 2008;103:1120.)
© 2008 American Heart Association, Inc.

Cellular Biology

Inducible Nitric Oxide Synthase Expression and Cardiomyocyte Dysfunction During Sustained Moderate Ischemia in Pigs

Frank R. Heinzel, Petra Gres, Kerstin Boengler, Alexej Duschin, Ina Konietzka, Tienush Rassaf, Julia Snedovskaya, Stephanie Meyer, Andreas Skyschally, Malte Kelm, Gerd Heusch, Rainer Schulz

From the Institute of Pathophysiology (F.R.H., P.G., K.B., A.D., I.K., J.S., S.M., A.S., G.H., R.S.), University of Essen Medical School, Germany; Department of Cardiology, Pneumology, and Vascular Medicine (T.R., M.K.), University Hospital Aachen, Germany; and Abteilung für Kardiologie (F.R.H.), Medizinische Universität Graz, Austria.

Correspondence to Dr Rainer Schulz, University of Essen Medical School, Institute of Pathophysiology, Hufelandstrasse 55, Essen 45122, Germany. E-mail rainer.schulz{at}uk-essen.de

In acute myocardial ischemia, regional blood flow and function are proportionally reduced. With prolongation of ischemia, function further declines at unchanged blood flow. We studied the involvement of an inflammatory signal cascade in such progressive dysfunction and whether dysfunction is intrinsic to cardiomyocytes. In 10 pigs, ischemia was induced by adjusting inflow into the cannulated left anterior coronary artery to reduce coronary arterial pressure to 45 mm Hg (ISCH); 4 pigs received the inducible nitric oxide synthase (iNOS) inhibitors aminoguanidine or L-N⁶-(1-iminoethyl)-lysine during ISCH (ISCH+iNOS-Inhib); 6 pigs served as controls (SHAM). Anterior (AW) and posterior (PW) systolic wall thickening (sonomicrometry) were measured. After 6 hours, nitric oxide (NO) synthase (NOS) protein expression, NOS activity, and NO metabolites (nitrite/nitrate/nitroso species) were quantified in biopsies isolated from AW and PW. Cardiomyocyte shortening and intracellular calcium (Indo-1 acetoxymethyl ester) were measured without and with the NOS substrate L-arginine (100 µmol/L). In ISCH, AW wall thickening decreased from 42±4% (baseline) to 16±3% (6 hours). Wall thickening remained unchanged in ISCH-PW and SHAM-AW/PW. NOS2 (iNOS) protein expression and activity, but not NOS3 (endothelial NO synthase), were increased in ISCH-AW and ISCH-PW. iNOS expression correlated with increased nitrite contents. Cardiomyocyte shortening was reduced in ISCH-AW versus SHAM-AW (4.4±0.3% versus 5.6±0.3%). L-Arginine reduced cardiomyocyte shortening further in ISCH-AW (to 2.8±0.2%) and ISCH-PW (3.4±0.4% versus 5.4±0.4%) but not in SHAM or in ISCH+iNOS-Inhib; intracellular [Ca²⁺] remained unchanged. With L-arginine, in vitro AW cardiomyocyte shortening correlated with in vivo AW wall thickening (r=0.72). In conclusion, sustained regional ischemia induces myocardial iNOS expression in pigs, which contributes to contractile dysfunction at the cardiomyocyte level.

Key Words: myocardial ischemia • nitric oxide • iNOS • sphingosine • TNF-