Inhibition of matrix metalloproteinases prevents peroxynitrite-induced contractile dysfunction in the isolated cardiac myocyte

Abstract

Background and purpose: The potent oxidant peroxynitrite (ONOO -) induces mechanical dysfunction in the intact heart in part through activation of matrix metalloproteinase-2 (MMP-2). This effect may be independent of the proteolytic actions of MMPs on extracellular matrix proteins. The purpose of this study was to examine the effects of ONOO - on contractile function at the level of the single cardiac myocyte and whether this includes the action of MMPs. Experimental approach: Freshly isolated ventricular myocytes from adult rats were superfused with Krebs-Henseleit buffer at 21°C and paced at 0.5 Hz. Contractility was measured using a video edge-detector. ONOO - or decomposed ONOO - (vehicle control) were co-infused over 40 min to evaluate the contraction cease time (CCT). The effects of ONOO - on intracellular [Ca 2+] were determined in myocytes loaded with calcium green-1 AM. MMP-2 activity was measured by gelatin zymography. Key results: ONOO - (30-600 μM) caused a concentration-dependent reduction in CCT. Myocytes subjected to 300 μM ONOO - had a shorter CCT than decomposed ONOO - (14.9+1.5 vs 32.2+3.5 min, n=7-8; P<0.05) and showed increased MMP-2 activity. The MMP inhibitors doxycycline (100 μM) or PD 166793 (2 μM) reduced the decline in CCT induced by 300 μM ONOO -. ONOO - caused shorter calcium transient cease time and significant alterations in intracellular [Ca 2+] homoeostasis which were partially prevented by doxycycline. Conclusions and implications: This is the first demonstration that inhibition of MMPs protects the cardiac myocyte from ONOO --induced contractile failure via an action unrelated to proteolysis of extracellular matrix proteins. © 2008 Nature Publishing Group All rights reserved.