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

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 degrees 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 [Ca2+] were determined in myocytes loaded with calcium green-1 AM. MMP-2 activity was measured by gelatin zymography. Key results: ONOO- (30-600 mu M) caused a concentration-dependent reduction in CCT. Myocytes subjected to 300 mM 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 mu M) or PD 166793 (2 mu M) reduced the decline in CCT induced by 300 mM ONOO-. ONOO- caused shorter calcium transient cease time and significant alterations in intracellular [Ca2+] 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.