Inhibition of matrix metalloproteinases prevents peroxynitrite-induced contractile dysfunction in the isolated cardiac myocyte
Inhibition of matrix metalloproteinases prevents peroxynitrite-induced contractile dysfunction in the isolated cardiac myocyte
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DOI：
10.1038/sj.bjp.0707621
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发表时间：
2008-02-01
影响因子：
7.3
通讯作者：
Schulz, R.
中科院分区：
文献类型：
--
作者：
Leon, H.;Baczko, I.;Schulz, R.
背景和目的：强效氧化剂过氧亚硝酸盐 (ONOO-) 部分通过激活基质金属蛋白酶-2 (MMP-2) 诱导完整心脏机械功能障碍。这种作用可能与 MMP 对细胞外基质蛋白的蛋白水解作用无关。本研究的目的是在单个心肌细胞水平上检查 ONOO- 对收缩功能的影响，以及这是否包括 MMP 的作用。 实验方法：从成年大鼠中新鲜分离的心室肌细胞在 21°C 下用 Krebs-Henseleit 缓冲液进行灌注，并以 0.5 Hz 起搏。使用视频边缘检测器测量收缩性。 ONOO-或分解的ONOO-（载体对照）共同输注40分钟以上，以评估收缩停止时间（CCT）。 ONOO- 对细胞内 [Ca2+] 的影响是在负载有钙绿-1 AM 的心肌细胞中测定的。通过明胶酶谱法测量 MMP-2 活性。主要结果：ONOO-（30-600 μM）导致 CCT 浓度依赖性降低。接受 300 mM ONOO- 处理的肌细胞的 CCT 比分解的 ONOO- 短（14.9+1.5 vs 32.2+3.5 分钟，n = 7-8；P < 0.05），并且显示出 MMP-2 活性增加。 MMP 抑制剂多西环素 (100 μM) 或 PD 166793 (2 μM) 减少了 300 mM ONOO- 诱导的 CCT 下降。 ONOO- 导致钙瞬时停止时间缩短，并显着改变细胞内 [Ca2+] 稳态，而强力霉素可部分阻止这种变化。 结论和意义：这是第一个证明，抑制 MMP 可通过与细胞外基质蛋白的蛋白水解无关的作用，保护心肌细胞免受 ONOO- 诱导的收缩衰竭。
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.