During myocardial ischemia and reperfusion (IR) injury matrix metalloproteinase-2 (MMP-2) is rapidly activated in response to oxidative stress. MMP-2 is a multifunctional protease that cleaves both extracellular and intracellular proteins. Oxidative stress also impairs mitochondrial function which is regulated by different proteins, including mitofusin-2 (Mfn-2), which is lost in IR injury. Oxidative stress and mitochondrial dysfunction trigger the NLRP3 inflammasome and the innate immune response which invokes the de novo expression of an N-terminal truncated isoform of MMP-2 (NTT-MMP-2) at or near mitochondria. We hypothesized that MMP-2 proteolyzes Mfn-2 during myocardial IR injury, impairing mitochondrial function and enhancing the inflammasome response. Isolated hearts from mice subjected to IR injury (30 min ischemia/40 min reperfusion) showed a significant reduction in left ventricular developed pressure (LVDP) compared to aerobically perfused hearts. IR injury increased MMP-2 activity as observed by gelatin zymography and increased degradation of troponin I, an intracellular MMP-2 target. MMP-2 preferring inhibitors, ARP-100 or ONO-4817, improved post-ischemic recovery of LVDP compared to vehicle perfused IR hearts. In muscle fibers isolated from IR hearts the rates of mitochondrial oxygen consumption and ATP production were impaired compared to those from aerobic hearts, whereas ARP-100 or ONO-4817 attenuated these reductions. IR hearts showed higher levels of NLRP3, cleaved caspase-1 and interleukin-1β in the cytosolic fraction, while the mitochondria-enriched fraction showed reduced levels of Mfn-2, compared to aerobic hearts. ARP-100 or ONO-4817 attenuated these changes. Co-immunoprecipitation showed that MMP-2 is associated with Mfn-2 in aerobic and IR hearts. ARP-100 or ONO-4817 also reduced infarct size and cell death in hearts subjected to 45 min ischemia/120 min reperfusion. Following myocardial IR injury, impaired contractile function and mitochondrial respiration and elevated inflammasome response could be attributed, at least in part, to MMP-2 activation, which targets and cleaves mitochondrial Mfn-2. Inhibition of MMP-2 activity protects against cardiac contractile dysfunction in IR injury in part by preserving Mfn-2 and suppressing inflammation.

在心肌缺血和再灌注 (IR) 损伤期间，基质金属蛋白酶 2 (MMP-2) 会因氧化应激而迅速激活。MMP-2 是一种多功能蛋白酶，可裂解细胞外和细胞内蛋白质。氧化应激还会损害由不同蛋白质调节的线粒体功能，包括在 IR 损伤中丢失的线粒体融合蛋白-2 (Mfn-2)。氧化应激和线粒体功能障碍会触发 NLRP3 炎症小体和先天免疫反应，从而引发 MMP-2 N 端截短亚型 (NTT-MMP-2) 在线粒体或线粒体附近的从头表达。我们假设 MMP-2 在心肌 IR 损伤过程中蛋白水解 Mfn-2，损害线粒体功能并增强炎症反应。与有氧灌注的心脏相比，受到 IR 损伤（30 分钟缺血/40 分钟再灌注）的小鼠离体心脏显示左心室发展压 (LVDP) 显着降低。通过明胶酶谱观察，IR 损伤增加了 MMP-2 活性，并增加了细胞内 MMP-2 靶标肌钙蛋白 I 的降解。与载体灌注 IR 心脏相比，MMP-2 优先抑制剂 ARP-100 或 ONO-4817 改善了 LVDP 的缺血后恢复。与有氧心脏相比，从 IR 心脏分离的肌纤维中，线粒体耗氧量和 ATP 生成率受到损害，而 ARP-100 或 ONO-4817 减弱了这些降低。与需氧心脏相比，IR 心脏的胞质部分中 NLRP3、裂解的 caspase-1 和白细胞介素 1β 水平较高，而线粒体富集部分中的 Mfn-2 水平较低。ARP-100 或 ONO-4817 减弱了这些变化。免疫共沉淀显示 MMP-2 在有氧心脏和 IR 心脏中与 Mfn-2 相关。ARP-100 或 ONO-4817 还可以减少 45 分钟缺血/120 分钟再灌注心脏中的梗塞面积和细胞死亡。心肌 IR 损伤后，收缩功能和线粒体呼吸受损以及炎症反应升高可能至少部分归因于 MMP-2 激活，MMP-2 靶向并裂解线粒体 Mfn-2。抑制 MMP-2 活性可部分通过保护 Mfn-2 和抑制炎症来防止 IR 损伤中的心脏收缩功能障碍。