Transgenic MMP-2 expression induces latent cardiac mitochondrial dysfunction

Abstract

Matrix metalloproteinases (MMPs) are central to the development and progression of dysfunctional ventricular remodeling after tissue injury. We studied 6 month old heterozygous mice with cardiac-specific transgenic expression of active MMP-2 (MMP-2 Tg). MMP-2 Tg hearts showed no substantial gross alteration of cardiac phenotype compared to age-matched wild-type littermates. However, buffer perfused MMP-2 Tg hearts subjected to 30 min of global ischemia followed by 30 min of reperfusion had a larger infarct size and greater depression in contractile performance compared to wild-type hearts. Importantly, cardioprotection mediated by ischemic preconditioning (IPC) was completely abolished in MMP-2 Tg hearts, as shown by abnormalities in mitochondrial ultrastructure and impaired respiration, increased lipid peroxidation, cell necrosis and persistently reduced recovery of contractile performance during post-ischemic reperfusion. We conclude that MMP-2 functions not only as a proteolytic enzyme but also as a previously unrecognized active negative regulator of mitochondrial function during superimposed oxidative stress.

Fig. 1

Measurement in MMP-2 Tg and WT hearts of myocardial contractile performance before and after ischemia/reperfusion (I/R) or in hearts subjected to ischemic preconditioning followed by ischemia–reperfusion (IPC+I/R). (*P < 0.05, **P < 0.01 vs. WT respectively, n = 13.)

Fig. 2

Measurement of infarct size and creatine kinase (CK) release in MMP-2 Tg and WT hearts after ischemia reperfusion (I/R) or in the hearts subjected to ischemic preconditioning followed by I/R (IPC+I/R). (*P < 0.05, **P < 0.01 vs. WT, n = 6–8.)

Fig. 3

Mitochondrial oxygen consumption rates (OCR, A) and respiratory control ratios (RCR, B) from normoxic and ischemia–reperfusion (I/R) hearts, and hearts subjected to ischemic preconditioning (IPC) followed by I/R. (*P < 0.05, **P < 0.01 vs. respective WT controls; ⁺P < 0.05 vs. both normoxic and IPC+I/R WT, n = 7.) (C) Myocardial malondialdehyde (MDA) content in MMP-2 Tg and WT heart tissue after ischemia–reperfusion (I/R) and in hearts subjected to ischemic preconditioning before I/R (IPC+I/R) (*P < 0.05, **P < 0.01 vs. respective WT controls; ⁺P < 0.05 vs. WT I/R, n = 6–7).

Fig. 4

Representative electron micrographs of ultrastructure in WT and MMP-2 Tg hearts tissue before and after I/R injury. Samples were from the non-infarcted zone of the LV myocardium. (A) Normoxic WT heart (NX-WT): the sarcoplasm is filled with branching myofibrils. Mitochondria are precisely located between myofibrils and Z-lines at the level of the A-band. Both outer and inner membranes are intact. (B) Normoxic transgenic heart (NX-MMP-2 Tg) showed similar baseline morphology and electron density of mitochondria as seen in WT except for a slight increase in collagen. (C) WT heart subjected to ischemia–reperfusion (I/R-WT, ischemic zone): mitochondria are severely edematous with disorganized cristae. They are detached from myofibrils, clustered and swollen with clearly broken membranes. (D) MMP-2 Tg heart subjected to ischemia–reperfusion: (I/R-MMP-2 Tg, ischemic zone): mitochondria are edematous and detached from myofibrils as shown in I/R-WT hearts. (E) Ischemic preconditioned WT heart after ischemia–reperfusion (IPC+I/R, ischemic zone): although some myofibrils are distorted, the baseline morphology of mitochondria and organelle contents are substantially preserved, indicating that IPC reduces the severity of mitochondrial impairment after I/R. (F) Ischemic preconditioned MMP-2 Tg heart (IPC+I/R, ischemic zone): characteristics are similar to those in the MMP-2 Tg heart subjected to I/R without IPC (D), indicating preconditioning did not attenuate damage to the myocardial ultrastructure (EM amplification 6700×).