Loss of Mitochondrial Dynamics Proteins Mitofusin‐2 and Drp‐1 in Myocardial Ischemia‐Reperfusion Injury Is Prevented by Matrix Metalloproteinase‐2 Preferring Inhibitors
Bibliographic record
Abstract
Objective Matrix metalloproteinase‐2 (MMP‐2) is a ubiquitous protease that cleaves several extracellular and intracellular proteins. MMP‐2 is activated intracellularly in response to enhanced oxidative stress resulting from myocardial ischemia/reperfusion (I/R) injury. Oxidative stress impairs mitochondrial function which is regulated by different proteins including mitofusin‐2 (Mfn‐2) and dynamin‐related protein‐1 (Drp‐1) which control mitochondrial dynamics involving fusion and fission, respectively. As both proteins are localized at the mitochondrial outer membrane and MMP‐2 is localized at the mitochondrial‐endoplasmic reticulum associated membrane we hypothesized that MMP‐2 may proteolyze Mfn‐2 and Drp‐1. We therefore investigated whether inhibition of MMP‐2 could protect against the loss of mitochondrial dynamics proteins during I/R injury. Methods Hearts isolated from 3 month old C57BL/6J mice were perfused according to Langendorff at constant pressure and subjected to I/R injury (30 min ischemia and 40 min reperfusion) in absence or presence of MMP‐2 preferring inhibitors ARP‐100 (10 µM) or ONO‐4817 (50 µM) or their vehicle (n=5 hearts per group). Effects on mechanical function were recorded. At the end of reperfusion, the hearts were homogenized and subcellular fractions were prepared. The degradation of troponin I (TnI), an intracellular MMP‐2 target, was measured in the cytosolic fraction as a marker of MMP‐2 activity. Levels of Mfn‐2 and Drp‐1 in the mitochondrial fraction were also measured using western blot. In silico analysis of potential MMP‐2 cleavage sites was performed using Procleave. Results ARP‐100 or ONO‐4817 significantly increased left ventricular developed pressure compared to vehicle‐treated I/R hearts (% of pre‐ischemic baseline value at R40: IR+vehicle 25.5±3.2, IR+ARP 50.1±3.2, IR+ONO 57.3±1.6%, p<0.05). Similarly, both inhibitors significantly improved the rates of contraction and relaxation (+/‐dP/dt). TnI loss was increased in I/R hearts as shown by a significant reduction in TnI (~30 kDa) and the appearance of an ~22 kDa band. ARP‐100 or ONO‐4817 attenuated TnI loss, indicating MMP‐2 activation. Levels of Mfn‐2 and Drp‐1 in the mitochondrial fraction were significantly reduced in the I/R group and ARP‐100 or ONO‐4817 attenuated this reduction (p<0.05). Similar results were obtained using a different Mfn‐2 antibody that binds to its C‐terminus. In silico analysis of both mouse Mfn‐2 and Drp‐1 sequences showed several potential sites that could be targeted by MMP‐2. Conclusions During myocardial I/R injury, MMP‐2 may affect mitochondrial dynamics by proteolysis of Mfn‐2 and Drp‐1. Inhibition of MMP‐2 activity could protect against cardiac contractile dysfunction in part by preserving these proteins affecting mitochondrial fusion and fission.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".