Myocardial Matrix Metalloproteinase‐2 Activation Impairs Amplitude and Frequency of Spontaneous Intracellular Ca <sup>2+</sup> Transients in Doxorubicin Cardiotoxicity
Notice bibliographique
Résumé
Anthracyclines, such as doxorubicin (DXR), are effective antineoplastic agents prescribed in many cancer chemotherapeutic regimens. However, 11% of cancer patients treated with DXR develop heart failure. There is no treatment to prevent DXR cardiotoxicity beyond limiting cumulative dosage and hence compromising its therapeutic efficacy. The mechanism of DXR cardiotoxicity is incompletely understood but two key characteristics are elevated oxidative stress and impaired Ca 2+ signaling (notably decreased intracellular Ca 2+ release, a key component of excitation‐contraction coupling). Elevated oxidative stress can activate matrix metalloproteinase‐2 (MMP‐2), a protease which can cleave sarcomeric proteins (e.g. α‐actinin and troponin I) and the Ca 2+ regulatory protein calreticulin. We hypothesize that MMP‐2 plays a role in DXR cardiotoxicity by proteolyzing these proteins upon activation by oxidative stress, impairing Ca 2+ signaling and cardiac contractile function. To test this, neonatal rat ventricular myocytes (NRVM) were treated with 0.5μM DXR ± the MMP inhibitors ARP‐100 or ONO‐4817 (1μM each) for 2–24h. This concentration of DXR is equivalent to that found in the blood of patients undergoing chemotherapy. 24h DXR caused no cell death in NRVM, but, as expected for a chemotherapeutic drug, this treatment protocol caused 15% of human fibrosarcoma HT1080 cells to undergo necrosis. By 12h of DXR treatment, NRVM exhibited increased oxidative stress, as shown by a 20% reduction in aconitase activity, and increased intracellular MMP‐2 activity by threefold relative to control. Intracellular MMP‐2 activation was accompanied by a twofold increase in MMP‐2 protein levels. ARP‐100 or ONO‐4817 attenuated the DXR‐increased MMP‐2 activity by 60%, but not MMP‐2 protein expression. 24h DXR reduced levels of the known MMP‐2 target troponin I by 40%, but this was not prevented by MMP inhibitors, indicating that this effect on troponin I was not due to MMP‐2. The effect of DXR‐induced MMP‐2 activation on intracellular Ca 2+ transients, likely indicative of sarcoplasmic reticulum Ca 2+ release and re‐uptake, was visualized in live NRVM using confocal microscopy and the Ca 2+ ‐binding fluorescent dye Fluo‐8L AM. 24h DXR decreased the amplitude of Ca 2+ transients by 72% and 63% under basal and isoproterenol‐stimulated (100nM) conditions, respectively. DXR also reduced the frequency of both basal and stimulated Ca 2+ transients by 73%. ARP‐100 restored both the DXR‐reduced amplitude and frequency of Ca 2+ transients. To understand how inhibiting MMP‐2 may protect against DXR‐impaired intracellular Ca 2+ transients, we measured the levels of calreticulin in NRVM lysates. However, calreticulin was unaffected by DXR. We hypothesized that SERCA, required for Ca 2+ uptake by the sarcoplasmic reticulum, could be proteolyzed by MMP‐2. MMP‐2 successfully proteolyzed SERCA in vitro, yielding a 50kDa fragment, confirmed by mass spectrometry. Future experiments will investigate the effect of DXR on SERCA levels in membrane fractions. In conclusion, DXR, at a clinically relevant concentration, increases oxidative stress and acutely activates myocardial MMP‐2. MMP‐2 activation appears to impair spontaneous intracellular Ca 2+ release in cardiomyocytes. Thus, the results of this study provide mechanistic insight into the role of MMP‐2 in DXR cardiotoxicity, and support the potential use of MMP inhibitors as an adjuvant therapy for patients receiving DXR chemotherapy. Support or Funding Information Canadian Institutes of Health Research Foundation Scheme Grant (to RS); Women and Children's Health Research Institute and Novartis Pharmaceuticals Canada Inc Graduate Studentships (to BC).
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Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,001 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,002 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».