The present study is the first to demonstrate myocardial preconditioning against infarction by the lanthanide cation, gadolinium (Gd). We have previously demonstrated that Gd attenuates contractile dysfunction associated with various pathophysiologic conditions, including abnormal stretch, ischemia-reperfusion (stunning) and cardiomyopathy (1
). In the current study, administration of Gd in the setting of ischemia-reperfusion (IR) reduced infarct size in a dose-dependent manner by a mechanism involving activation of both JAK/STAT and p44 MAPK pathways, as well as KATP
channels. The infarct reducing effect of Gd is dose-dependent, with maximum reduction in infarct size manifest at a dose of 20 μmol/kg. We did not observe a reduction in infarct size with Gd at 40 μmol/kg, the dose used by Gysembergh et al
to block volume overload-induced cardioprotection (5
), however we did observe protection with Gd at doses of 5–30 μmol/kg. These findings illustrate the dose-dependent nature of protection against infarction with Gd and highlight the need to perform dose-response studies when determining the potential cardioprotective properties of novel agents.
The temporal effects of Gd with respect to IR observed in the present study are important, as they suggest potential broad applications for the use of Gd in different clinical settings. Similar to previous observations on stunned myocardium (3
), Gd was effective in the present study at protecting the heart from infarction when given prior to onset of ischemia or when given during ischemia (in particular, when given only one minute prior to reperfusion). Thus Gd may offer cardioprotection when administered prior to ischemia in planned settings where IR might occur, such as elective coronary bypass surgery (CABG) or elective percutaneous intervention (PCI). It may also be useful however, in acute coronary syndromes, where administering Gd immediately prior to reperfusion (by either emergency CABG or PCI) could help reduce infarct size. The magnitude of immediate protection afforded by Gd in the current study is less than the protection reported with ischemic preconditioning (31% vs. 86% reduction in infarct size) (11
), but the fact that Gd can be given one minute before reperfusion and still provide significant infarct reduction makes it perhaps more clinically important than ischemic preconditioning. In addition to these temporal relationships, we observed that a single dose of Gd confers delayed cardioprotection for up to 72 hours to an extent that is comparable to delayed cardioprotection observed with ischemic preconditioning (66% vs. 67% reduction in infarct size) (12
). This effect may also have important clinical applications in revascularization procedures, where recurrent ischemia may occur early after the procedure from graft occlusion or PCI failure.
The cardioprotective effects of Gd observed in the present study appear to be mediated through multiple mechanisms, including both the JAK/STAT pathway and the p42/p44 MAPK component of the Reperfusion Injury Salvage Kinase pathway. The JAK/STAT pathway plays an integral role in the response of the myocardium to various insults, including myocardial infarction. Furthermore it has a prominent role in cardioprotective therapies such as ischemic preconditioning. Further studies are needed to determine if other components of preconditioning such as PKC, adenosine receptors, and Akt/PI3
kinase play a role in Gd-induced cardioprotection. The JAK/STAT pathway also plays an essential role in the development of delayed preconditioning (13
). Our study demonstrates that activation of JAK/STAT is necessary for both the immediate and delayed cardioprotective effects of Gd to be manifest. We propose that Gd stimulates the heart to conduct a signal via
the JAK/STAT pathway to the nucleus to activate nuclear transcription factors, resulting in the synthesis of genes such as inducible nitric oxide synthase, hemoxygenase-1, aldose reductase, and Mn superoxide dismutase to confer delayed protection against injury from myocardial infarction. As JAK resides in the vicinity of the sarcolemma, we speculate that Gd may be acting through a cell surface receptor to activate JAK/STAT. The notion that Gd can act as a ligand to a receptor protein is supported by recent studies showing the metabotropic glutamate receptor 1α binds Gd resulting in activation of two distinct pathways involving calcium (mediated through the Gq family of heterotrimeric guanine nucleotide-binding proteins) and adenosine 3′,5′-monophosphate (cAMP, mediated through Gs proteins) (14
). Furthermore it is known that the JAK/STAT and p42/44 MAPK pathways can be simultaneously activated in the heart (15
) to confer protection against injury from ischemia/reperfusion.
The present study suggests that the cardioprotective effects of Gd are also mediated in part through ATP-dependent potassium (KATP
) channels. These channels, highly expressed in myocardial sarcolemmal and thought to be expressed in myocardial mitochondria, have been found to serve as mediators of cardioprotection (17
). Gadolinium is known to exert effects on a broad range of ion channels in the heart. Recently Babich et al
) proposed that Gd binding stabilizes ion channels in an inactivated state similar to the actions of other calcium channel blockers such as the dihydropyridines. In addition to L-type calcium channels and the sodium-calcium exchanger, another pathway exists for entry of calcium into the ventricular myocyte that is gadolinium-sensitive and distinct from the B-type channel (21
). Furthermore Gd is commonly used to interrupt stretch-mediated changes in intracellular ion flux in the heart through a presumed blockade of specific stretch-activated ion channels (22
). Since mechanical stretch causes pathophysiologic changes in the myocardium (1
), and regional myocardial ischemia results in abnormal stretch (2
), the effects of Gd observed herein may also be related to modulation of ion fluxes that are caused by ischemia-induced stretch. Effects on KATP
and perhaps other ion channels may be of particular importance when Gd is administered one minute before reperfusion, as cell signaling to induce gene expression (as postulated above) would be a less likely factor in such a short time frame.
The effects of Gd on infarct size were not altered by treatment with free radical scavengers, suggesting that the mechanism underlying the immediate cardioprotective effects of Gd is not related to oxidative stress. The selection of the in vivo
doses for the radical scavengers 2-mercaptoproprionyl glycine (20mg/kg) and N-acetyl cysteine (150 mg/kg) were based on previous studies from our lab (29
) and from another laboratory, respectively (30
In summary, a single treatment with Gd conferred immediate protection against injury caused by regional ischemia and reperfusion in the rat heart, with protection mediated partially by JAK-2, STAT3, p42/p44 MAPK and KATP channels. Gd also conferred delayed cardioprotection by a mechanism associated with the activation of JAK-2. Further studies are warranted to identify additional pathways by which Gd confers cardioprotection. Our results suggest that Gd directly protects the heart and may represent a novel approach for the treatment of myocardial IR injury.