Protection by Nitric Oxide Donors of Isolated Rat Hearts Is Associated with Activation of Redox Metabolism and Ferritin Accumulation.

Preconditioning (PC) procedures (ischemic or pharmacological) are powerful procedures used for attaining protection against prolonged ischemia and reperfusion (I/R) injury, in a variety of organs, including the heart. The detailed molecular mechanisms underlying the protection by PC are however, complex and only partially understood. Recently, an 'iron-based mechanism' (IBM), that includes de novo ferritin synthesis and accumulation, was proposed to explain the specific steps in cardioprotection generated by IPC. The current study investigated whether nitric oxide (NO), generated by exogenous NO-donors, could play a role in the observed IBM of cardioprotection by IPC. Therefore, three distinct NO-donors were investigated at different concentrations (1-10 μM): sodium nitroprusside (SNP), 3-morpholinosydnonimine (SIN-1) and S-nitroso-N-acetylpenicillamine (SNAP). Isolated rat hearts were retrogradely perfused using the Langendorff configuration and subjected to prolonged ischemia and reperfusion with or without pretreatment by NO-donors. Hemodynamic parameters, infarct sizes and proteins of the methionine-centered redox cycle (MCRC) were analyzed, as well as cytosolic aconitase (CA) activity and ferritin protein levels. All NO-donors had significant effects on proteins involved in the MCRC system. Nonetheless, pretreatment with 10 μM SNAP was found to evoke the strongest effects on Msr activity, thioredoxin and thioredoxin reductase protein levels. These effects were accompanied with a significant reduction in infarct size, increased CA activity, and ferritin accumulation. Conversely, pretreatment with 2 μM SIN-1 increased infarct size and was associated with slightly lower ferritin protein levels. In conclusion, the abovementioned findings indicate that NO, depending on its bio-active redox form, can regulate iron metabolism and plays a role in the IBM of cardioprotection against reperfusion injury

Experimental protocols.

<p>Experimental protocols.</p

Activity of cytosolic aconitase.

<p>Activity of cytosolic aconitase.</p

Infarct sizes of the rat hearts after exposure to the different experimental protocols.

<p>Data are presented as Mean ± SEM.* p < 0.05 versus I/R.</p

Ferritin and iron content at various times of the perfusion protocol.

<p>Ferritin and iron content at various times of the perfusion protocol.</p

Thioredoxin and thioredoxin-reductase protein levels.

<p>Thioredoxin and thioredoxin-reductase protein levels.</p

Activities of methionine sulfoxide reductase.

<p>Activities of methionine sulfoxide reductase.</p

Hemodynamic parameters of the rat hearts after exposure to the different experimental protocols.

<p>Hemodynamic parameters of the rat hearts after exposure to the different experimental protocols.</p

The Methionine-Centered Redox Cycle.

<p>The formation of methionine sulfoxide (MetO) can result from the oxidation of free methionine, or a methionyl residue of a protein. Additional oxidation will generate methionine sulfone (MetO₂), a product that is almost irreversible in biological systems, and can cause protein denaturation. MetO can be reduced by methionine sulfoxide reductases (MsrA or MsrB isoform), through thioredoxin (Trx). Thioredoxin reductase (TrxR) regenerates the oxidized Trx (Trx_ox) via critical components of the cellular redox system, NADP/NADP(H) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0159951#pone.0159951.ref032" target="_blank">32</a>].</p