Trimetazidine: a novel protective role via maintenance of Na+/K+-ATPase activity?

See article by El Banani et al. [31] (pages 688–696) in this issue. Trimetazidine (1-[2,3,4-trimethoxybenzyl]-piperazine; TMZ) is a piperazine compound that has been demonstrated to protect against ischaemic injury in a variety of tissues [1–4]. As a cardiovascular therapeutic agent, TMZ is protective during coronary artery bypass graft surgery [5] and catheter angioplasty [6], has been effective in the treatment of angina [7,8] and reduces infarct size when given before thrombolysis in patients with anterior myocardial infarction [9]. There have been insights into some of the molecular mechanisms of action of TMZ, however, new mechanisms continue to be discovered [10–12]. The best known action of TMZ is inhibition of the β-oxidation pathway of fatty acid metabolism [13]. Inhibition of the β-oxidation pathway during ischaemia and reperfusion concomitantly increases glucose oxidation [13]. Protection from ischaemic injury by TMZ is hypothesized to be due to the increase in glucose oxidation leading to lower rates of glycolysis and less H+ production [14–17]. Increased fatty acid oxidation and glycolysis do result in greater H+ production [18,19] and ischaemic intracellular pH changes have been observed in TMZ-treated hearts [14]. The detrimental effect of glycolytic H+ production in the ischaemic myocardium is thought to be due to increased Na+/H+ exchange activity [20–22]. H+ activate the Na+/H+ exchanger [23], causing Na+ influx. Na+ efflux via the Na+/K+-ATPase is attenuated during ischaemia [24], therefore activation of the Na+/H+ exchanger leads to increased intracellular Na+[25–27]. The increased intracellular Na+ stimulates Na+/Ca2+ exchange [28], resulting in increased intracellular Ca2+[25,29] and it is this high Ca2+ which exacerbates myocardial injury [21,25,29,30]. TMZ-mediated alterations in fatty acid and glucose metabolism should therefore lead to changes in ion homeostasis. Specifically, one would predict a decrease in ischaemic intracellular H+, Na+ and Ca2+ with TMZ. In a study published in this issue of Cardiovascular Research, El Banani et al. [31], examined the effects of TMZ on ionic changes during low-flow and zero-flow (total) ischaemia in isolated fatty-acid perfused rat hearts using 31P and 23Na nuclear magnetic resonance (NMR) spectroscopy. Like others, El Banani et al. [31] found perfusion with 1 μM TMZ to be cardioprotective, resulting in a 2-fold increase in post-ischaemic recovery of contractile function after low-flow ischaemia and a 5-fold increase after zero-flow ischaemia. However, they also found that the ionic effects of TMZ differed during low vs. zero-flow ischaemic protocols. In the low-flow ischaemic hearts, TMZ attenuated the ischaemic decrease in intracellular pH, consistent with the predicted attenuation of glycolysis subsequent to inhibition of β-oxidation by TMZ. However, the TMZ-treated low-flow ischaemic hearts did not exhibit an attenuation in ischaemic Na+, implying that TMZ-mediated protection was not via decreased Na+/H+ exchange activity during ischaemia. Also, contrary to the findings in low-flow hearts, there was no effect of TMZ treatment on ischaemic intracellular pH in zero-flow hearts, however, ischaemic Na+ was attenuated. The lack of a pH effect of TMZ in zero-flow hearts may be due to rapid glycolytic inhibition by a faster accumulation of metabolic products in zero vs. low-flow hearts, but in the absence of a pH effect, the role of glycolysis and Na+/H+ exchange activity in TMZ-mediated protection is questioned. During reperfusion, protective effects of TMZ were apparent in both low-flow and zero-flow hearts. In both groups, TMZ treatment resulted in greater increases in LVDP and faster decreases in both LVEDP and intracellular Na+. These findings may imply that TMZ-mediated alterations in glucose metabolism occur primarily during reperfusion, however, the absence of a higher pH during reperfusion in TMZ-treated vs. untreated low or zero-flow hearts again argues against decreased glycolysis as the mechanism of TMZ-mediated protection. Interestingly, positive effects of TMZ have been demonstrated previously in the absence of changes in glycolysis during ischaemia or reperfusion [32]. So how is the protective effect of TMZ mediated if not via changes in glycolysis and H+ production? An intriguing potential mechanism is implicated by the intracellular Na+ measurements of El Banani et al. [31]. In both low-flow and zero-flow hearts, intracellular Na+ levels during reperfusion were lower in TMZ-treated than in untreated hearts. In fact, in untreated zero-flow hearts, intracellular Na+ increased for the first 7 min of reperfusion, whilst in the TMZ-treated hearts, intracellular Na+ decreased immediately upon reflow (within the first 30-s measurement). Van Emous et al. [33] demonstrated a similar increase in intracellular Na+ during the first few minutes of reperfusion when the Na+/K+-ATPase was inhibited by ouabain. The possibility exists, therefore, that the Na+/K+-ATPase is inhibited in fatty acid-perfused hearts and this inhibition is relieved by TMZ. Interestingly, the Na+/K+-ATPase is reversibly inhibited by palmitoyl carnitine [34] and TMZ, at the same concentration used this study [31], attenuates the ischaemia-induced increase in levels of long chain acyl carnitines, such as palmitoyl carnitine [16]. In addition, unpublished observations from the same laboratory indicate a similar rapid decrease of intracellular Na+ upon reperfusion of fatty-acid perfused hearts treated with etomoxir, a carnitine palmitoyltransferase-1 inhibitor which also decreases long chain acyl carnitines [35]. As inhibition of Na+/K+-ATPase activity leads to increased intracellular Na+ and Ca2+ and exacerbation of injury [24,36,37], maintenance of post-ischaemic Na+/K+-ATPase activity by reduction of acyl carnitine levels by TMZ may explain the beneficial effect of TMZ in the experiments of El Banani et al. [31] and others [32], in the absence of evidence of decreased glycolysis. This hypothesis is consistent with the observation in the El Banani study [31] that LVEDP was lower during reperfusion in TMZ-treated vs. untreated hearts; LVEDP being proportional to intracellular Ca2+[38]. Maintenance of Na+/K+-ATPase activity during ischaemia in the TMZ-treated zero-flow hearts may also explain the observation of lower ischaemic intracellular Na+ levels in the absence of alterations in pH. Although the documented increase in glucose oxidation, as well as other reported mechanisms such as decreased myocardial oxygen consumption with lower fatty acid catabolism [39] and TMZ-mediated antioxidant and mitochondrial effects [9–11], may explain some of the protection afforded by TMZ, the findings of El Banani et al. [31] indicate a novel and intriguing potential protective mechanism. Future studies measuring Na+/K+-ATPase activity during ischaemia and reperfusion in TMZ-treated and untreated hearts, concurrent with measurements of glycolysis and glucose oxidation, will determine whether maintenance of Na+/K+-ATPase activity indeed contributes to the protective effects of the powerful therapeutic agent, trimetazidine.