Original articleCardiomyocyte-specific ablation of CD36 improves post-ischemic functional recovery
Introduction
Myocardial ischemia occurs as a consequence of insufficient blood flow and subsequent oxygen delivery to the myocardium, resulting in a variety of clinical conditions ranging from mild angina to myocardial infarction. Myocardial injury and contractile dysfunction are directly correlated with the length and severity of an ischemic event. Although many mechanisms contribute to ischemic injury (see [1] for review), there is clear evidence that contractile dysfunction during and after myocardial ischemia is mediated, at least in part, by the predominant type of energy substrate metabolized by the heart [2], [3].
In the healthy heart, mitochondrial oxidation of long-chain fatty acids (FAs) provides the majority of ATP needed for proper cardiac function [4]. Notwithstanding this, pre-clinical evidence has suggested that partial inhibition of myocardial FA oxidation (FAO) and a subsequent switch to greater glucose oxidation for ATP production can prevent ischemia/reperfusion (I/R) injury (see [2], [4] for reviews). Consistent with this finding, partial inhibitors of FAO in clinical use, such as trimetazidine [5], [6] and ranolazine [7], [8], [9], [10], have shown to improve and preserve cardiac function in patients suffering from ischemic heart disease and I/R injury [11], [12]. However, more recent reports suggest that mechanistically both drugs may act via alternative pathways and thus exert their beneficial effects independent of partial inhibition of FAO [12], [13], [14]. There is also a growing body of evidence suggesting that partial inhibition of myocardial FAO may actually contribute to cardiac dysfunction [15], [16] as a result of mismatch between FA uptake into the cardiomyocyte and subsequent utilization, leading to excessive and pathological lipid accumulation. It thus remains unclear as to whether partial inhibition of FAO is truly beneficial to the injured or diseased myocardium.
To address the potential limitations inherent in existing pharmacological therapies aimed at optimizing myocardial energetics, genetically modified mouse models designed to partially inhibit FA uptake and/or oxidation could be utilized. One such model is the whole body CD36 knockout (totalCD36KO) mouse [17]. CD36 is a transmembrane sarcolemmal protein involved in facilitating approximately 50% of cardiomyocyte FA uptake and is consequently responsible for controlling 40–60% of FAO rates in the working mouse heart [18], [19], [20]. Although totalCD36KO mice appeared to be suitable to address how partial inhibition of FA uptake and oxidation can influence ischemic injury, conflicting reports about the extent of myocardial I/R injury in these mice have emerged [21], [22]. The precise reason(s) for the different outcomes is not known, but may include perfusion conditions, mouse genetic background, type of FA used, age of mice, etc. Additionally, germline deletion of CD36 modifies a variety of metabolic pathways in multiple tissues [23] and subsequently whole body metabolism, which may make it difficult to determine the effects of cardiomyocyte-specific CD36-mediated alterations in metabolism on I/R injury. Moreover, compensatory alterations resulting from chronic CD36 ablation in other metabolic processes within the cardiomyocyte may have occurred during development that could influence I/R injury.
To overcome the challenges inherent to the totalCD36KO mouse, we have generated a cardiomyocyte-specific and tamoxifen-inducible CD36 KO (icCD36KO) mouse. Using short-term inducible cardiomyocyte-specific CD36 ablation we tested the hypotheses that: 1) icCD36KO mouse hearts have reduced FA uptake, utilization and storage and 2) this alteration in cardiac substrate utilization leads to improved post-ischemic functional recovery. These studies will allow us to determine if the concept of a combined strategy of limiting FA uptake and partially inhibiting FAO is a beneficial therapeutic approach to reducing ischemic injury [24], [25].
Section snippets
Mice
The investigation conforms to the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication No. 85–23, revised 1996). The University of Alberta adheres to the principles for biomedical research involving animals developed by the Council for International Organizations of Medical Sciences and complies with the Canadian Council on Animal Care guidelines. The University of Alberta Health Sciences Animal Welfare Committee and the
Inducible cardiomyocyte-specific CD36 ablation does not alter cardiac morphology
A Cre/loxP recombination system was employed to generate icCD36KO mice and mice were studied 4–6 weeks following tamoxifen administration. Four weeks post tamoxifen administration, CD36 protein expression was markedly reduced in both ventricles (Fig. 1C) and isolated cardiomyocytes (Fig. 1D) from icCD36KO mice compared to control mice, confirming ablation of CD36 in icCD36KO mice. Ablation of CD36 was restricted to the cardiomyocytes as CD36 protein expression in other tissues, such as skeletal
Discussion
We have previously shown that global loss of CD36 and the subsequent partial inhibition of cardiomyocyte FAO do not negatively impact the ability of the heart to recover from an ischemic insult [21]. More recently, a selective CD36 ligand was shown to reduce myocardial FA uptake in vivo and lessen post-ischemic myocardial injury [25]. Although the latter suggests that inhibition of CD36 in the heart may be of benefit during ischemia, the protective effects induced by the CD36 ligand could occur
Conclusions
In summary, the data presented herein show for the first time that cardiomyocyte-specific CD36 ablation can significantly reduce FA uptake, FAO, and TAG storage in healthy hearts as well as improve myocardial efficiency and functional recovery following an ischemic injury. Our findings are consistent with recent studies that have suggested that contractile dysfunction during and after myocardial ischemia can be reduced by stimulating glucose oxidation either directly or secondary to inhibition
Disclosure statement
None.
Acknowledgments
This work was supported by grants from the Canadian Institute of Health Research to J.R.B.D. J.R.B.D. is an Alberta Heritage Foundation for Medical Research Scholar. J.N. is supported by doctoral studentships from the Mazankowski Alberta Heart Institute, Alberta Innovates Health Solutions (AiHS), and the Canadian Diabetes Association (CDA). P.C.K. is supported by postdoctoral fellowships from the Heart and Stroke Foundation of Canada, the CDA, and the AiHS. T.P. is supported by a postdoctoral
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