Journal of Molecular and Cellular Cardiology 69 (2014) 83–84
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Editorial
Role of extracellular vesicles in remote ischemic preconditioning: ‘Good things come in small packages’?
Remote ischemic preconditioning (RIPC) is the phenomenon whereby brief episodes of ischemia–reperfusion applied in distant tissues or organs initiate a cardioprotective phenotype and render the myocardium resistant to a subsequent sustained ischemic insult [1]. The unique feature that discriminates RIPC from other conditioning paradigms (pre- and postconditioning) is communication: i.e., the transfer of a protective stimulus from the remote tissue to the heart [1–3]. In the two decades since the first description of RIPC [4,5], multiple and potentially synergetic mechanisms of inter-organ communication have been proposed, including the release of one or more circulating humoral factors (most notably, an elusive ‘small, b15 kDa hydrophobic molecule’ [6,7]), activation of autonomic reflex pathways [8], and/or posttranslational or transcriptional modification of circulating immune cells [1–3]. Most recently, this constellation of mechanisms has been expanded to include a new concept: Giricz and colleagues report novel evidence that extracellular vesicles participate in the RIPCinitiated communication of protective signals to the heart [9].
Extracellular vesicles as vectors for inter-organ communication Extracellular vesicles (members of the family of bioactive vesicles encompassing exosomes, microvesicles and microparticles) are nanosized lipid membrane-bound structures actively secreted or shed from multiple cell types. Initially regarded as debris, growing attention has focused on the theory that extracellular vesicles, released into the circulation, may serve as a form of intercellular and inter-organ communication by conveying their cargo of proteins, lipids, messenger and microRNAs to distant targets [10–13]. Although vesicle-mediated communication was first investigated in the fields of cancer and infectious disease [10], emerging data suggests that circulating exosomes and microparticles may play important roles in multiple facets of cardiovascular (patho)physiology, including attenuation of myocardial ischemia– reperfusion injury as well as repair, healing and angiogenesis postinfarction [11–15]. Moreover, in a recent editorial, Yellon and Davidson postulate that extracellular vesicles (and, in particular, exosomes), may “… contribute to the humoral transmission of the cardioprotective state induced by cardioprotective modalities, such as remote ischemic preconditioning” [13]. Giricz et al. provide seminal data in support for this hypothesis [9]. Using the isolated buffer-perfused rat heart model, the authors first demonstrate that repeated episodes of brief preconditioning ischemia was associated with the release of extracellular vesicles into the coronary effluent, as detected by electron microscopy, dynamic light scattering and immunoblotting for Hsp60, an established component of exosomes [15]. Second, and as expected [16], coronary effluent 0022-2828/$ – see front matter © 2014 Published by Elsevier Ltd. http://dx.doi.org/10.1016/j.yjmcc.2014.01.020
collected from preconditioned hearts (and confirmed to contain extracellular vesicles) had a significant infarct-sparing effect when administered to naïve acceptor hearts. Finally, effluent from nonischemic control hearts – and, most importantly, effluent from preconditioned hearts which had been depleted of extracellular vesicles – was ineffective in initiating a remote cardioprotective effect [9].
‘Communication via extracellular vesicles’: a paradigm shift in RIPC? The results obtained by Giricz and colleagues may, potentially, portend a paradigm shift in our understanding of RIPC. However, in order to establish ‘communication via extracellular vesicles’ as a tenable mechanism for the inter-organ transfer of a remote cardioprotective stimulus, multiple questions will require resolution. The first, critical issue is in vivo relevance: are the observations made by Giricz et al. [9] an anomaly of the isolated buffer-perfused heart model, possibly reflecting increased secretion of extracellular vesicles in response to the high coronary flow rates (and thus high shear forces [17]) that characterize this preparation? Evidence of alterations in circulating extracellular vesicles in response to a standard in vivo RIPC stimulus will be required, manifest as either an increase in plasma concentrations beyond the reportedly normal physiologic levels of 1 × 1010 per mL [13,18]), or, alternatively, a change in composition of the vesicles. Importantly, this must be coupled with evidence of cause-and-effect: definitive documentation must be provided that the in vivo infarctsparing effect of RIPC can be attributed to this augmented population of extracellular vesicles. Recent preliminary data published in abstract form have yielded progress in this regard: Vicencio and colleagues have shown an increased plasma concentration of extracellular vesicles (more specifically, exosomes) following repeated brief limb ischemia in rats and in human subjects, and observed that in vivo administration of purified exosomes reduced infarct size in the rat [19]. A second, related and daunting question is: what is the cellular source of the protective population of extracellular vesicles? Plausible candidates in the isolated buffer-perfused heart model include both cardiomyocytes and endothelial cells [15,17,20]. However, if extracellular vesicles are confirmed to play a mechanistic role in vivo, in the presence of platelets and other circulating blood elements known to serve as sources of exosomes and microparticles [21], the list of possibilities, and possible confounders, is substantially increased. Third, which specific subset(s) of bioactive vesicles are responsible for conveying the cardioprotective signal? Although Giricz et al. did not address this issue, exosomes (rather than microparticles) have, in general, been associated with protection [13]. Indeed, in the one study that has specifically interrogated the role of microvesicles in RIPC, brief repeated
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episodes of limb ischemia initiated a significant increase in plasma concentrations of platelet- and endothelium-derived microparticles, but these RIPC-induced microparticles were neither necessary nor sufficient to evoke the resultant infarct-sparing effect [22]. A final, conceptual question is: how does ‘communication via extracellular vesicles’ integrate with our current (and still-limited) knowledge of the mechanisms of RIPC? Aside from preliminary evidence of a link to classic ‘survival kinase’ signaling [1–3] – specifically, an increase in cardiac expression of phospho-ERK (phosphorylated extracellular signal regulated kinase) following administration of purified exosomes [19] – this fundamental issue has not been explored. In addition, no insight has been provided into the identity of the cargo, conveyed on or within the extracellular vesicles, that is responsible for initiating a cardioprotective phenotype. In fact, as acknowledged by the authors [9], it is possible that high concentrations of extracellular vesicles per se, irrespective of their composition and origin, may, for unknown reasons, protect the heart against ischemia–reperfusion injury. In summary, Giricz and colleagues [9] provide intriguing evidence that, for the communication phase of RIPC, the adage ‘good things come in small packages’ may hold true. Although many questions remain unanswered, these observations have the potential to propel RIPC in a new direction, and provide the impetus for future investigations aimed at elucidating the in vivo relevance and mechanistic details of extracellular vesicle-mediated inter-organ transfer of cardioprotective signals. Disclosures statement None.
[9]
[10]
[11] [12] [13] [14] [15]
[16]
[17]
[18]
[19]
[20] [21] [22]
dependent on the activity of vagal pre-ganglionic neurones. Cardiovasc Res 2012;95:487–94. Giricz Z, Varga ZV, Baranyai T, Sipos P, Paloczi K, Kittel A, et al. Cardioprotection by remote ischemic preconditioning of the rat heart is mediated by extracellular vesicles. J Mol Cell Cardiol 2014;68:75–8. Fleming A, Sampey G, Chung MC, Bailey C, van Hoek ML, Kashanchi F, et al. The carrying pigeons of the cell:exosomes and their role in infectious diseases caused by human pathogens. Pathog Dis 2014 [Epub 23 Jan]. Sahoo S, Losordo DW. Exosomes and cardiac repair after myocardial infarction. Circ Res 2014;114:333–44. Waldenstrom A, Ronquist G. Role of exosomes in myocardial remodeling. Circ Res 2014;114:315–24. Yellon DM, Davidson SM. Exosomes: nanoparticles involved in cardioprotection? Circ Res 2014;114:325–32. Lai RC, Arslan F, Lee MM, Sze NS, Choo A, Chen TS, et al. Exosome secreted by MSC reduces myocardial ischemia/reperfusion injury. Stem Cell Res 2010;4:214–22. Malik ZA, Kott KS, Poe AJ, Kuo T, Chen L, Ferrara KW, et al. Cardiac myocyte exosomes: stability, HSP60, and proteomics. Am J Physiol Heart Circ Physiol 2013;304:H954–65. Dickson EW, Lorbar M, Porcaro WA, Fenton RA, Reinhardt CP, Gysembergh A, et al. Rabbit heart can be “preconditioned” via transfer of coronary effluent. Am J Physiol 1999;277:H2451–7. Hergenreider E, Heydt S, Treguer K, Boettger T, Horrevoets AJ, Zeiher AM, et al. Atheroprotective communication between endothelial cells and smooth muscle cells through miRNAs. Nat Cell Biol 2012;14:249–56. Dragovic RA, Gardiner C, Brooks AS, Tannetta DS, Ferguson DJ, Hole P, et al. Sizing and phenotyping of cellular vesicles using nanoparticle tracking analysis. Nanomedicine 2011;7:780–8. Vicencio JM, Boi-Doku C, Das D, Sivaraman V, Kearney J, Hall AR, et al. Protecting the heart at a distance: exosomes for nano-sized cardioprotection. Heart 2014;100(Suppl. 1):A9. Zheng Y, Vicencio JM, Yellon DM, Davidson SM. Exosomes released from endothelial cells are cardioprotective. Heart 2014;100(Suppl. 1):A10. Hargett LA, Bauer NN. On the origin of microparticles: from “platelet dust” to mediators of intercellular communication. Pulm Circ 2013;3:329–40. Jeanneteau J, Hibert P, Martinez MC, Tual-Chalot S, Tamareille S, Furber A, et al. Microparticle release in remote ischemic conditioning mechanism. Am J Physiol Heart Circ Physiol 2012;303:H871–7.
References [1] Przyklenk K, Whittaker P. Remote ischemic preconditioning: current knowledge, unresolved questions, and future priorities. J Cardiovasc Pharmacol Ther 2011;16:255–9. [2] Przyklenk K. Reduction of myocardial infarct size with ischemic “conditioning”: physiologic and technical considerations. Anesth Analg 2013;117:891–901. [3] Przyklenk K, Whittaker P. Genesis of remote conditioning: action at a distance —‘hypotheses non fingo’? J Cardiovasc Med (Hagerstown) 2013;14:180–6. [4] Przyklenk K, Bauer B, Ovize M, Kloner RA, Whittaker P. Regional ischemic ‘preconditioning’ protects remote virgin myocardium from subsequent sustained coronary occlusion. Circulation 1993;87:893–9. [5] Whittaker P, Przyklenk K. Reduction of infarct size in vivo with ischemic preconditioning: mathematical evidence for protection via non-ischemic tissue. Basic Res Cardiol 1994;89:6–15. [6] Dickson EW, Blehar DJ, Carraway RE, Heard SO, Steinberg G, Przyklenk K. Naloxone blocks transferred preconditioning in isolated rabbit hearts. J Mol Cell Cardiol 2001;33:1751–6. [7] Shimizu M, Tropak M, Diaz RJ, Suto F, Surendra H, Kuzmin E, et al. Transient limb ischaemia remotely preconditions through a humoral mechanism acting directly on the myocardium: evidence suggesting cross-species protection. Clin Sci (Lond) 2009;117:191–200. [8] Mastitskaya S, Marina N, Gourine A, Gilbey MP, Spyer KM, Teschemacher AG, et al. Cardioprotection evoked by remote ischaemic preconditioning is critically
Karin Przyklenk Cardiovascular Research Institute, Wayne State University School of Medicine, Detroit, MI, USA Department of Physiology, Wayne State University School of Medicine, Detroit, MI, USA Department of Emergency Medicine, Wayne State University School of Medicine, Detroit, MI, USA Cardiovascular Research Institute, Wayne State University School of Medicine, Elliman Building, Room 1107, 421 E Canfield, Detroit, MI 48201, USA. Tel.: +1 313 577 9047; fax: +1 313 577 8615. E-mail address:
[email protected].
28 January 2014