Original articleAdiponectin regulates SR Ca2 + cycling following ischemia/reperfusion via sphingosine 1-phosphate-CaMKII signaling in mice
Introduction
Cardiovascular disease affects more than 82 million people in the US and is the leading cause of death in the developed world [1]. Newly published data revealed a total coronary heart disease (CHD) prevalence of 7.0% (i.e., 16.3 million people) among adults aged ≥ 20 years in the US [1]. Early reperfusion of the ischemic myocardium is the most effective intervention for restoring cardiac function in the event of acute myocardial infarction (AMI); however, it is invariably accompanied by reperfusion-induced cellular damage and mechanical/electrical impairment of ventricular function [2], [3]. The causes of these deleterious effects are multifactorial, but it has been proposed that disturbances in Ca2 + homeostasis are central contributors to post-ischemic injury [4].
The sarcoplasmic reticulum (SR) is the main intracellular Ca2 + store in cardiac myocytes. It orchestrates excitation–contraction coupling through a rapid Ca2 + uptake (Ca2 +-ATPase, SERCA) and release transport system (ryanodine receptor, RyR) and is facilitated by a close juxtaposition with T-tubules and mitochondria [5]. Alterations in [Ca2 +]i handling, especially Ca2 + release from the SR, are known to be involved in the cytosolic Ca2 + overload and cardiac dysfunction observed during myocardial ischemia/reperfusion (I/R) and heart failure [3], [6], [7], [8]. Ca2 + uptake into the SR is considered to be depressed in myocardial I/R; however, the causes of this and whether enhanced Ca2 + uptake into the SR would benefit for ischemic myocytes remain controversial [4], [9]. SERCA2 activity in the SR is specifically regulated by phospholamban (PLB). In its dephosphorylated form, PLB exerts an inhibitory effect that can be reversed by its phosphorylation by protein kinase A (PKA) or Ca2 +/calmodulin-dependent protein kinase II (CaMKII). Because ischemia induces PLB dephosphorylation, regulating PLB phosphorylation has garnered attention as a potential therapeutic strategy for reducing reperfusion injury [10], [11].
Adiponectin (APN), a protein produced by adipocytes, protects the heart against I/R injury [12], [13]. However, although strong evidence supports the cardioprotective effects of APN during I/R, the underlying mechanisms remain incompletely understood. APN activates the intracellular AMP activated protein kinase (AMPK) pathway which acts as a metabolic switch for several intracellular systems, including glucose uptake, glucose transporter 4 generation, fatty acid oxidation, and mitochondrial genesis [14]. The precise intracellular molecules that mediate the AMPK-independent anti-apoptotic action of APN remain undefined [14], [15], [16]. Our previous study found that APN significantly reduced I/R-induced superoxide/NO overproduction and blocked inducible NOS (iNOS)/NOX-2 expression and apoptosis in wild-type as well as transgenic mice, which were cardiomyocyte-specific dominant negative for AMPK. One study reported that APN exerted its cardioprotection partially through AMPK-independent cyclooxygenase-2 activation and subsequent TNFα inhibition [13]. In cardiac myocytes, APN increased sphingosine 1-phosphate (S1P) and protected cells from apoptosis via either palmitate or C2-ceramide [17]. In addition, APN has been shown to induce cyclooxygenase-2 expression in cardiac myocytes via a mechanism that was AMPK independent but sphingosine kinase-1/S1P dependent [13], [18]. The potential role of S1P in the cytoprotective effects of APN remains largely unclear, however [17].
APN induces an intracellular Ca2 + influx in skeletal muscle that is necessary for the subsequent activation of Ca2 +/calmodulin dependent protein kinase kinase (CaMKKβ), AMPK, and mitochondrial biogenesis in myocytes [19], [20]. However, the effect of APN on Ca2 + activity, especially [Ca2 +]i transients in cardiomyocytes, has never been identified. Here, we identified a novel role for APN in cardiomyocytes and its underlying mechanism. Using a myocardial I/R (MI/R) protocol in vivo and individual cardiomyocytes in vitro, we demonstrated for the first time that APN is a natural modulator of SR SERCA2 activity. Mechanistic studies revealed that the SR SERCA2 activity enhanced by APN is dependent on the S1P/S1P receptor (S1PR)-CaMKII-PLB but independent of AMPK. Pharmacological inhibition of S1P/S1PR and SERCA2 siRNA suppressed the APN-mediated cardioprotective effects against I/R injury. These results support the role of S1P as a key mediator of APN cardioprotection and, more importantly, reveal novel cellular mechanisms by which APN and S1P protect the heart against I/R injury.
Section snippets
Animal and experimental protocols
Age-matched (12–14 weeks old) male wild-type (WT) and APN knockout (APN KO) mice were used for the experiments in this study. The generation, breeding, phenotypic characteristics, and genotyping of these mice have been previously described in detail [21]. The experimental protocols adhered to the National Institutes of Health Guidelines for the Use of Laboratory Animals and were approved by the Fourth Military Medical University Committee on Animal Care.
Mice were anesthetized with 2% isoflurane,
APN stimulated CaMKII and PLB (Thr17) phosphorylation and SR SERCA2 activity in the heart during I/R
During simulated I/R, cardiac SR SERCA2 activity is remarkably suppressed, which leads to reperfusion-induced hypercontracture, a hallmark of acute reperfusion injury [2]. CaMKII and PLB activities play a central role in regulating the activity of cardiac SR SERCA2 [27], [29], and site-specific Thr17 phosphorylation of PLB is mediated by the CaMKII pathway [30]. Despite some controversy [31], a few studies have demonstrated that CaMKII is inactivated in response to MI/R (30 min/30 min) [27], [28]
Discussion
APN is emerging as a protein hormone with insulin-sensitizing, anti-inflammatory and antiapoptotic functions, which are mostly mediated by AMPK. Several clinical observations have demonstrated that plasma APN concentrations are significantly reduced in patients with AMI and are inversely correlated with the risk of coronary artery disease [34], [35]. We have previously demonstrated that MI/R injury is markedly exacerbated in APN KO mice and that exogenous supplementation of APN is
Acknowledgments
This work was supported by the Program of National Science Fund for Distinguished Young Scholars of China (Grant No. 81225001), National Key Basic Research Program of China (973 Program, 2013CB531204), New Century Excellent Talents in University (Grant No. NCET-11-0870), National Science Funds of China (Grants Nos. 81070676 and 81170186), Program for Changjiang Scholars and Innovative Research Team in University (No. PCSIRT1053) and Major Science and Technology Project of China “Significant New
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2018, Biochimica et Biophysica Acta - Molecular Cell ResearchCitation Excerpt :In these cells, S1P coupled to HDL also induced an increase in intracellular calcium [137]. Furthermore, S1P induces calcium overload in neonatal rat cardiac myocytes through activation of S1P1 [138], whereas S1P evokes cardioprotection against reperfusion-induced injury in isolated neonatal mice hearts [139], and mediates adiponectin-evoked cardioprotection by a mechanism dependent on CaMKII-PLB and activation of SERCA2 [140]. An interesting finding was that pressure induced an increase in myogenic tone in rabbit posterior cerebral arteries through SphK1/S1P-dependent calcium signaling and increased Rho kinase-evoked phosphorylation of the 20 kDa myosin light chain [141].
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These three authors contributed equally to this work.