The chromogranin A-derived peptides Vasostatin-1 and catestatin in the context of cardioprotection

To limit ischemia/reperfusion (I/R) injury in the heart, survival mechanisms can be triggered by short, non-lethal periods of ischemia/reperfusion, applied either before (preconditioning, PreC) or immediately after (postconditioning, PostC) the infarcting ischemia. PreC is a robust cardioprotective intervention, but has limited clinical efficacy. Conversely, PostC provides cardioprotection in clinically feasible settings, i.e., during coronary angioplasty. The existence of PostC cardioprotection confirms that about 50% of the final reperfused myocardial infarct may actually be due to myocardial reperfusion injury. Also peptides, such as bradykinin or opioids applied either before or after prolonged ischemia can trigger protective pathways (Penna et al., 2008, 2009). The Reperfusion Injury Salvage Kinase (RISK) pathway and the Survival Activating Factor Enhancement (SAFE) pathways are two apparently distinct signalling pathways which actually interact to convey the PostC stimulus from the cell surface to the mitochondria. It has been reported that pertussis toxin (PT) abolishes the cardioprotective effect of ischemic preconditioning in mammalian hearts. PT-sensitive, receptor-independent activation of G protein-dependent pathways have been proposed as mediator of the effects of vasostatin-I (VS-1) and catestatin (CTS) on vascular and cardiac responses. We reported that Vasostatin–I, triggers preconditioning-like protective effects via an adenosine/nitric oxide pathway (Cappello et al., 2006). Whether CTS may limit injury or contributes to ischemia/reperfusion injury is not clear. Since CTS activates some elements of the RISK pathway, including nitric oxide synthase, we propose that CTS may improve post-ischemic cardiac function and cell survival. To verify this hypothesis isolated ischemic/reperfused rat hearts were either preconditioned or postconditioned by ischemic episodes (I-PreC or I-PostC) or with exogenous Catestatin (CTS-PreC or CTS-PostC) performed either before ischemia or at the onset of reperfusion, respectively. In particular, CTS-PreC hearts underwent, 30-40 min stabilization, 20-min Catestatin (CTS, 75nM) infusion and 30-min global ischemia followed by 120-min reperfusion. In CTS-PostC, after stabilization and 30-min ischemia, hearts received 20-min CTS (75nM) infusion at the beginning of the 120-min reperfusion. Ischemic Prec (I-Prec) was obtained with 3 cycles of 3-min I/R before 30-min ischemia. Ischemic PostC (I-PostC) was obtained with 5 cycles of 10-sec I/R at the beginning of 120-min reperfusion. Perfusion pressure and left ventricular pressure (LVP) were monitored. Infarct size was evaluated with nitroblue-tetrazolium staining. In CTS-PostC group only, we assessed the phosphorylation level of key kinases of RISK by Western blot analysis. The CTS (5 nM) effects were also tested in simulated ischemia/reperfusion experiments on cardiomyocytes isolated from young-adult rats, evaluating cell survival with propidium-iodide labelling. We found that infarct size was 61±6% of risk area in hearts subjected to I/R only. Infarct size reduction by CTS-PostC was much stronger than that induced by CTS-PreC. Both CTS-PreC and CTS-PostC reduced diastolic LVP, an index of contracture. However only CTS-PostC dramatically improved post-ischemic recovery of developed LVP. In samples of these hearts the level of phospho-Aκt, -ERK ½, and -GSK3β resulted higher than the level of phosphorylation in I/R and Sham samples. In isolated cardiomyocytes CTS increased the cell viability rate after simulated ischemia/reperfusion. In conclusions, the cardio-protective influence exerted on post-ischemic mechanical performance and against I/R injury, point to CTS as a novel cardiac modulator, able to protect the heart via the phosphorylation/activation of several protective kinases. Catestatin applied in the reperfusion phase, as PostC agent seems more protective than Catestatin applied as PreC mimetic. These effects support a distinct activity with regard to the well evident preconditioning effect of Vasostatin.