We studied the role of two powerful molecular signalling mechanisms involved in the cardioprotective effect of sphingosine-1-phosphate (S1P), a major component of high density lipoprotein (HDL) against myocardial ischaemic-reperfusion injury, namely the RISK pathway (Akt/Erk), including its downstream target FOXO-1 and, the SAFE pathway (TNF/STAT-3).

Control hearts from wildtype, TNF deficient (TNF^−/−) or cardiomyocyte STAT-3 deficient (STAT-3^−/−) male mice were perfused on a Langendorff apparatus (35 min global ischaemia and 45 min reperfusion). S1P (10 nM) was given at the onset of reperfusion for the first 7 min, with/without STAT-3 or Akt inhibitors, AG490 and wortmannin (W), respectively.

S1P reduced myocardial infarct size in wildtype hearts (39.3 ± 4.4% in control vs 17.3 ± 3.1% in S1P-treated hearts; n ≥ 6; p < 0.05) but not in STAT-3^−/− or TNF^−/− mice (34.2 ± 4.3% in STAT-3^−/− and 34.1 ± 2.0% in TNF^−/− mice; n ≥ 6; p = ns vs. their respective control). Both STAT-3 and Akt inhibitors abolished the protective effects of S1P (33.7 ± 3.3% in S1P + AG490 and 36.6 ± 4.9% in S1P + W; n = 6; p = ns vs. their respective control). Increased nuclear levels of phosphorylated STAT-3 (pSTAT-3), Akt and FOXO-1 were observed at 15 min reperfusion in wildtype mice with Western Blot analysis (53% STAT-3, 47% Akt, 41% FOXO-1; p < 0.05 vs control) but not in STAT-3−/− mice or in wiltype hearts treated with the Akt inhibitor. Interestingly, an activation of pSTAT-3 was noticed in the mitochondria at 7 min but not 15 min of reperfusion.

In conclusion, S1P activates both the SAFE and RISK pathways, therefore suggesting a dual protective signalling in S1P-induced cardioprotection.