Opposing Roles of S1P3 Receptors in Myocardial Function

Abstract

Sphingosine-1-phosphate (S1P) is a lysophospholipid mediator with diverse biological function mediated by S1P_1-5 receptors. Whereas S1P was shown to protect the heart against ischemia/reperfusion (I/R) injury, other studies highlighted its vasoconstrictor effects. We aimed to separate the beneficial and potentially deleterious cardiac effects of S1P during I/R and identify the signaling pathways involved. Wild type (WT), S1P₂-KO and S1P₃-KO Langendorff-perfused murine hearts were exposed to intravascular S1P, I/R, or both. S1P induced a 45% decrease of coronary flow (CF) in WT-hearts. The presence of S1P-chaperon albumin did not modify this effect. CF reduction diminished in S1P₃-KO but not in S1P₂-KO hearts, indicating that in our model S1P₃ mediates coronary vasoconstriction. In I/R experiments, S1P₃ deficiency had no influence on postischemic CF but diminished functional recovery and increased infarct size, indicating a cardioprotective effect of S1P₃. Preischemic S1P exposure resulted in a substantial reduction of postischemic CF and cardiac performance and increased the infarcted area. Although S1P₃ deficiency increased postischemic CF, this failed to improve cardiac performance. These results indicate a dual role of S1P₃ involving a direct protective action on the myocardium and a cardiosuppressive effect due to coronary vasoconstriction. In acute coronary syndrome when S1P may be released abundantly, intravascular and myocardial S1P production might have competing influences on myocardial function via activation of S1P₃ receptors.

Keywords: albumin; cardioprotection; coronary flow; ischemia/reperfusion; myocardial function; myocardial infarct; sphingosine-1-phosphate; vasoconstriction.

Figure 1

Dose-dependent effects of S1P on coronary flow of isolated murine hearts infused alone or in the presence of S1P-carrier albumin. In these experiments S1P was applied in a range of 10⁻⁹ to 10⁻⁵ M in cumulative doses without (S1P) or in the presence of its carrier, human serum albumin (S1P + albumin), and its effects on coronary flow were investigated. Albumin was present in a concentration twice that of S1P. ED₅₀ values were 1.17 × 10⁻⁶ M (S1P) and 1.85 × 10⁻⁷ M (S1P + albumin). Mean ± SEM; n = 9; 8. Non-linear regression analysis and comparison of Fits using GraphPad Prism 7.0.

Figure 2

Effects of S1P on coronary flow (CF) (A), left ventricular developed pressure (LVDevP) (B), +dLVP/dt_max (C) and −dLVP/dt_max (D) of isolated mouse hearts. S1P (10⁻⁶ M) or its vehicle was administered to the perfusate of isolated wild-type (WT) murine hearts for 5 min. The infusion was followed by a 20-min wash-out period. Administration of S1P resulted in a remarkable decrease in CF, which prevailed throughout the infusion and the wash-out period (p < 0.0001). CF reduction compromised left ventricular contractile performance as evidenced by a concomitant decrease in LVDevP, +dLVP/dt_max and −dLVP/dt_max (p < 0.0001). Mean ± SEM; n = 6, 9; #### p < 0.0001 vs. baseline (pre-infusion value), * p < 0.05 vs. vehicle; two-way repeated measurement ANOVA and Dunnett’s post hoc test.

Figure 3

Effects of S1P on coronary flow (CF) (A–C) and left ventricular developed pressure (LVDevP) (D–F) of hearts isolated from wild-type (WT) and S1P₂ knock-out (KO) mice. S1P (10⁻⁶ M) was administered to the perfusate of isolated WT and S1P₂-KO murine hearts for 5 min. The infusion was followed by a 20-min wash-out period. CF and LVDevP were monitored during the entire experiment (panels A and D). Maximal decrease in CF and LVDevP compared to preinfusion baseline are shown in panels B and E. Values of area over the curve (AOC) during S1P infusion are shown in panels C and F. In S1P₂-KO hearts S1P-induced CF and LVDevP reduction was similar to that observed in WT hearts. Mean ± SEM; n = 10, 8; #### p < 0.0001 vs. baseline (preinfusion value) in both groups, two-way repeated measurement ANOVA followed by Dunnett’s post hoc test.

Figure 4

Effects of S1P on coronary flow (CF) (A–C) and left ventricular developed pressure (LVDevP) (D–F) of hearts isolated from wild-type (WT) and S1P₃ knock-out (KO) mice. S1P (10⁻⁶ M) was administered to the perfusate of isolated WT and S1P₃-KO murine hearts for 5 min. The infusion was followed by a 20-min wash-out period. CF and LVDevP are shown in panels A and D. Maximal decrease in CF and LVDevP compared to preinfusion baseline are shown in panels B and E. Values of area over the curve (AOC) during S1P infusion are shown in panels C and F. In S1P₃-KO hearts, the S1P-induced CF and LVDevP reduction was significantly reduced. Mean ± SEM; n = 6, 8; #### p < 0.0001 vs. baseline (preinfusion value); * p < 0.05, ** p < 0.01 vs. WT; two-way repeated measurement ANOVA and Dunnett’s post hoc test (A,D) and unpaired t-test (B–F).

Figure 5

Postischemic coronary flow (CF) (A), left ventricular developed pressure (LVDevP) (B), +dLVP/dt_max (C), −dLVP/dt_max (D) and left ventricular diastolic pressure (LVDiastP) (E) in isolated WT and S1P₃ knock-out (KO) mouse hearts without (left panels: A1–E1) or with S1P administration (middle panels: A2–E2) for 5 min to the perfusate at 10⁻⁶ M before the induction of a 20-min ischemia followed by a 120-min reperfusion period. The right panels (A3–E3) demonstrate statistical comparison of the parameters captured at the end of the reperfusion period. Mean ± SEM; n = 6, 8, 7, 7; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, with two-way repeated measurement ANOVA and Dunnett’s post hoc test in the graphs and two-way ANOVA followed by Sidak’s post hoc test in the table insets.

Figure 6

Relative infarct size (A,B) and representative sections (C,D) from hearts subjected to ischemia/reperfusion without (A & C) or with (B & D) 10⁻⁶ M S1P infusion. Mean ± SEM; n = 6, 8, 7, 7; * p < 0.05, ** p < 0.01, **** p < 0.0001, with unpaired t-test (A,B) or two-way ANOVA and Sidak’s multiple comparison test (E).

Figure 7

Events in acute coronary syndrome related to S1P₃-mediated alterations of cardiac function.