Propofol Induces Cardioprotection Against Ischemia-Reperfusion Injury via Suppression of Transient Receptor Potential Vanilloid 4 Channel

Figures

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  Figure 1. The protocol for myocardial I/R injury in isolated mice hearts.

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  Figure 2. The dose-dependent effect of propofol on myocardial I/R injury. The recovery of LVDP (A), +dP/dt max (B), -dP/dt max (C), and RPP (D) 60 minutes after reperfusion displayed as the percentage of respective preischemia values. (E) The amount of LDH release in coronary effluent at different times after reperfusion. (F) Representative images of TTC stained LV slices. Infarct size is quantified. Values are presented as mean ± SD, n = 6 for all groups, a one-way ANOVA followed by Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. INTRA. #p < 0.05, ###p < 0.001 vs. POP 50.

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  Figure 3. The effect of TRPV4 agonist GSK1016790A and TRPV4 antagonist HC-067047 on myocardial I/R injury. The recovery of LVDP (A), +dP/dt max (B), -dP/dt max (C), and RPP (D) 60 minutes after reperfusion displayed as the percentage of respective preischemia values. (E) The amount of LDH release in the coronary effluent at different times after reperfusion. (F) Representative images of TTC stained LV slices. Infarct size is quantified. Values are presented as mean ± SD, n = 6 for all groups, a one-way ANOVA followed by Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. DMSO.

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  Figure 4. The effect of propofol on TRPV4 agonist-induced myocardial I/R injury. Propofol or intralipid is given 5 minutes before applying TRPV4 agonist or DMSO. The recovery of LVDP (A), +dP/dt max (B), -dP/dt max (C), and RPP (D) 60 minutes after reperfusion displayed as the percentage of respective preischemia values. (E) The amount of LDH release in the coronary effluent at different times after reperfusion. (F) Representative images of TTC stained LV slices. Infarct size is quantified. Values are presented as mean ± SD, n = 6 for all groups, a one-way ANOVA followed by a Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. INTRA + DMSO. #p < 0.05, ###p < 0.001 vs. INTRA + GSK. &p < 0.05, &&&p < 0001 vs. POP50 + GSK.

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  Figure 5. The effect of the combination of propofol and TRPV4 antagonist on myocardial I/R injury. Propofol or intralipid is given 5 minutes before applying TRPV4 antagonist or DMSO. The recovery of LVDP (A), +dP/dt max (B), -dP/dt max (C), and RPP (D) 60 minutes after reperfusion displayed as the percentage of respective preischemia values. (E) The amount of LDH release in the coronary effluent at different times after reperfusion. (F) Representative images of TTC stained LV slices. Infarct size is quantified. Values are presented as mean ± SD, n = 6 for all groups, a one-way ANOVA followed by a Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. INTRA + DMSO.

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  Figure 6. The effect of the combination of propofol and TRPV4 agonist/TRPV4-siRNA on the [Ca2+]i and cell viability in H9C2 cells subjected to H/R. Pretreatment with propofol dose-dependently attenuates [Ca2+]i (A, n = 14 in N group, n = 16 in N + POP and H/R groups, n = 17 in H/R + POP 25, H/R + POP 50, and H/R + POP 100 groups) and increases cell viability (B, n = 6 for all groups). (C, n = 17 in H/R, H/R + GSK + POP,H/R + POP, H/R + HC, and H/R + HC + POP groups, n = 15 in H/R + GSK group) and (D, n = 6 for all groups) show the effects of propofol in the presence of TRPV4 agonist GSK1016790A. (E, n = 16 in H/R, H/R + siRNA, and H/R + siRNA + POP groups, n = 10 in H/R + Con siRNA and H/R + POP groups) and (F, n = 6 for all groups) show the protective effects of TRPV4-siRNA combined with propofol. Values are presented as mean ± SD, a one-way ANOVA followed by a Bonferroni test, ***p < 0.001 vs. N. #p < 0.05, ###p < 0.001 vs. H/R. &&p < 0.01, &&&p < 0.001 vs. H/R+GSK. ^^^p < 0.001 vs. HR+Con siRNA, NS. means no statistical significance.

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  Figure 7. The effect propofol on TRPV4 agonist-induced [Ca2+]i and the LDH release in ARVMs subjected to H/R. Pretreatment with 50 µM propofol attenuates [Ca2+]i (A) and LDH release (B), (C) and (D) show the effects of propofol in the presence of TRPV4 agonist GSK1016790A. Values are presented as mean ± SD, n = 6 for all groups, a one-way ANOVA followed by a Bonferroni test, ***p < 0.001 vs. N. ###p< 0.001 vs. H/R. &&&p < 0.001 vs. H/R+GSK.

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  Figure 8. Propofol dose-dependently suppresses the Ca2+ influx induced by TRPV4 agonists in HEK-293T cells transfected with hTRPV4. The above graphs show intracellular Ca2+ level change induced by 300 nM GSK1016790A (A) and 3 μM 4α-PDD (B). Arrows indicate when GSK1016790A and 4α-PDD are applied. Quantitative analyses of the relative changes in Ca2+ influx induced by GSK1016790A (C) and 4α-PDD (D) (∆F/F0). The dose-response curve fitted by the Hill function showing the inhibition of Ca2+ influx induced by GSK1016790A (E) and 4α-PDD (F). Values are presented as mean ± SD, GSK1016790A: n = 15 in the absence of POP, n = 12 in the presence of propofol at 12.5 µM, n = 15 in the presence of propofol at 25, 50 and 100 µM, 4α-PDD: n = 18 for all groups, a one-way ANOVA followed by a Bonferroni test, *p < 0.05, **p < 0.01, ***p < 0.001 vs. without propofol.

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  Figure 9. Propofol suppresses the Ca2+ influx induced by TRPV4 agonists in cardiomyocytes. Original time-lapse Ca2+ influx traces and quantification of GSK1016790A -induced [Ca2+]i response in H9C2 cells (A-C) and ARVMs (D-F). Arrows indicate when GSK1016790A (300 nM) and Ionomycin (1 μM) are applied. Cells are pretreated with 50 μM propofol for 30 minutes. Ionomycin is set as a positive control. Values are presented as mean ± SD, n = 6 for all groups, a two-tailed t test, ***p < 0.001 vs. DMSO, NS. means no statistical significance