GLP-1R agonist liraglutide activates cytoprotective pathways and improves outcomes after experimental myocardial infarction in mice

Abstract

Objective: Glucagon-like peptide-1 receptor (GLP-1R) agonists are used to treat type 2 diabetes, and transient GLP-1 administration improved cardiac function in humans after acute myocardial infarction (MI) and percutaneous revascularization. However, the consequences of GLP-1R activation before ischemic myocardial injury remain unclear.

Research design and methods: We assessed the pathophysiology and outcome of coronary artery occlusion in normal and diabetic mice pretreated with the GLP-1R agonist liraglutide.

Results: Male C57BL/6 mice were treated twice daily for 7 days with liraglutide or saline followed by induction of MI. Survival was significantly higher in liraglutide-treated mice. Liraglutide reduced cardiac rupture (12 of 60 versus 46 of 60; P = 0.0001) and infarct size (21 +/- 2% versus 29 +/- 3%, P = 0.02) and improved cardiac output (12.4 +/- 0.6 versus 9.7 +/- 0.6 ml/min; P = 0.002). Liraglutide also modulated the expression and activity of cardioprotective genes in the mouse heart, including Akt, GSK3beta, PPARbeta-delta, Nrf-2, and HO-1. The effects of liraglutide on survival were independent of weight loss. Moreover, liraglutide conferred cardioprotection and survival advantages over metformin, despite equivalent glycemic control, in diabetic mice with experimental MI. The cardioprotective effects of liraglutide remained detectable 4 days after cessation of therapy and may be partly direct, because liraglutide increased cyclic AMP formation and reduced the extent of caspase-3 activation in cardiomyocytes in a GLP-1R-dependent manner in vitro.

Conclusions: These findings demonstrate that GLP-1R activation engages prosurvival pathways in the normal and diabetic mouse heart, leading to improved outcomes and enhanced survival after MI in vivo.

Figures

FIG. 1.

Liraglutide pretreatment improves outcomes after MI in mice. Kaplan-Meier survival curves show survival after MI in nondiabetic mice (A): sham (n = 20), liraglutide-MI (75 μg/kg, n = 35 or 200 μg/kg, n = 60), PBS-MI (n = 60), and pair-fed mice (n = 25), P = 0.0001 for LIR 75 and 200 versus PBS; and diabetic mice (B): sham (n = 15, 5/treatment group), PBS-MI (n = 18), metformin-MI (n = 18), liraglutide-MI (75 μg/kg, n = 18), P = 0.04 for LIR 75 versus PBS. C: Frequency and timing of cardiac rupture in nondiabetic mice is shown as a percentage of total group. One-week pretreatment with liraglutide (200 or 75 μg/kg i.p. twice daily) had no significant effects on random blood glucose levels in adult nondiabetic mice (PBS: 7.6 ± 4.0 versus LIR 200: 6.3 ± 0.6 versus LIR 75: 6.1 ± 0.4 mmol/l; P = 0.72). Blood glucose levels in diabetic mice are shown in supplemental Table 1 (available in the online appendix).

FIG. 2.

Effects of liraglutide pretreatment on infarct size and heart weight. A: Representative photomicrographs of H-E–stained hearts 28 days post-MI depict decreased infarct size (arrows) in liraglutide- (n = 36) versus PBS-treated mice (n = 21) as confirmed by morphometric quantification of percent total LV circumference (*P = 0.025). B: HW/BW ratio was reduced in liraglutide- versus PBS-treated mice 28 days post-MI (n = 36 and 21, respectively; **P = 0.001). Data shown are means ± SE. (A high-quality digital representation of this figure is available in the online issue.)

FIG. 3.

Effects of liraglutide pretreatment on levels of cardiac genes and proteins. The expression of genes/proteins before left anterior descending ligation is analyzed in liraglutide-treated hearts. A: Representative Western blots for wild-type mice treated with LIR 200 as outlined in supplemental Fig. 1 (available in the online appendix). Corresponding densitometric quantification (n = 6/group) of fold changes in phosphorylation of prosurvival kinases Akt and GSK3β and in expression of PPAR-β/δ, Nrf2, and HO-1 are shown in the top five panels. The bottom panel depicts a representative agarose gel (n = 6/group) showing HO-1–specific mRNA levels by RT-PCR. Data shown are means ± SE; *P < 0.05. B–D: Representative Western blot analysis of cardiac prosurvival kinases in LIR 75–treated wild-type (B), LIR 75–treated Glp1r+/+ littermate controls (C), and LIR 75–treated Glp1r-/- (D) mice, respectively.

FIG. 4.

Effects of liraglutide pretreatment on cardioprotective signaling pathways after MI in mice. Liraglutide pretreatment for 7 days has persistent cardioprotective effects detectable 4 days post-MI as indicated by representative Western blots and corresponding densitometric quantification (n = 6/group) of fold changes in phosphorylation of Akt and GSK3β, cleavage of caspase-3, and expression of ANP and activity of MMP-9 by zymography in liraglutide- versus PBS-treated mice. Data shown are means ± SE; *P < 0.05.

FIG. 5.

Effects of liraglutide on recovery of LV function after I/R injury in isolated hearts. A: Effect of direct infusions of liraglutide- (0.3, 3, and 30 nmol/l, n = 5/group) or no treatment (vehicle, n = 21) either pre- or postischemia in isolated murine hearts subjected to experimental I/R ex vivo. B: Two additional groups of mice (n = 5/group) received i.p. injections of liraglutide b.i.d. for 1 or 7 days before ex vivo experiments. Data shown are means ± SE; *P < 0.01 compared with untreated controls.

FIG. 6.

Liraglutide induces cAMP formation and reduces caspase-3 activation in murine cardiomyocytes in vitro. A: Liraglutide (100 nmol/l) increases cAMP formation in cultured neonatal cardiomyocytes. The actions of liraglutide were abolished by the GLP-1R antagonist exendin9–39. ***P < 0.001. B: Liraglutide (L: 10–1,000 nmol/l) reduced TNF-α–induced activation of caspase-3 in a dose-dependent manner in cultured neonatal mouse cardiomyocytes. Cotreatment with exendin9–39 (Ex: 10 μmol/l) abolished the protective effects of liraglutide. Positive control represents treatment of cells with the potent apoptosis-inducing agent H2O2. Data shown are means ± SE; *P < 0.01 and **P < 0.001 versus cultures only treated with TNFα.