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A small molecule activator of AKT does not reduce ischemic injury of the rat heart.

Moreira JB, Wohlwend M, Alves MN, Wisløff U, Bye A - J Transl Med (2015)

Bottom Line: Similarly, mitochondrial enzyme activity was not affected by SC79.Finally, SC79 failed to reduce infarct size or release of cardiac injury biomarkers at reperfusion.We conclude that selective AKT activation by the synthetic molecule SC79 does not protect the rat heart against ischemic injury, indicating that acute pharmacological activation of AKT is not sufficient for cardioprotection.

View Article: PubMed Central - PubMed

Affiliation: K.G. Jebsen Center of Exercise in Medicine, Department of Circulation and Medical Imaging, St. Olavs Hospital, Norwegian University of Science and Technology (NTNU), Prinsesse Kristinas gt. 3, 7006, Trondheim, Norway. jose.moreira@ntnu.no.

ABSTRACT

Background: Activation of protein kinase AKT is required for cardioprotection by ischemic preconditioning, and transgenic overexpression of AKT protects the heart against ischemia. However, it is unknown whether acute pharmacological activation of AKT alone, using a therapeutically relevant strategy, induces cardioprotection. In this study we provide the first evidence to clarify this question.

Methods: We used a recently described specific activator of AKT, the small molecule SC79, to treat rat hearts submitted to ischemia and reperfusion. Initially, isolated rat hearts were perfused with increasing doses of SC79 to verify the magnitude of AKT activation. Low and high doses were determined and used to treat hearts submitted to ischemia (35 minutes) and reperfusion (60 minutes), in a randomized and blinded design. AKT activation was verified by western immunobloting. Metabolic profile was determined by cardiac ATP content and mitochondrial enzyme activity, while cytosolic levels of cytochrome C and caspase-3 activity were used as markers of apoptosis. Ischemic injury was assessed by quantification of infarct size and cardiac release of creatine kinase and lactate dehydrogenase.

Results: SC79 activated cardiac AKT within 30 minutes in a dose-dependent fashion. ATP content was largely reduced by ischemia, but was not rescued by SC79. Similarly, mitochondrial enzyme activity was not affected by SC79. SC79 administered before ischemia or at reperfusion did not prevent cytosolic accumulation of cytochrome C and overactivation of caspase-3. Finally, SC79 failed to reduce infarct size or release of cardiac injury biomarkers at reperfusion.

Conclusion: We conclude that selective AKT activation by the synthetic molecule SC79 does not protect the rat heart against ischemic injury, indicating that acute pharmacological activation of AKT is not sufficient for cardioprotection.

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Related in: MedlinePlus

Ischemic injury.A: Abundance of creatine kinase in the perfusion buffer at reperfusion. B: Activity of lactate dehydrogenase (LDH) in the perfusion buffer at reperfusion. C: Infarct size as measured by histology after staining with Triphenyltetrazolium chloride. D: Representative histological sections of the left ventricle; Each dot within the bars represents one animal of the group. Note that IR-Low-Post shows 10 animals in panels “A” and “B” because one rat was detected as an outlier for these measurements. Other groups show data from all 11 animals. Data are shown as mean ± SE. &, p < 0.05 all other groups.
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Fig5: Ischemic injury.A: Abundance of creatine kinase in the perfusion buffer at reperfusion. B: Activity of lactate dehydrogenase (LDH) in the perfusion buffer at reperfusion. C: Infarct size as measured by histology after staining with Triphenyltetrazolium chloride. D: Representative histological sections of the left ventricle; Each dot within the bars represents one animal of the group. Note that IR-Low-Post shows 10 animals in panels “A” and “B” because one rat was detected as an outlier for these measurements. Other groups show data from all 11 animals. Data are shown as mean ± SE. &, p < 0.05 all other groups.

Mentions: Cardiac release of intracellular enzymes and infarct size are important predictors of prognosis and mortality after MI, and are used as primary outcome in clinical studies [5,6]. Abundance of CK and LDH was below detection limit in most of the hearts submitted to normoxic perfusion (Figures 5A and B). SC79 administered before ischemia or at reperfusion did not significantly reduce cardiac release of CK (Figure 5A) and LDH (Figure 5B). Similarly, SC79 did not promote survival of myocardial tissue after ischemia, as evidenced by the data on infarct size (Figure 5C). Figure 5D shows representative cross sections of the left ventricle after TTC staining.Figure 5


A small molecule activator of AKT does not reduce ischemic injury of the rat heart.

Moreira JB, Wohlwend M, Alves MN, Wisløff U, Bye A - J Transl Med (2015)

Ischemic injury.A: Abundance of creatine kinase in the perfusion buffer at reperfusion. B: Activity of lactate dehydrogenase (LDH) in the perfusion buffer at reperfusion. C: Infarct size as measured by histology after staining with Triphenyltetrazolium chloride. D: Representative histological sections of the left ventricle; Each dot within the bars represents one animal of the group. Note that IR-Low-Post shows 10 animals in panels “A” and “B” because one rat was detected as an outlier for these measurements. Other groups show data from all 11 animals. Data are shown as mean ± SE. &, p < 0.05 all other groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4352273&req=5

Fig5: Ischemic injury.A: Abundance of creatine kinase in the perfusion buffer at reperfusion. B: Activity of lactate dehydrogenase (LDH) in the perfusion buffer at reperfusion. C: Infarct size as measured by histology after staining with Triphenyltetrazolium chloride. D: Representative histological sections of the left ventricle; Each dot within the bars represents one animal of the group. Note that IR-Low-Post shows 10 animals in panels “A” and “B” because one rat was detected as an outlier for these measurements. Other groups show data from all 11 animals. Data are shown as mean ± SE. &, p < 0.05 all other groups.
Mentions: Cardiac release of intracellular enzymes and infarct size are important predictors of prognosis and mortality after MI, and are used as primary outcome in clinical studies [5,6]. Abundance of CK and LDH was below detection limit in most of the hearts submitted to normoxic perfusion (Figures 5A and B). SC79 administered before ischemia or at reperfusion did not significantly reduce cardiac release of CK (Figure 5A) and LDH (Figure 5B). Similarly, SC79 did not promote survival of myocardial tissue after ischemia, as evidenced by the data on infarct size (Figure 5C). Figure 5D shows representative cross sections of the left ventricle after TTC staining.Figure 5

Bottom Line: Similarly, mitochondrial enzyme activity was not affected by SC79.Finally, SC79 failed to reduce infarct size or release of cardiac injury biomarkers at reperfusion.We conclude that selective AKT activation by the synthetic molecule SC79 does not protect the rat heart against ischemic injury, indicating that acute pharmacological activation of AKT is not sufficient for cardioprotection.

View Article: PubMed Central - PubMed

Affiliation: K.G. Jebsen Center of Exercise in Medicine, Department of Circulation and Medical Imaging, St. Olavs Hospital, Norwegian University of Science and Technology (NTNU), Prinsesse Kristinas gt. 3, 7006, Trondheim, Norway. jose.moreira@ntnu.no.

ABSTRACT

Background: Activation of protein kinase AKT is required for cardioprotection by ischemic preconditioning, and transgenic overexpression of AKT protects the heart against ischemia. However, it is unknown whether acute pharmacological activation of AKT alone, using a therapeutically relevant strategy, induces cardioprotection. In this study we provide the first evidence to clarify this question.

Methods: We used a recently described specific activator of AKT, the small molecule SC79, to treat rat hearts submitted to ischemia and reperfusion. Initially, isolated rat hearts were perfused with increasing doses of SC79 to verify the magnitude of AKT activation. Low and high doses were determined and used to treat hearts submitted to ischemia (35 minutes) and reperfusion (60 minutes), in a randomized and blinded design. AKT activation was verified by western immunobloting. Metabolic profile was determined by cardiac ATP content and mitochondrial enzyme activity, while cytosolic levels of cytochrome C and caspase-3 activity were used as markers of apoptosis. Ischemic injury was assessed by quantification of infarct size and cardiac release of creatine kinase and lactate dehydrogenase.

Results: SC79 activated cardiac AKT within 30 minutes in a dose-dependent fashion. ATP content was largely reduced by ischemia, but was not rescued by SC79. Similarly, mitochondrial enzyme activity was not affected by SC79. SC79 administered before ischemia or at reperfusion did not prevent cytosolic accumulation of cytochrome C and overactivation of caspase-3. Finally, SC79 failed to reduce infarct size or release of cardiac injury biomarkers at reperfusion.

Conclusion: We conclude that selective AKT activation by the synthetic molecule SC79 does not protect the rat heart against ischemic injury, indicating that acute pharmacological activation of AKT is not sufficient for cardioprotection.

Show MeSH
Related in: MedlinePlus