Abstract 150: FAK/Pyk2 Inhibitor Prevents Mitochondrial Ca2+ Overload and Cardiac Injury during Adrenergic Stimulation.
Introduction: Proline-rich tyrosine kinase 2 (Pyk2) and focal adhesion kinase (FAK) are abundantly expressed in cancer cells. In addition, these kinases are the potent therapeutic targets for cancer treatment and currently several selective FAK/Pyk2 inhibitors are in clinical trials. Recent study revealed that Pyk2 is also highly expressed in heart tissue and significantly activated during human heart failure. We have recently reported that α1-adrenoceptor (α1-AR) stimulation accelerates mitochondrial Ca2+ uptake through Pyk2-dependent phospholylation of mitochondrial Ca2+ uniporter (MCU). However, the roles of Pyk2 in cardiac mitochondrial physiology and pathophysiology have not been well established.
Hypothesis: Persistent adrenergic signaling activates cell death signaling via Pyk2-dependent MCU activation and mitochondrial Ca2+ overload.
Methods: Using H9C2 cardiac myoblasts, mitochondrial Ca2+ and reactive oxygen species (ROS) were measured using mitochondrial matrix-targeted Ca2+-sensitive inverse pericam and MitoSOX, respectively. Mitochondrial permeability transition pore (mPTP) activity was observed by measuring the amount of cytochrome c in cytosol by Western blotting or by monitoring the release of GFP-tagged mitochondrial protein, Smac-GFP using confocal microscopy.
Results: Pyk2 was not only expressed in cytosol, but also in cardiac mitochondria. α1-AR agonist phenylephrine activated mitochondrial Pyk2 and enhanced mitochondrial Ca2+ uptake via Pyk2-dependent MCU phosphorylation. In addition, persistent α1-AR stimulation increases ROS, activity of mPTP. These effects were abolished by co-expression of dominant-negative MCU or kinase-dead Pyk2, suggesting that Pyk2-dependent MCU activation followed by mitochondrial Ca2+ overload are critical for this mechanism. Moreover, pretreatment of a potent FAK/Pyk2 inhibitor PF-431396 also effectively inhibited α1-AR-mediated ROS generation and mPTP activation.
Conclusion: FAK/Pyk2 inhibitor prevents mitochondrial Ca2+ overload, oxidative stress and mitochondrial injury under persistent adrenergic stimulation. Thus, Pyk2 may become a novel potent therapeutic target for preventing cardiac cell injury and death during heart failure.
- © 2013 by American Heart Association, Inc.