Antihypertrophic Effects of Small Molecules that Maintain Mitochondrial ATP Levels Under Hypoxia.

Since impaired mitochondrial ATP production in cardiomyocytes is thought to lead to heart failure, a drug that protects mitochondria and improves ATP production under disease conditions would be an attractive treatment option. In this study, we identified small-molecule drugs, including the anti-parasitic agent, ivermectin, that maintain mitochondrial ATP levels under hypoxia in cardiomyocytes. Mechanistically, transcriptomic analysis and gene silencing experiments revealed that ivermectin increased mitochondrial ATP production by inducing Cox6a2, a subunit of the mitochondrial respiratory chain. Furthermore, ivermectin inhibited the hypertrophic response of human induced pluripotent stem cell-derived cardiomyocytes. Pharmacological inhibition of importin ß, one of the targets of ivermectin, exhibited protection against mitochondrial ATP decline and cardiomyocyte hypertrophy. These findings indicate that maintaining mitochondrial ATP under hypoxia may prevent hypertrophy and improve cardiac function, providing therapeutic options for mitochondrial dysfunction.

A pyridone derivative activates SERCA2a by attenuating the inhibitory effect of phospholamban.

The cardiac sarco/endoplasmic reticulum Ca2+-dependent ATPase 2a (SERCA2a) plays a central role in Ca2+ handling within cardiomyocytes and is negatively regulated by phospholamban (PLN), a sarcoplasmic reticulum (SR) membrane protein. The activation of SERCA2a, which has been reported to improve cardiac dysfunction in heart failure, is a potential therapeutic approach for heart failure. Therefore, we developed a novel small molecule, compound A and characterized it both in vitro and in vivo. Compound A activated the Ca2+-dependent ATPase activity of cardiac SR vesicles but not that of skeletal muscle SR vesicles that lack PLN. The surface plasmon resonance assay revealed a direct interaction between compound A and PLN, suggesting that the binding of compound A to PLN attenuates its inhibition of SERCA2a, resulting in SERCA2a activation. This was substantiated by inhibition of the compound A-mediated increase in Ca2+ levels within the SR of HL-1 cells by thapsigargin, a SERCA inhibitor. Compound A also increased the Ca2+ transients and contraction and relaxation of isolated adult rat cardiomyocytes. In isolated perfused rat hearts, the compound A enhanced systolic and diastolic functions. Further, an infusion of compound A (30mg/kg, i.v. bolus followed by 2mg/kg/min, i.v. infusion) significantly enhanced the diastolic function in anesthetized normal rats. These results indicate that compound A is a novel SERCA2a activator, which attenuates PLN inhibition and enhances the systolic and diastolic functions of the heart in vitro and in vivo. Therefore, compound A might be a novel therapeutic lead for heart failure.

Effects of 4(1H)-quinolinone derivative, a novel non-nucleotide allosteric purinergic P2Y 2 agonist, on cardiomyocytes in neonatal rats.

Purinergic P2Y 2 receptors, G-protein coupled receptors that primarily couple with Gαq/11-proteins, are activated equipotently by adenosine-5'-triphosphate (ATP) and uridine-5'-triphosphate. Evidence suggests that P2Y 2 agonists make potential drug candidates for the treatment of cardiovascular diseases. However, selective non-nucleotide, small-molecule P2Y 2 agonists have yet to be developed. In this report, we discuss Compound 89, a novel non-nucleotide allosteric P2Y 2 agonist that was active in signal transduction and gene induction, and in our in vitro cardiac hypertrophy model. Compound 89 exhibited selective P2Y 2 agonistic activity and potentiated responses to the endogenous agonist ATP, while exhibiting no agonistic activities for four other Gαq/11-coupled human P2Y (hP2Y) receptors and one representative Gαi/o-coupled hP2Y12 receptor. Its P2Y 2 agonistic effect on mouse P2Y 2 receptors suggested non-species-specific activity. Compound 89 acted as a pure positive allosteric modulator in a Ca2+ mobilization assay of neonatal rat cardiomyocytes; it potentiated ATP-induced expression of genes in the nuclear receptor 4A family (negative regulators of hypertrophic stimuli in cardiomyocytes). Additionally, Compound 89 attenuated isoproterenol-induced cardiac hypertrophy, presumably through dose-dependent interaction with pericellular ATP. These results indicate that Compound 89 is potentially efficacious against cardiomyocytes and therefore a good proof-of-concept tool for elucidating the therapeutic potential of P2Y2 activation in various cardiovascular diseases.

Plasminogen activator inhibitor-1 deficiency retards diabetic nephropathy.

BACKGROUND: Plasminogen activator inhibitor-1 (PAI-1) is increased in kidneys of humans and animals with diabetic nephropathy and is associated with extracellular matrix (ECM) accumulation. PAI-1 may promote ECM buildup by preventing plasmin and matrix metalloproteinase (MMP) activation. However, the importance and mechanism of PAI-1 action in the pathogenesis of diabetic nephropathy is unknown. METHODS: We investigated the effect of streptozotocin (STZ)-induced diabetes in wild-type (PAI-1(+/+)) mice and mice null for PAI-1 (PAI-1(-/-)). After 1 month of diabetes, animals were placed in metabolic cages for 24-hour urine collection. Total RNA was isolated from kidney cortex for reverse transcription-polymerase chain reaction (RT-PCR) and Northern blot analysis, and Western blots were quantitated from cortical protein. Primary mesangial cells were grown from Sprague-Dawley rats and used in signal transduction studies. RESULTS: Urinary albumin excretion (UAE) in diabetic PAI-1(+/+) mice increased >threefold, but remained unchanged in PAI-1(-/-) mice. Transforming growth factor-beta (TGF-beta) and fibronectin message and protein levels were lower in diabetic PAI-1(-/-) vs. PAI-1(+/+) mice, suggesting that PAI-1 deficiency impaired TGF-beta expression despite diabetes. Indeed, recombinant PAI-1 directly stimulated TGF-beta message and protein via mitogen-activated protein kinase (MAPK) signal transduction in cultured mesangial cells. Urokinase plasminogen activator (uPA) inhibited this PAI-1 action in a dose-dependent manner. The inhibitory effect of antibody to uPA receptor (uPAR) on PAI-1-induced TGF-beta function suggested that uPAR mediated the cellular effect of PAI-1. CONCLUSION: PAI-1 can regulate TGF-beta expression by binding to uPAR and activating the extracellular-regulated signal kinase (ERK)/MAPK pathway. Therefore, PAI-1 contributes to diabetic nephropathy by regulating TGF-beta and renal ECM production and may be a therapeutic target in diabetic nephropathy.