Protective effects of Araloside C against myocardial ischaemia/reperfusion injury: potential involvement of heat shock protein 90.

The present study was designed to investigate whether Araloside C, one of the major triterpenoid compounds isolated from Aralia elata known to be cardioprotective, can improve heart function following ischaemia/reperfusion (I/R) injury and elucidate its underlying mechanisms. We observed that Araloside C concentration-dependently improved cardiac function and depressed oxidative stress induced by I/R. Similar protection was confirmed in isolated cardiomyocytes characterized by maintaining Ca2+ transients and cell shortening against I/R. Moreover, the potential targets of Araloside C were predicted using the DDI-CPI server and Discovery Studio software. Molecular docking analysis revealed that Araloside C could be stably docked into the ATP/ADP-binding domain of the heat shock protein 90 (Hsp90) protein via the formation of hydrogen bonds. The binding affinity of Hsp90 to Araloside C was detected using nanopore optical interferometry and yielded KD values of 29 µM. Araloside C also up-regulated the expression levels of Hsp90 and improved cell viability in hypoxia/reoxygenation-treated H9c2 cardiomyocytes, whereas the addition of 17-AAG, a pharmacologic inhibitor of Hsp90, attenuated Araloside C-induced cardioprotective effect. These findings reveal that Araloside C can efficiently attenuate myocardial I/R injury by reducing I/R-induced oxidative stress and [Ca2+ ]i overload, which was possibly related to its binding to the Hsp90 protein.

Salvianolic Acid A Protects H9c2 Cells from Arsenic Trioxide-Induced Injury via Inhibition of the MAPK Signaling Pathway.

BACKGROUND/AIMS: This study aimed to investigate whether Salvianolic acid A (Sal A) conferred cardiac protection against Arsenic trioxide (ATO)-induced cardiotoxicity in H9c2 cells by inhibiting MAPK pathways activation. METHODS: H9c2 cardiac cells were exposed to 10 µM ATO for 24 h to induce cytotoxicity. The cells were pretreated with Sal A for 4 h before exposure to ATO. Cell viability was determined utilizing the MTT assay. The percentage of apoptosis was measured by a FITC-Annexin V/PI apoptosis kit for flow cytometry. Mitochondrial membrane potential (∆Ψm) was detected by JC-1. The intracellular ROS levels were measured using an Image-iTTM LIVE Green Reactive Oxygen Species Detection Kit. The apoptosis-related proteins and the MAPK signaling pathways proteins expression were quantified by Western blotting. RESULTS: Sal A pretreatment increased cell viability, suppressed ATO-induced mitochondrial membrane depolarization, and significantly altered the apoptotic rate by enhancing endogenous antioxidative enzyme activity and ROS generation. Signal transduction studies indicated that Sal A suppressed the ATO-induced activation of the MAPK pathway. More importantly, JNK, ERK, and p38 inhibitors mimicked the cytoprotective activity of Sal A against ATO-induced injury in H9c2 cells by increasing cell viability, up-regulating Bcl-2 protein expression, and down-regulating both Bax and caspase-3 protein expression. CONCLUSION: Sal A decreases the ATO-induced apoptosis and necrosis of H9c2 cells, and the underlying mechanisms of this protective effect of Sal A may be connected with the MAPK pathways.

Salvianolic Acid A Ameliorates Arsenic Trioxide-Induced Cardiotoxicity Through Decreasing Cardiac Mitochondrial Injury and Promotes Its Anticancer Activity.

Arsenic trioxide (ATO) is used as a therapeutic agent in the treatment of acute promyelocytic leukemia (APL). The therapeutic use of arsenic is limited due to its severe cardiovascular side effects. The cardio-protective effect of salvianolic acid A (Sal A) against ATO cardiotoxicity has been reported. However, the distinct role of the mitochondria in the cardio-protection of Sal A is not understood. The aim of this study was to determine whether Sal A preconditioning protects against ATO-induced heart injury by maintaining cardiac mitochondrial function and biogenesis. For the in vivo study, BALB/c mice were treated with ATO and/or Sal A. For the in vitro study, we determined the effects of ATO and/or Sal A in H9c2 cardiomyocytes. Our results showed that ATO induced mitochondrial structural damage, abnormal mitochondrial permeability transition pore (mPTP) opening, overproduction of mitochondrial reactive oxygen species (ROS), and decreased the ATP content. Sal A pretreatment alleviated the ATO-induced mitochondrial structural and functional damage. In this study, ATO decreased the expression level of the peroxisome proliferator activator receptor gamma-coactivator 1 (PGC-1α) and disrupted the normal division and fusion of mitochondria. Sal A pretreatment improved the dynamic balance of the damaged mitochondrial biogenesis. Moreover, the combination treatment of Sal A and ATO significantly enhanced the ATO-induced cytotoxicity of SGC7901, HepaRG, K562 and HL60 cells in vitro. These results indicated that Sal A protects the heart from ATO-induced injury, which correlates with the modulation of mitochondrial function, and the maintenance of normal mitochondrial biogenesis.

Myricitrin Protects Cardiomyocytes from Hypoxia/Reoxygenation Injury: Involvement of Heat Shock Protein 90.

Modulation of oxidative stress is therapeutically effective in ischemia/reperfusion (I/R) injury. Myricitrin, a naturally occurring phenolic compound, is a potent antioxidant. However, little is known about its effect on I/R injury to cardiac myocytes. The present study was performed to investigate the potential protective effect of myricitrin against hypoxia/reoxygenation (H/R)-induced H9c2 cardiomyocyte injury and its underlying mechanisms. Myricitrin pretreatment improved cardiomyocyte viability, inhibited ROS generation, maintained the mitochondrial membrane potential, reduced apoptotic cardiomyocytes, decreased the caspase-3 activity, upregulated antiapoptotic proteins and downregulated proapoptotic proteins during H/R injury. Moreover, the potential targets of myricitrin was predicted using Discovery Studio software, and heat shock protein 90 (Hsp90) was identified as the main disease-related target. Further mechanistic investigation revealed that 17-AAG, a pharmacologic inhibitor of Hsp90, significantly blocked the myricitrin-induced cardioprotective effect demonstrated by increased apoptosis and ROS generation. These results suggested that myricitrin provides protection to H9c2 cardiomyocytes against H/R-induced oxidative stress and apoptosis, most likely via increased expression of Hsp90.

Calenduloside E Analogues Protecting H9c2 Cardiomyocytes Against H2O2-Induced Apoptosis: Design, Synthesis and Biological Evaluation.

Modulation of apoptosis is therapeutically effective in cardiomyocytes damage. Calenduloside E (CE), a naturally occurring triterpenoid saponin, is a potent anti-apoptotic agent. However, little is known about its synthetic analogues on the protective effects in apoptosis of cardiomyocytes. The present research was performed to investigate the potential protective effect of CE analogues against H2O2-induced apoptosis in H9c2 cardiomyocytes and the underlying mechanisms. Sixteen novel CE anologues have been designed, synthesized and biological evaluation. Among the 16 CE anologues, as well as the positive control CE tested, compound 5d was the most effective in improving cardiomyocytes viability. Pretreatment with anologue 5d inhibited ROS generation, maintained the mitochondrial membrane potential and reduced apoptotic cardiomyocytes. Moreover, exposure to H2O2 significantly increased the levels of Bax, cleaved caspase-3, and cleaved PARP, and decreased the level of Bcl-2, resulting in cell apoptosis. Pretreatment with anologue 5d (0.02-0.5 µg/mL) dose-dependently upregulated antiapoptotic proteins and downregulated proapoptotic proteins mentioned above during H2O2-induced apoptosis. These results suggested that CE analogues provide protection to H9c2 cardiomyocytes against H2O2-induced oxidative stress and apoptosis, most likely via anti-apoptotic mechanism, and provided the basis for the further optimization of the CE analogues.

Arsenic trioxide triggered calcium homeostasis imbalance and induced endoplasmic reticulum stress-mediated apoptosis in adult rat ventricular myocytes.

Arsenic trioxide (ATO) is a potent anticancer drug agent but its clinical use is often limited by severe cardiotoxicity. However, its exact mechanism remains poorly understood. In this study, we simultaneously explored the direct effect of ATO on cardiac contraction in adult rat ventricular myocytes and its effects on Ca2+ transient in real time by using an IonOptix MyoCam system. The results showed that ATO increased the amplitude of sarcomere shortening, the maximal velocity of relengthening and shortening (-dL/dtmax and +dL/dtmax), time-to-90% relengthening (TR90), and time-to-peak shortening (TPS), resulting in abnormal cardiomyocyte contraction. Meanwhile, ATO markedly increased the resting Ca2+ ratio, amplitude/resting calcium, the maximal velocity of Ca2+ shortening and relaxation (+d[Ca2+]/dtmax and -d[Ca2+]/dtmax), time-to-50% peak [Ca2+] i and the decay rate of [Ca2+] i transients, suggesting that ATO leads to intracellular imbalance of calcium homeostasis. ATO also inhibited sarcoplasmic reticulum Ca2+-ATPase 2a (SERCA2a) activity in a time-dependent manner and activated the endoplasmic reticulum (ER) stress reaction. These results revealed that ATO dramatically aggravates Ca2+ overload and promotes ER stress, eventually causing abnormal cardiomyocyte contraction in a dose-dependent and time-dependent manner.