Protective effects of Myrica rubra flavonoids against hypoxia/reoxygenation-induced cardiomyocyte injury via the regulation of the PI3K/Akt/GSK3ß pathway.

Myrica rubra is well known for its delicious taste and high nutritional value. The present study investigated the potential protective effects and mechanisms of M. rubra flavonoids (MRF) extract on isoproterenol (ISO)­induced myocardial injury in rats and hypoxia/reoxygenation (H/R) injury in H9c2 cardiomyocytes. An in vivo study revealed that MRF decreased serum cardiac enzyme levels, ameliorated pathological heart alterations and increased the antioxidant potential. The in vitro investigation demonstrated that MRF inhibited cell death, reactive oxygen species (ROS) accumulation, mitochondrial membrane depolarization, apoptosis rate and caspase­3 activation and enhanced the Bcl­2/Bax ratio during H/R injury. These effects were accompanied by the phosphorylation of protein kinase B (Akt) and glycogen synthase kinase (GSK)­3ß. Further mechanism studies demonstrated that LY294002, a specific inhibitor of phosphoinositide 3­kinase (PI3K), abolished the MRF­mediated cardioprotection against H/R­induced apoptosis and ROS overproduction. Collectively, these results suggested that MRF exerts cardioprotective effects by attenuating oxidative damage and cardiomyocyte apoptosis most likely via a PI3K/Akt/GSK3ß­dependent mechanism.

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.

Salvianolic acid B attenuates doxorubicin-induced ER stress by inhibiting TRPC3 and TRPC6 mediated Ca2+ overload in rat cardiomyocytes.

Doxorubicin (DOX)-induced cardiotoxicity is a clinically complex syndrome that leads to significant pain to cancer survivors. Endoplasmic reticulum (ER) stress has been suggested to be an important contributor to myocardium dysfunction during this phenomenon. Our previous study proved that Salvianolic acid B (Sal B) protected against doxorubicin induced cardiac dysfunction by inhibiting ER stress and cardiomyocyte apoptosis. However, the underlying molecular mechanism is not yet clearly. In this study, we investigated the protective effect and mechanisms of Sal B againest DOX-induced cardiac injury and ER stress in vivo and in vitro. After pretreatment with Sal B (0.25, 0.5, 1mg/kg i.v.) for 7 days, male SD rats were intraperitoneally injected with a single dose of DOX (3mg/kg) every 2 days for three injections. The cardioprotective effect of Sal B was observed 2 weeks after the first administration. Adult rat ventricular myocytes were isolated and treated with Sal B (20µg/ml) for 6h and then exposed in DOX (1µm) for 4h. The cardiomyocyte contractility and the level of intracellular Ca2+ were determined. Sal B ameliorated DOX-induced apoptosis damage in heart tissues. In vitro studies showed that DOX induced adult rat ventricular myocytes contractile dysfunction and intracellular Ca2+ handling derangement, disrupted mitochondrial membrane potential, raised the level of ER stress related proteins. However, Sal B pretreatment suppressed all of these adverse effects of DOX. The effects of Sal B were closely related to the inhibition of transient receptor potential canonical (TRPC) channels, as characterized by inhibiting the expression of TRPC 3 and TRPC6. These results indicate that Sal B protects against DOX-induced cardiac apoptosis and ER stress via TRPC3 and TRPC6 inhibition.

Myricitrin attenuates endothelial cell apoptosis to prevent atherosclerosis: An insight into PI3K/Akt activation and STAT3 signaling pathways.

Blood vessel endothelial dysfunction induced by oxidized low-density lipoprotein (ox-LDL) has been implicated in the pathogenesis of atherosclerosis and vasculopathy. The ox-LDL-elicited reactive oxygen species (ROS) release has been assumed to serve a critical function in endothelial damage. Myricitrin (from Myrica cerifera) is a natural antioxidant that has strong anti-oxidative, anti-inflammatory, and anti-nociceptive activities. However, the protective effect of myricitrin on ROS-induced endothelial cell injury and its related molecular mechanisms have never been investigated. This study demonstrates that myricitrin can inhibit ox-LDL-induced endothelial apoptosis and prevent plaque formation at an early stage in an atherosclerotic mouse model. The administration of myricitrin in vivo decreases the thickness of the vascular wall in the aortic arch of ApoE-/- mice. In vitro study shows that ox-LDL-induced human umbilical vein endothelial cell apoptosis can be reduced upon receiving myricitrin pre-treatment. Treatment with myricitrin significantly attenuated ox-LDL-induced endothelial cell apoptosis by inhibiting LOX-1 expression and by increasing the activation of the STAT3 and PI3K/Akt/eNOS signaling pathways. At the same time, our result demonstrates that myricitrin treatment optimizes the balance of pro/anti-apoptosis proteins, including Bax, Bad, XIAP, cIAP-2, and survivin. Our study suggests that myricitrin treatment can effectively protect cells from ox-LDL-induced endothelial cell apoptosis, which results in reduced atherosclerotic plaque formation. This result indicates that myricitrin can be used as a drug candidate for the treatment of cardiovascular diseases.

Inhibitory effects of myricitrin on oxidative stress-induced endothelial damage and early atherosclerosis in ApoE-/- mice.

Atherosclerosis (AS) is a state of heightened oxidative stress characterized by lipid and protein oxidation in vascular walls. Oxidative stress-induced vascular endothelial cell (VEC) injury is a major factor in the pathogenesis of AS. Myricitrin, a natural flavonoid isolated from the root bark of Myrica cerifera, was recently found to have a strong antioxidative effect. However, its use for treating cardiovascular diseases, especially AS is still unreported. Consequently, we evaluated the cytoprotective effect of myricitrin on AS by assessing oxidative stress-induced VEC damage. The in vivo study using an ApoE-/-mouse model of AS demonstrated that myricitrin treatment protects against VEC damage and inhibits early AS plaque formation. This effect is associated with the antioxidative effect of myricitrin, as observed in a hydrogen peroxide (H2O2)-induced rat model of artery endothelial injury and primary cultured human VECs. Myricitrin treatment also prevents and attenuates H2O2-induced endothelial injury. Further investigation of the cytoprotective effects of myricitrin demonstrated that myricitrin exerts its function by scavenging for reactive oxygen species, as well as reducing lipid peroxidation, blocking NO release, and maintaining mitochondrial transmembrane potential. Myricitrin treatment also significantly decreased H2O2-induced apoptosis in VECs, which was associated with significant inhibition of p53 gene expression, activation of caspase-3 and the MAPK signaling pathway, and alteration of the patterns of pro-apoptotic and anti-apoptotic gene expression. The resulting significantly increased bcl-2/bax ratio indicates that myricitrin may prevent the apoptosis induced by oxidative stress injury.

Oxidative stress suppression by luteolin-induced heme oxygenase-1 expression.

Luteolin, a flavonoid that exhibits antioxidative properties, exerts myocardial protection effects. However, the underlying molecular mechanisms are not yet fully understood. To investigate the effects of luteolin on myocardial injury protection and its possible mechanisms, a myocardial injury model was established with intragastric administration of 4 mg/kg isoproterenol (ISO) to male Sprague-Dawley rats (200-220 g) daily for 2 days. We found that pretreatment of luteolin (160, 80 and 40 mg/kg, i.g., respectively) daily for 15 days can prevent ISO-induced myocardial damage, including decrease of serum cardiac enzymes, improvement electrocardiography and heart vacuolation. Luteolin also improved the free radical scavenging and antioxidant potential, suggesting one possible mechanism of luteolin-induced cardio-protection is mediated by blocking the oxidative stress. To clarify the mechanisms, we performed the in vitro study by hydrogen peroxide (H(2)O(2))-induced cytotoxicty model in H9c2 cells. We found that luteolin pretreatment prevented apoptosis, increased the expression of heme oxygenase-1 (HO-1), and enhanced the binding of Nrf2 to the antioxidant response element, providing an adaptive survival response against H(2)O(2)-derived oxidative cytotoxicity. The addition of Znpp, a selective HO-1 competitive inhibitor, reduced the cytoprotective ability of luteolin, indicating the vital role of HO-1 on these effects. Luteolin also activated Akt and ERK, whereas the addition of LY294002 and U0126, the pharmacologic inhibitors of PI3K and ERK, attenuated luteolin-induced HO-1 expression and cytoprotective effect. Taken together, the above findings suggest that luteolin protects against myocardial injury and enhances cellular antioxidant defense capacity through the activation of Akt and ERK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction.