Cardiocyte Apoptosis Research Articles

Protective effects of ginsenoside F2 on isoproterenol-induced myocardial infarction by activating the Nrf2/HO-1 and PI3K/Akt signaling pathways

BackgroundGinsenoside F2 (GF2) serves as the principal intestinal metabolite resulting from the oral intake of Panax ginseng and Panax quinquefolius, exhibiting antioxidative, hypolipidemic, antitumor, and anti-inflammatory properties. Nevertheless, its effect on myocardial infarction (MI) is still unknown. PurposeThe purpose of this study is to investigate the protective effect and the underlying mechanisms of GF2 against isoproterenol (ISO)-induced MI. MethodsISO-induced H9c2 cardiomyocytes and MI rat models were utilized as in vitro and in vivo models to evaluate the impact of anti-MI of GF2. The underlying mechanisms were investigated using a variety of methodologies, including electrocardiography, Western blot analysis, histopathological examination, immunofluorescence, immunohistochemistry, and ELISA techniques. ResultsIn vivo experiments, our results indicated that GF2 significantly ameliorated ISO-induced electrocardiographic (ECG) abnormalities, myocardial fiber necrosis, rupture, fibrosis of myocardial tissues, and suppressed cardiac enzyme activities. Meanwhile, GF2 notably raised the activity of antioxidant enzymes like CAT, GSH, and SOD. Furthermore, it downregulated Keap1 expression level while upregulating NQO1, Nrf2, and HO-1 expression levels. Additionally, GF2 suppressed the expression of the cleaved caspase-3 and pro-apoptotic protein Bax while promoting the expression of anti-apoptotic proteins Bcl-2, p-PI3K, and p-Akt. TUNEL fluorescence results also demonstrated that GF2 effectively inhibited cardiomyocyte apoptosis. Furthermore, consistent with the results of animal experiments, GF2 considerably attenuated ROS generation, changed apoptosis and mitochondrial function, and reduced oxidative stress in ISO-induced H9c2 cardiomyocytes through activating Nrf2/HO-1 and PI3K/Akt signaling pathways. ConclusionTaken together, GF2 ameliorated MI by preventing cardiocyte apoptosis, oxidative stress, and mitochondrial dysfunction via modulating the Nrf2/HO-1 and PI3K/Akt signaling pathways, showing potential as a treatment strategy for treating MI.

Ferulic Acid Ameliorates Isoproterenol-Induced Heart Failure by Decreasing Oxidative Stress and Inhibiting Cardiocyte Apoptosis via Activating Nrf2 Signaling Pathway in Rats.

Ferulic acid (FA) has potential therapeutic effects in multiple diseases including cardiovascular diseases. However, the effect and molecular basis of FA in heart failure (HF) has not been thoroughly elucidated. Herein, we investigated the roles and mechanisms of FA in HF in isoproterenol (ISO)-induced HF rat model. Results found that FA ameliorated cardiac dysfunction, alleviated oxidative stress, reduced cell/myocardium injury-related enzyme plasma level, inhibited cardiocyte apoptosis in ISO-induced HF rat models. Moreover, FA reduced the co-localization of Keap1 and nuclear factor-E2-related factor 2 (Nrf2) in heart tissues of ISO-induced HF rats, and FA alleviated the inhibitory effects of ISO on expressions of p-Nrf2, heme oxygenase-1 (HO-1) and reduced nicotinamide adenine dinucleotide phosphate quinone dehydrogenase 1 (NQO1). Additionally, Nrf2 signaling pathway inhibitor ML385 showed adverse effects. FA weakened the effects of ML385 in ISO-induced HF rat models. Collectively, FA ameliorated HF by decreasing oxidative stress and inhibiting cardiocyte apoptosis via activating Nrf2 pathway in ISO-induced HF rats. Our data elucidated the underling molecular mechanism and provided a novel insight into the cardioprotective function of FA, thus suggested the therapeutic potential of FA in HF treatment.

Pyrrolidine Dithiocarbamate Attenuates Cardiocyte Apoptosis and Ameliorates Heart Failure Following Coronary Microembolization in Rats

Background:Nuclear factor-kB is highly activated in cardiovascular disorders. However, few articles have targeted at the role of nuclear factor-kB inhibitor in heart failure.Aims:To evaluate the effects of nuclear factor-kB inhibitor pyrrolidine dithiocarbamate on cardiocyte apoptosis and cardiac function in a rat heart failure model.Study Design:Animal experiment.Methods:A stable and reproducible rat heart failure model (n=64) was prepared by injecting homologous microthrombotic particles into the left ventricle of Sprague–Dawley rats while obstructing the ascending aorta to produce coronary microembolization. Rats with heart failure were randomized into untreated (HFu) and pyrrolidine dithiocarbamate-treated (HFp) groups; the latter received an intraperitoneal injection of pyrrolidine dithiocarbamate (100 mg/kg/day) 1 h prior to surgery as well as on postoperative days 1-7. The sham group comprised 32 Sprague–Dawley rats. Eight rats from each group were sacrificed on days 1, 3, 7, and 14 postoperatively. Masson’s trichrome staining was used to determine the micro-fibrotic area to indicate the severity of myocardial loss. Terminal transferase uridine triphosphate nick end labeling staining was used to detect apoptotic cardiomyocytes. Echocardiography and hemodynamics were performed to evaluate left ventricular function.Results:Rats with heart failure exhibited pathological changes evidenced by patchy myocardial fibrosis, remarkably elevated severity of myocardial loss, and persistently reduced left ventricular function. At the end of the study, compared with the HFu group, myocardial infarct size was reduced by 28% (p=0.001), cardiocyte apoptosis was suppressed (7.17%±1.47% vs 2.83%±0.75%, p<0.001), cardiac function parameters such as left ventricular ejection fraction (80%±4% vs 61%±6%), left ventricular + dP/dt max (4828±289 vs 2918±76 mmHg.s−1), left ventricular - dP/dt max (4398±269 vs 2481±365 mmHg.s−1), and left ventricular systolic pressure (126±13 vs 100±10 mmHg) were significantly increased, and left ventricular end-diastolic pressure was reduced (18±2 vs 13±1 mmHg) (p<0.001, for all) in the HFu group.Conclusion:Our rat model can adequately mimic heart failure via coronary vessel embolization. Moreover, pyrrolidine dithiocarbamate treatment can reduce cardiocyte apoptosis and improve cardiac function, which may be beneficial for patients with heart failure secondary to myocardial infarction.

Microarray profiling analysis identifies the mechanism of miR-200b-3p/mRNA-CD36 affecting diabetic cardiomyopathy via peroxisome proliferator activated receptor-γ signaling pathway.

Current study focused on the influence of miR-200b-3p on cardiocyte apoptosis of diabetic cardiomyopathy (DCM) by regulating CD36 and peroxisome proliferator-activated receptor γ (PPAR-γ) signaling pathway. Bioinformatic analysis was used to analyze differentially expressed microRNA (miRNAs), messenger RNAs (mRNAs) and activated pathways in DCM. And then quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to verify expression of miR-200b-3p and CD36 in DCM model rats and glucose treated H9c2 cell line. Luciferase reporter assay was used to verify the transcriptional regulation of agomiR-200b-3p and investigate the relationship between miR-200b-3p and CD36. Flow cytometry was performed to assess cardiocyte apoptosis in different interference conditions. Echocardiography was used to illustrate the ejection fraction rate and fraction shortening rate of DCM model rats. Next, hematoxylin-eosin (H&E) staining assay was carried out to reveal structures of cardiocyte tissues with transfection in different conditions. Masson trichrome staining was used to evaluate myocardial fibrosis. Western blot analysis was used to detect the expression levels of PPAR-γ signaling-related protein PPAR-γ and Bcl-2. miR-200b-3p was low-expressed while CD36 was overexpressed in DCM. AgomiR-200b-3p could inhibit the expression of CD36 to regulate cardiocyte apoptosis in DCM. CD36 activated PPAR-γ signaling pathway in DCM. Silencing CD36 or GW9662 treatment protect rat against DCM. miR-200b-3p targeted CD36 to regulate cardiocyte apoptosis of DCM by activating PPAR-γ signaling pathway.

Retracted: Upregulated microRNA-132 rescues cardiac fibrosis and restores cardiocyte proliferation in dilated cardiomyopathy through the phosphatase and tensin homolog-mediated PI3K/Akt signal transduction pathway.

Cardiac fibrosis is known to be present in dilated cardiomyopathy (DCM) and it predicts the occurrence of sudden death and congestive heart failure. The aim of our study is to investigate the expression of microRNA-132 (miR-132) and its effect on cardiocyte proliferation, apoptosis, and cardiac fibrosis by binding to phosphatase and tensin homolog (PTEN) through the phosphateidylinositol 3-kinase (PI3K)/protein kinase (Akt) signal transduction pathway in DCM rats. DCM rat models induced by doxorubicin were established and confirmed by an ultrasonic cardiogram. Epithelial cells were treated with inhibitors, activators, and small interfering RNAs to identify the mechanisms by which miR-132 controls cardiocyte activity and cardiac fibrosis. Angiotensin II (Ang II) and aldosterone (ALD) expressions were detected by an enzyme-linked immunosorbent assay. The relationship between PTEN and miR-132 was verified by a dual-luciferase reporter assay. Cell proliferation and apoptosis were tested by the MTT assay and flow cytometry. PTEN was determined to be the target gene of miR-132. Rat models of DCM exhibited a lower level of miR-132, PI3K, Akt, B-cell lymphoma 2, collagen I, and collagen III, but a higher level of PTEN, Bcl-2-associated X protein, and proliferating cell nuclear antigen as well as inflammatory response (Ang II and ALD), accompanied by declined cardiocyte proliferation and elevated apoptosis and cardiac fibrosis. Upregulated miR-132 or silenced PTEN activated the PI3K/Akt pathway, thus facilitating cardiocyte proliferation and repressing cardiocyte apoptosis and cardiac fibrosis, as well as inflammatory responses. Downregulated miR-132 reversed this tendency. These findings indicate that miR-132 activates the PI3K/Akt pathway by inhibiting PTEN expression, thus facilitating cardiocyte proliferation and inhibiting apoptosis and cardiac fibrosis in DCM rats.

Activation of novel estrogen receptor GPER results in inhibition of cardiocyte apoptosis and cardioprotection.

Several studies have recently demonstrated that G protein-coupled estrogen receptor (GPER) 30 directly binds to estrogen and mediates its action. The aim of the present study was to investigate the effects of GPER on cardiocyte apoptosis following ischemia/reperfusion injury (MIRI) in H9C2 myocardial cells. H9C2 cells were treated with a specific GPER agonist (G1), 17β-estradiol (E2) or the vehicle. The cells were subjected to 20 min of myocardial ischemia followed by 120 min of reperfusion. They were then randomly assigned to three experimental groups: Control, G1, E2. B-cell lymphoma 2 (Bcl-2) and Bcl-2 associated X (Bax) levels were measured, Hoechst 33258 staining was performed to assess apoptosis, and superoxide dismutase (SOD), tumor necrosis factor (TNF)-α and adenosine triphosphatase (ATPase) levels were determined. To test the specificity of G1, GPER-knockout cells were treated with G1 and analyzed as stated above. Compared with the vehicle-treated groups, G1 and E2-treated groups exhibited elevated Bcl-2 levels, decreased Bax levels and cell apoptosis, significantly increased SOD and ATP levels and decreased TNF-α levels following ischemia-reperfusion. However, G1 had no evident effects on the GPER-knockout cells. In conclusion, the present study suggested that GPER activation provided a cardioprotective effect following ischemia-reperfusion by inhibiting cardiocyte apoptosis.

Protective effect of proanthocyanidins against myocardial damage induced by mechanical trauma in rats

Objective To evaluate the effect of proanthocyanidins on cardiocyte apoptosis and cardiac dysfunction following mechanical trauma (MT) in rats and discuss its mechanism. Methods A total of 120 SD rats were allocated to normal group, trauma group, and trauma solvent group (treated with proanthocyanidins) and trauma vehicle group (treated with saline solution) according to the random number table, with 30 rats per group. Small animal quantitative wound instrument was used to prepare the MT model in rats. Cardiocyte apoptosis was measured by TUNEL staining. Changes in left ventricular diastolic pressure were recorded by left ventricular intubation via the right common carotid artery. Reactive oxygen species (ROS) produced by H9c2 cells before and after proanthocyanidins administration was detected with flow cytometry. Level of Ca2+ in cardiocytes marked by Fluo-4AM was determined with laser scanning confocal microscope. Ultrastructure changes in cardiocytes were observed under electronic microscope. Results After the induction of MT, apoptosis index of cardiocytes in trauma group, trauma solvent group and trauma vehicle group was (7.96±1.16)%, (2.44±0.70)%, and (7.76±1.14)% respectively, far higher than (0.64±0.59)% in normal group (P 0.05) but substantial reduction was detected in trauma solvent group (P<0.05). The + dp/dtmax of cardiac function in trauma and trauma vehicle groups was (3294.55±180.95)mmHg/s and (3337.80±220.11)mmHg/s respectively lowered from (4030.04±293.70) mmHg/s in normal group (P<0.01), and was (3926.72±364.95)mmHg/s in trauma solvent group lowered in contrast with trauma group (P<0.05). The -dp/dtmax in trauma and trauma vehicle groups was (2294.55±298.69)mmHg/s and (2291.20±348.72)mmHg/s respectively lowered significantly from (2830.04±334.63) mmHg/s in normal group (P<0.05), and was (2801.52±266.25)mmHg/s increased significantly in contrast with trauma group (P<0.05). Compared with normal group, trauma group showed chromatin margination and condensation as well as marked vacuolization in the epithelium from intramuscular capillaries. However, the pathological changes were improved after proanthocyanidins therapy. Conclusion Proanthocyanidins is effective in heart protection after the mechanical trauma by attenuating the pathological changes of myocardial ultrastructure and myocardial apoptosis and improving cardiac function. Key words: Proanthocyanidins; Heart injuries; Oxidative stress

MicroRNA-21 protects against ischemia-reperfusion and hypoxia-reperfusion-induced cardiocyte apoptosis via the phosphatase and tensin homolog/Akt-dependent mechanism

Myocardial tissue injury caused by ischemia and hypoxia is a major cause of fatal diseases, including coronary atherosclerosis resulting from myocardial infarction and stroke. A number of microRNAs have been demonstrated to function as protectors against ischemia-reperfusion (I/R) and/or hypoxia-reperfusion (H/R)-induced myocardial injury, including microRNA-21 (miR-21). However, the protective mechanism of miR-21 has not been fully elucidated. The present study demonstrated that miR-21 had an anti-apoptotic role in I/R-induced myocardial damage in vivo and in H/R-induced H9C2 cell death in vitro. Of note, the present study indicates that a common molecular mechanism is likely to exist in I/R- and H/R-induced cardiocyte apoptosis. During I/R and H/R, forced expression of miR-21 upregulated the Akt signaling activity via suppressing the expression of phosphatase and tensin homolog (PTEN) and the increased activity of Akt signaling further inhibited apoptosis partially by increasing the ratio of B-cell lymphoma 2(Bcl-2)/Bcl-2-associated X protein, which further suppressed the expression of caspase-3. In conclusion, to the best of our knowledge, it was shown for the first time that miR-21 had a protective role in I/R- and H/R-induced cardiocyte apoptosis via the PTEN/Akt-dependent mechanism. The present study indicates that miR-21 may be a promising agent for the treatment of I/R and H/R-induced myocardial injury.

Protective effects of fasudil hydrochloride post-conditioning on acute myocardial ischemia/reperfusion injury in rats

In recent years, the alleviation of acute myocardial ischemia and reperfusion injury (MI/RI) during myocardial reperfusion has presented a significant clinical challenge. This study was performed to investigate the effects of fasudil hydrochloride (FH) postconditioning on MI/RI and the underlying mechanism. Seventy-two rats were randomly divided into four groups: a Sham group, an ischemia/reperfusion (I/R) group, a fasudil hydrochloride (FH) group, and a fasudil hydrochloride+PI3K inhibitor (FH+I) group. Myocardial infarct size, cell apoptotic index (AI), and myocardial tissue expression of Rho-associated coiled-coil containing protein kinase 1 (ROCK1), Bcl-2, Bcl-2 associated X protein (Bax), caspase-3, Akt and phosphorylated Akt (P-Akt) were detected. All these parameters, except Akt expression, were higher in the I/R group than in the Sham group (p < 0.05). Compared to the I/R group, myocardial infarct size, AI, Bax and caspase-3 expression were significantly reduced in the FH group (p < 0.05), while Bcl-2 expression was increased (p < 0.05). However, the myocardial infarct size and AI of the FH+I group were similar to those of the I/R group (p > 0.05). Compared to the FH group, Bcl-2 expression was reduced in the FH+I group, while Bax and caspase-3 expression was increased (p < 0.05). Furthermore, P-Akt expression in the FH group was significantly higher than that of the I/R group (p < 0.05). FH post-conditioning alleviated MI/RI, with narrowing of the infarct size and decreased apoptosis of ischemic cardiocytes. The mechanism was associated with activation of the PI3K-Akt signaling pathway.

Role of damaged mitochondria in cardiocyte apoptosis in septic rats

Objective To investigate the role of damaged mitochondria in cardiac cell apoptosis in septic rats and the possible mechanism involved.Methods Seventy-two Sprague-Dawley rats were randomly (random number) divided into negative control group (n =18) and septic group (further divided into three groups as per rats sacrificed 6 h,12 h,and 24 h after endotoxin injection intra-peritoneally,n =18).The hearts of rats were taken.The changes of cardiac morphology were observed under light microscope and scanning electron microscope.Cell apoptosis in situ were examined by using terminal transferase-mediated dUTP nick end-labeling assay and nuclear factor-kappa B (NF-κB) activation in myocardium was detected by using Western blotting to estimate myocardial cell apoptosis.Mitochondrial lipid and protein oxidation were measured to assess oxidative stress,and mitochondrial superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities were determined to estimate antioxidant defense.Results Septic induced inflammatory cells infiltration,myocardium degeneration and effusion in a time-dependent manner.A remarkable expansion of capillaries could be observed in the hearts of infected rats at post-challenge of 24 h.Compared with sham-treated rats,the percentage of apoptosis increased in a time-dependent manner in the hearts of infected rats at 6 h,12 h,24 h of post-injection (P ＜0.05).The concentration of NF-κB p65 in the cytosol decreased gradually and increased in the nucleus during sepsis in a time-dependent manner (P ＜0.05),indicating that septic challenge provoked progressive activation of NF-κB.Mitochondrial cristae disappeared in 6 h of challenge,and significant mitochondrial cristae disappearance,vacuolization,and rupture of mitochondria membrane became markedly obvious 12 and 24 h later.Both SOD and GPx activities decreased,while mitochondrial lipid and protein oxidation increased in a time-dependent manner after 6-24 h of challenge (P ＜ 0.05).Conclusions Septic challenge induced myocardial apoptosis and mitochondrial damage.Further,damaged mitochondria might play an important role by means of alteration of defenses against reactive oxygen species in myocardial cell apoptosis during sepsis. Key words: Sepsis; Myocardial injury; Cell apoptosis; Mitochondrial; Nuclear factor-kappa B

Inhibitory effect of resveratol on ischemia reperfusion-induced cardiocyte apoptosis and its relationship with PI3K-Akt signaling pathway

To study the effect of resveratol on ischemia reperfusion-induced cardiocyte apoptosis and its relationship with PI3K-Akt signaling pathway. Fifty male SD rats were divided randomly into five groups: the control group (SH group), the ischemia reperfusion group (I/R group), the resveratol pretreatment group (Res group), the resveratol pretreatment + wortmannin group (Res +Wom group) and the ischemia reperfusion + wortmannin group (I/R + Wom group). The myocardial ischemia model was established by ligating left coronary artery for 45 min followed by 120 min reperfution, in order to observe the contents of NOS and NO. Cardiac myocyte apoptosis was determined by terminal deoxynueleotidyl transferase-mediated dUTP nick end labeling (TUNEL). Bcl-2 and Bax proteins were detected by immunohistochemistry. The t-Akt and p-Akt signaling protein expressions were determined by Western blotting analysis. Compared with the I/R groups and the Res + Wom group, the Res group showed significant increase in the expressions of NOS, NO, Bcl-2 protein and p-Akt and notable decrease in cardiocyte apoptosis and Bax/Bcl-2. The difference of above indicators showed a statistical significance (P<0.05). Furthermore, above changes can be blocked by wortmannin, a specific blocker of PI3K-Akt signaling pathway, indicating a statistical significance in their changes (P<0.05). Resveratol can inhibit the ischemia reperfusion-induced cardiocyte apoptosis, in which PI3K-Akt signaling pathway gets involved.

The Effect of 17-methoxyl-7-hydroxy-benzene-furanchalcone Isolated from Millettia pulchra on Myocardial Ischemia In Vitro and In Vivo

The effect of 17-methoxyl-7-hydroxy-benzene-furanchalcone isolated from the roots of Millettia pulchra (Benth.) Kurz var. Laxior (Dunn) Z. Wei on rat myocardial ischemia has been investigated. An in vitro cardiocyte apoptosis model and an in vivo myocardial ischemia model were used to elucidate the mechanism of 17-methoxyl-7-hydroxy-benzene-furanchalcone. In contrast to hydrogen peroxide (H2O2, 100 µmol/L), 17-methoxyl-7-hydroxy-benzene-furanchalcone in vitro (255 and 510 µmol/L) increased the quantity of total superoxide dismutase and the protein expression of B cell lymphoma/leukemia-2, while it inhibited cardiocyte apoptosis, the release of malondialdehyde and tumor necrosis factor α, and protein expression of nuclear factor κ Bp65 and Bcl-2-associated X protein. Furthermore, pretreatment with MHBFC in vivo (10 and 20 mg/kg) decreased heart rate, systolic pressure, diastolic pressure, average pressure, left ventricular systolic pressure, the largest upstroke velocity of the left ventricular pressure (+ dp/dtmax), total antioxidative capability, myoglobin isoenzyme of creatine kinase, and inducible nitric oxide synthase, while it increased endothelial nitric oxide synthase, ATPases, left ventricular diastolic pressure, left ventricular end-diastolic pressure, the largest descendent velocity of the left ventricular pressure (-dp/dtmax) and the interval from the beginning of left ventricular contraction to +dp/dtmax (t - dp/dtmax), all in a dose-dependent manner. Our present results suggest that 17-methoxyl-7-hydroxy-benzene-furanchalcone is an attractive antimyocardial ischemia agent mostly because of its negative heart rate and negative inotropic effects, the reduction in myocardial oxidative damage, and the modulating expression of genes associated with apoptosis, which improves diastolic function.

The protective effects of insulin on cardiocyte apoptosis challenged by burn serum

To investigate the protective effects of insulin on burn serum-challenged cardiocyte apoptosis and its mechanism. Burn-serum challenged cardiocytes were pretreated with insulin and inhibitors to pathway SB203580 and LY294002. The expression of cardiac myofilament proteins cleaved-caspase-3, Bax and phosphorylation nuclear factor-ΚB inhibitive factor α (p-IΚBα) were examined by Western blotting. The mRNA expression of tumor necrosis factor-α (TNF-α) was determined by real-time reverse transcription-polymerase chain reaction (RT-PCR). Apoptosis of cardiocyte was observed after Hoechst 33258 staining. Further blocking experiments were used to investigate the cytoprotective pathway of insulin. Insulin could significantly decrease the expression of cleaved-caspase-3 (2.22 ± 0.30 vs. 4.84 ± 0.74, P < 0.01), Bax (1.33 ± 0.35 vs. 3.74 ± 0.65, P < 0.01), p-IΚBα (1.43 ± 0.62 vs. 3.62 ± 0.74, P < 0.01), TNF-α (0.72 ± 0.27 vs. 2.02 ± 0.63, P < 0.01) and the cardiocyte apoptosis rate [(9.4 ± 3.4)% vs. (19.1 ± 5.6)%, P < 0.01] in cardiocytes challenged by burn serum. Further blocking experiments showed that LY294002, phosphatidylinositol-3-kinase (PI3K)/Akt activation inhibitor, could mitigate the protective effects of insulin. Meanwhile, SB203580, an inhibitor of p38 mitogen-activated protein kinase (p38MAPK) pathway, was able to inhibit cardiocyte injury challenged by burn serum, and it was as effective as insulin. For cardiocytes challenged by burn serum, insulin may decrease inflammatory cytokine expression and apoptosis via regulating PI3K/Akt and p38MAPK pathway.

Testosterone improves cardiac function and alters angiotensin II receptors in isoproterenol-induced heart failure

The renin-angiotensin-aldosterone system is known to play an important role in the pathophysiology and development of heart failure. Several studies have reported the benefits of testosterone in heart failure. However, the mechanisms of testosterone-induced effects on heart failure require further study. To determine the effects of castration and testosterone administration on cardiac function and angiotensin II receptor function in rats with isoproterenol-induced heart failure. Wistar rats were divided randomly into control and heart failure groups. The heart failure groups were further divided into the following groups: castration; castration+testosterone replacement; and sham castration. Echocardiography and haemodynamic measurements were used to evaluate cardiac function. Cardiocyte apoptosis and fibrosis were determined using terminal deoxyribonucleotide transferase-mediated dUTP nick-end labelling (TUNEL) staining and Masson's Trichrome staining, respectively. Angiotensin II receptor (AT1 and AT2) messenger ribonucleic acid (mRNA) expression levels were assayed using real-time reverse transcriptase-polymerase chain reactions, while Western immunoblotting was used to estimate Bcl-2 protein expression levels. Castration significantly increased cardiomyocyte apoptosis and fibrosis that was normally induced by isoproterenol (P<0.05). AT2 receptor mRNA expression in the castration group was increased and Bcl-2 protein expression was decreased compared with the castration+testosterone replacement group (P<0.05). These data suggest that androgen therapy could play an important role in pathophysiological changes in heart failure and have beneficial effects for its treatment.

Functional networks for Salvia miltiorrhiza and Panax notoginseng in combination explored with text mining and bioinformatical approach

Salvia miltiorrhiza (SM) and Panax notoginseng (PN) in combination (SMPN) have been widely used primarily in Traditional Chinese Medicine (TCM), for the treatment of coronary heart disease, and its pharmacological activity should be complicated because of its multiple components. Here, we combine text mining with bioinformatics to predict functional networks for the combination. 53 genes related with SMPN were found with text mining. Protein-protein interaction information for these genes from databases and Literature data was searched. Eight highly-connected regions were detected by IPCA algorithm to infer significant complexes or pathways in this network. Over-represented Gene Ontology categories of highly-connected regions by biological network gene ontology tool involved in small GTPase mediated signal transduction, apoptosis, regulation of immune effector process, phosphorylation about enzyme linked receptor protein signaling pathway, positive regulation of biological process. Integrate expression data from six microarray experiments about coronary heart disease into the SMPN network, and use the jActiveModules tool to find active subnetworks in differential expression conditions. The most relevant functions and pathways extracted from these subnetworks were related to proliferation and apoptosis of endothelial cell, apoptosis of arterial smooth muscle cell, apoptosis and regulation of immune system process within macrophages during foam cell formation, cardiocyte apoptosis. Analysis of the subnetwork composition indicated that there were in each subnetworks, and in the most subnetworks were dominant, the nodes came from SM network more than from PN network. It was suggested that, therapeutic efficacies of SMPN should be results of interaction between SM and PN in the multiple pathways and biological processes, and SM maybe play a principal role and PN serve as adjuvant one to assist the effects during the treatment of coronary heart disease. Key words: Panax notoginseng, salvia miltiorrhiza, herbal combination, bioinformatics, text mining, pharamcological activity.

Protective action of l-carnitine on cardiac mitochondrial function and structure against fatty acid stress

Cardiovascular risks are frequently accompanied by high serum fatty acid levels. Although recent studies have shown that fatty acids affect mitochondrial function and induce cell apoptosis, l-carnitine is essential for the uptake of fatty acids by mitochondria, and may attenuate the mitochondrial dysfunction and apoptosis of cardiocytes. This study aimed to elucidate the activity of l-carnitine in the prevention on fatty acid-induced mitochondrial membrane permeability transition and cytochrome c release using isolated cardiac mitochondria from rats. Palmitoyl-CoA-induced mitochondrial respiration that was observed with l-carnitine was inhibited with oligomycin. The palmitoyl-CoA-induced mitochondrial membrane depolarization and swelling were greatly inhibited by the presence of l-carnitine. In ultrastructural observations, terminally swollen and ruptured mitochondria with little or no distinguishable cristae structures were induced by treatment with palmitoyl-CoA. However, the severe morphological damage in cardiac mitochondria was dramatically inhibited by pretreatment with l-carnitine. Treatment with l-carnitine also attenuated 4-hydroxy- l-phenylglycine- and rotenone-induced mitochondrial swelling even when the l-carnitine could not protect against the decrease in oxygen consumption associated with these inhibitors. Furthermore, l-carnitine completely inhibited palmitoyl-CoA-induced cytochrome c release. We concluded that l-carnitine is essential for cardiac mitochondria to attenuate the membrane permeability transition, and to maintain the ultrastructure and membrane stabilization, in the presence of high fatty acid β-oxidation. Consequently, the cells may be protected against apoptosis by l-carnitine through inhibition of the fatty acid-induced cytochrome c release.

Effect of late reperfusion of infarct-related coronary artery on cardiocyte apoptosis in a dog acute myocardial infarction model

To investigate the effect of late reperfusion of infarct-related coronary artery on cardiomyocyte apoptosis and expressions of bcl-2/bax in the risk areas in a dog acute myocardial infarction (AMI) model, a total of 28 dogs were randomly divided into the three groups: SHAM group (n=8), AMI group (n=10) and late reperfusion (LR) group (n=10). Coronary artery was exposed after thoracotomy. The left anterior descending arteries were occluded for 12 h in the AMI group, and for 6 h in the LR group followed by reperfusion for 6 h, respectively. The cardiocyte apoptosis was measured by TUNEL assay. The protein and mRNA expressions of bcl-2 and bax were determined by western blot assay, immunohistochemistry and PCR, respectively. The apoptotic index in the LR group was significantly higher than in the sham group (P<0.01), but lower than the AMI group (P<0.05). The mRNA and protein expressions of in the LR group was markedly higher than in the SHAM group (P<0.01), but lower than in the AMI group (P<0.05). Late reperfusion of infarct-related artery can decrease cardiomyocyte apoptosis in the risk area of AMI which may be related to the reduced mRNA and protein expressions of bax. Key words: Acute myocardial infarction, later reperfusion, cardiomyocyte apoptosis, bax, bcl-2.

Cardioprotective Effect of Sulphonated Formononetin on Acute Myocardial Infarction in Rats

This study was designed to investigate the therapeutic effect of sodium formononetin-3'-sulphonate (Sul-F), a water-soluble derivate of formononetin, on acute myocardial infarction in rats. The results showed that treatment with Sul-F significantly prevented the elevation of ST-segment level, decreased the contents of creatine kinase-MB, lactate dehydrogenase, alanine aminotransferase and cardiac troponin T in serum and reduced the myocardium necrosis scores. The number of apoptosis cardiocytes is well accordance with the up-regulated expression of Bcl-2 and the down-regulated expression of Bax. Meanwhile, Sul-F significantly increased the cardiac mitochondrial ATP content, improved ATP synthase activity, decreased thiobarbituric acid-reactive substances content and attenuated the decrease in superoxide dismutase and glutathione peroxidase activities. These findings indicate that Sul-F has a protective potential against myocardial infarction injury. A possible mechanism for the protective effect is the elevated expression of endogenous antioxidant defence enzymes degraded lipid peroxidation products and improved energy metholism of cardiac mitochondrial, thus attenuating cardiocyte apoptosis.

The defects in development and apoptosis of cardiomyocytes in mice lacking the transcriptional factor Pax-8

BackgroudCardiac-specific deletion of ALK3 is lethal in mid-gestation with ventricular septum malformations (VSM). This study was designed to define the Pax-8's role in heart development and cardiomyocyte apoptosis. MethodsPathologic changes in the hearts of Pax-8 or ALK3 knockout and wild type control mice were determined by light and electron microscopy. Analysis of cardiomyocyte apoptosis was performed by TUNEL. The effect of Pax-8 gene deficiency on caspase-3 activity was examined after transfecting Pax-8 siRNA into cultured myoblast cell line. ResultsMice with ALK3 or Pax-8 gene knockout but not wild type control animals showed the development of VSM. Increased cardiomyocyte apoptosis was found in homozygotes. Echocardiography showed that Pax-8 homozygote mice developed malfunction of the heart. Furthermore, the caspase-3 activity was significantly higher in the cells treated with Pax-8 siRNA as compared to those treated with negative control siRNA in H9C2 (2-1) cell line. ConclusionsThe Pax-8 gene may play a crucial role in heart development and regulating cardiocyte apoptosis. Knockout of Pax-8 may exert a similar effect on myocardial morphology and apoptosis as those seen in ALK3 knockouts. Furthermore, the ventricular septum malformations could be partially attributed to accelerated cardiomyocyte apoptosis.

Is overall blockade superior to selective blockade of adrenergic receptor subtypes in suppressing left ventricular remodeling in spontaneously hypertensive rats?

To test the hypothesis that nonselective blockade of adrenergic receptor (AR) subtypes is superior to selective blockade of AR subtypes in suppressing left ventricular (LV) remodeling induced by hypertension. Sixty-four spontaneously hypertensive rats (SHR) were randomly divided into four groups: bisoprolol-treated, propranolol-treated, carvedilol-treated and no treatment groups (n=16, each). Sixteen Wistar-Kyoto (WKY) rats served as a control group. Echocardiography and cardiac catheterization were carried out to record the mitral flow velocity ratio of E wave to A wave (E/A), LV mass index (LVMI), maximal rising (dp/dt(max)) and falling (-dp/dt(max)) rate of the LV pressure and LV relaxation time constant (τ). The mRNA and protein expression levels of AR, protein kinase(PK) and G-protein subtypes, intracellular free calcium (Ca) concentration and cardiocyte apoptoisis rate were determined. Three drug-treated groups showed higher velocity ratio of E wave to A wave (E/A) and -dp/dt(max) and lower systolic blood pressure (SBP), LVMI, τ, apoptosis rate and intracellular free Ca(2+) concentration than the no treatment group. The mRNA expression levels of AR-α(1B) in the carvedilol group were significantly lower than the other two drug-treated groups. The mRNA expression levels of AR-β(1), AR-β(2) and Gsα were significantly higher in the three drug-treated groups than in the no treatment group, with the expression levels of AR-β(2) being the highest in the carvedilol-treated group. The protein expression levels of PKA and PKC subtype α and δ were lower in the three drug-treated groups than in the no treatment group. Overall blockade of AR subtypes is not superior to selective blockade of AR subtypes in suppressing LV remodeling in SHR. Although carvedilol is the most effective in attenuating cardiocyte apoptosis, normalizing AR-α(1B) and Gsα expression and increasing AR-β(2) expression.