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Parkinson's disease (PD) is a common neurodegenerative disorder associated with striatal dopaminergic neuronal loss in the Substantia nigra. Oxidative stress plays a significant role in several neurodegenerative diseases. Paraquat (PQ) is considered a potential neurotoxin that affects the brain leading to the death of dopaminergic neurons mimicking the PD phenotype. Various scientific reports have proven that cryptotanshinone possesses antioxidant and anti-inflammatory properties. We hypothesized that cryptotanshinone could extend its neuroprotective activity by exerting antioxidant effects. This study was designed to evaluate the effects of cryptotanshinone in both cellular and animal models of PQ-induced PD. Annexin V-PI double staining and immunoblotting were used to detect apoptosis and oxidative stress proteins, respectively. Reactive oxygen species kits were used to evaluate oxidative stress in cells. For in vivo studies, 18 B6 mice were divided into three groups. The rotarod data revealed the motor function and immunostaining showed the survival of TH+ neurons in SNpc region. Our study showed that cryptotanshinone attenuated paraquat-induced oxidative stress by upregulating anti-oxidant markers in vitro, and restored behavioral deficits and survival of dopaminergic neurons in vivo, demonstrating its therapeutic potential.

Pathological cardiac hypertrophy is associated with many diseases including hypertension. Recent studies have identified important roles for microRNAs (miRNAs) in many cardiac pathophysiological processes, including the regulation of cardiomyocyte hypertrophy. However, the role of miR-145-5p in the cardiac setting is still unclear. In this study, H9C2 cells were overexpressed with microRNA-145-5p, and then treated with Ang-II for 24 h, to study the effect of miR-145-5p on Ang-II-induced myocardial hypertrophy in vitro. Results showed that Ang-II treatment down-regulated miR-145-5p expression were revered after miR-145-5p overexpression. Based on results of bioinformatics algorithms, paxillin was predicted as a candidate target gene of miR-145-5p, luciferase activity assay revealed that the luciferase activity of cells was substantial downregulated the following co-transfection with wild paxillin 3'UTR and miR-145-5p compared to that in scramble control, while the inhibitory effect of miR-145-5p was abolished after transfection of mutant paxillin 3'UTR. Additionally, overexpression of miR-145-5p markedly inhibited activation of Rac-1/ JNK /c-jun/ NFATc3 and ANP expression and induced SIRT1 expression in Ang-II treated H9c2 cells. Jointly, our study suggested that miR-145-5p inhibited cardiac hypertrophy by targeting paxillin and through modulating Rac-1/ JNK /c-jun/ NFATc3/ ANP / Sirt1 signaling, therefore proving novel downstream molecular pathway of miR-145-5p in cardiac hypertrophy.

Obesity in aged population have surges the occurrence of various metabolic disorders including Nonalcoholic fatty liver disease (NAFLD). Apoptosis in the liver is one of the causative factors for NAFLD-induced liver damage. Plants derived bioactive peptides have been shown as an alternative treatment approach for the treating NAFLD due to its less toxicity. Moderate exercise has been reported to improve cellular physiological function prevent age associated metabolic disorders. In the present study, we evaluate the effects of bioactive dipeptide (IF) derived from alcalase potato-protein hydrolysates and swimming exercise in preventing High Fat Diet (HFD)-induced liver damage in senescence accelerated mouse-prone 8 (SAMP8) mice model. Mouse were fed with HFD for 6 weeks followed by oral IF administration or swimming exercise and both for 8 weeks. HFD induces significant structural changes in liver of HFD fed SAMP8 mouse. Both IF administration and exercise prevent the structural abnormalities induced by HFD, however, combined IF treatment and exercise offer better protection. Combined IF treatment and exercise activate PI3K/Akt cell survival protein and effectively inhibit Fas-FADD-induced apoptosis in HFD fed aged mouse. Oral supplementation of bioactive peptide IF combined with moderate swimming exercise effectively alleviate HFD-induced hepatic injury in aged mice.

High-glucose (HG) suppresses mesenchymal stem cell (MSC) functions, resulting in a decrease in cardiac regenerative capability for MSC in diabetes mellitus (DM). Resveratrol enhances MSC functions under stress. This study explores if cardiac regenerative capability can be enhanced in MSCs pretreated with resveratrol in DM rats receiving MSCs. In vitro evidence confirms that HG decreases MSCs capability through suppression of survival markers, AMP-activated protein kinase (AMPK)/Sirtuin 1 (Sirt1) axis, and expression of apoptotic markers. All of these markers are improved when MSCs are cocultured with resveratrol. Wistar male rats were randomly divided into Sham, DM (DM rats), DM rats with autologous transplantation of adipose-derived stem cells (DM + ADSC), and DM rats with resveratrol pretreated ADSC (DM + RSVL-ADSC). Compared to the Sham, DM induces pathological pathways (including fibrosis, hypertrophy, and apoptosis) and suppresses survival as well as the AMPK/Sirt1 axis in the DM group. DM + ADSC slightly improves the above pathways whereas DM + RSVL-ADSC significantly improves the above pathways when compared to the DM group. These results illustrate that resveratrol pretreated with MSCs may show clinical potential in the treatment of heart failure in patients with DM.

It has been increasingly recognized that accelerated atherosclerosis is a major cause of morbidity and mortality in patients with systemic lupus erythematosus, a multisystem autoimmune disease. In this study, we investigated the anti-apoptotic effects of heat-killed Lactobacillus reuteri GMNL-263 on the cardiac tissue of NZB/W F1 mice. The myocardial architecture of the mice heart was observed and evaluated using different staining techniques such as hematoxylin and eosin, TUNEL assay, Masson's trichrome, and fluorescent immunohistochemistry. Additionally, the probiotics-related pathway proteins were analyzed via western blot analysis. Our results showed prevention of enlarged interstitial spaces and abnormal myocardial structures in the hearts of NZB/W F1 mice with L. reuteri GMNL-263 feeding. Significant reduction in TUNEL-positive cells, Fas death receptor-related components, and apoptosis was also detected in the cardiac tissues of the NZB/W F1 mice after L. reuteri GMNL-263 feeding compared with the control group. These findings are the first to reveal the protective effects of L. reuteri GMNL-263 against cardiac abnormalities in NZB/W F1 mice and suggest the potential clinical applications of L. reuteri GMNL-263 in the treatment of SLE-related cardiovascular diseases.

In this study, we used ICI 182 780 (ICI), an estrogen receptor (ER) antagonist, to investigate the estrogenic activity of Danshen, and to further explored whether Danshen extract can block Leu27IGF-II-induced hypertrophy in H9c2 cardiomyoblast cells. We first used an IGF-II analog Leu27IGF-II, which specifically activates IGF2R signaling cascades and induces H9c2 cardiomyoblast cell hypertrophy. However, Danshen extract completely inhibited Leu27IGF-II-induced cell size increase, ANP and BNP hypertrophic marker expression, and IGF2R induction. We also observed that Danshen extract inhibited calcineurin protein expression and NFAT3 nuclear translocation, leading to suppression of Leu27IGF-II-induced cardiac hypertrophy. Moreover, the anti-Leu27IGF-II-IGF2R signaling effect of Danshen was totally reversed by ICI, which suggest the cardio protective effect of Danshen is mediated through estrogen receptors. Our study suggests that, Danshen exerts estrogenic activity, and thus, it could be used as a selective ER modulator in IGFIIR induced hypertrophy model.

Previous studies stated that stem cell functions are reduced under high glucose environment, leading to reduce stem cell capability of tissue regeneration. This study aimed to investigate if stem cells preconditioned with resveratrol show better therapeutic effect on the treatment of liver dysfunction in diabetic rats than stem cells without resveratrol precondition. Male Wistar rats were divided into four groups including sham, DM (diabetic rats), DM + ADSC (DM rats receiving autologous transplantation of adipose-derived stem cells), and DM + pre-R-ADSC (DM rats receiving ADSC preconditioned with resveratrol). Compared with sham group, experimental results showed that DM group induced suppression of survival, suppression of Sirt1, activation of apoptotic, and activation of fibrotic pathways, leading to liver dysfunction. Autologous transplantation of ADSC (DM + ADSC) improved above pathways except for fibrotic signaling. By contrast, transplantation of resveratrol preconditioned ADSC (DM + pre-R-ADSC) significantly improved above pathways including fibrosis. Supplemental evidences suggest that resveratrol precondition increases ADSC viability under high glucose stress via Sirt1 and IGF1R expressions. Furthermore, increased secretion of IGF1 via paracrine route also confirmed in ADSC preconditioned with resveratrol. The experimental results imply that ADSC preconditioned with resveratrol shows potential in the treatment of liver dysfunction in DM patients with liver dysfunction.

Cardiotoxicity by doxorubicin hampers its therapeutic potential as an anticancer drug, but mechanisms leading to cardiotoxicity remain contentious. Through this study, the functional contribution of insulin-like growth factor receptor type II α (IGF-IIRα) which is a novel stress-inducible protein was explored in doxorubicin-induced cardiac stress. Employing both in vitro H9c2 cells and in vivo transgenic rat models (SD-TG [IGF-IIRα]) overexpressing IGF-IIRα specifically in heart, we found that IGF-IIRα leads to cardiac structural abnormalities and functional perturbations that were severely aggravated by doxorubicin-induced cardiac stress. Overexpression of IGF-IIRα leads to cumulative elevation of stress associated cardiac hypertrophy and apoptosis factors. There was a significant reduction of survival associated proteins p-Akt and estrogen receptor β/α, and abnormal elevation of cardiac hypertrophy markers such as atrial natriuretic peptide, cardiac troponin-I, and apoptosis-inducing agents such as p53, Bax, and cytochrome C, respectively. IGF-IIRα also altered the expressions of AT1R, ERK1/2, and p38 proteins. Besides, IGF-IIRα also increased the reactive oxygen species production in H9c2 cells which were markedly aggravated by doxorubicin treatment. Together, we showed that IGF-IIRα is a novel stress-induced protein that perturbed cardiac homeostasis and cumulatively exacerbated the doxorubicin-induced cardiac injury that perturbed heart functions and ensuing cardiomyopathy.

The heart is a very dynamic pumping organ working perpetually to maintain a constant blood supply to the whole body to transport oxygen and nutrients. Unfortunately, it is also subjected to various stresses based on physiological or pathological conditions, particularly more vulnerable to damages caused by oxidative stress. In this study, we investigate the molecular mechanism and contribution of IGF-IIRα in endoplasmic reticulum stress induction in the heart under doxorubicin-induced cardiotoxicity. Using in vitro H9c2 cells, in vivo transgenic rat cardiac tissues, siRNAs against CHOP, chemical ER chaperone PBA, and western blot experiments, we found that IGF-IIRα overexpression enhanced ER stress markers ATF4, ATF6, IRE1α, and PERK which were further aggravated by DOX treatment. This was accompanied by a significant perturbation in stress-associated MAPKs such as p38 and JNK. Interestingly, PARKIN, a stress responsive cellular protective mediator was significantly downregulated by IGF-IIRα concomitant with decreased expression of ER chaperone GRP78. Furthermore, ER stress-associated pro-apoptotic factor CHOP was increased considerably in a dose-dependent manner followed by elevated c-caspase-12 and c-caspase-3 activities. Conversely, treatment of H9c2 cells with chemical ER chaperone PBA or siRNA against CHOP abolished the IGF-IIRα-induced ER stress responses. Altogether, these findings suggested that IGF-IIRα contributes to ER stress induction and inhibits cellular stress coping proteins while increasing pro-apoptotic factors feeding into a cardio myocyte damage program that eventually paves the way to heart failure.

Hepatocellular carcinoma is a common type of liver cancer. Resistance to chemotherapeutic agents is a major problem in cancer therapy. MicroRNAs have been reported in cancer development and tumor growth; however, the relationship between chemoresistance and hepatocellular carcinoma needs to be fully investigated. Here, we treated hepatocellular carcinoma cell line (HA22T) with a histone deacetylase inhibitor to establish hepatocellular carcinoma-resistant cells (HDACi-R) and investigated the molecular mechanisms of chemoresistance in HCC cells. Although histone deacetylase inhibitor could not enhance cell death in HDACi-R but upregulation of miR-107 decreased cell viability both in parental cells and resistance cells, decreased the expression of cofilin-1, enhanced ROS-induced cell apoptosis, and dose-dependently sensitized HDACi-R to HDACi. Further, miR-107 upregulation resulted in tumor cell disorganization in both HA22T and HDACi-R in a mice xenograft model. Our findings demonstrated that miR-107 downregulation leads to hepatocellular carcinoma cell resistance in HDACi via a cofilin-1-dependent molecular mechanism and ROS accumulation.

Ethnopharmacological relevance Platycodon grandiflorum (PG) is a Chinese medical plant used for decades as a traditional prescription to eliminate phlegm, relieve cough, reduce inflammation and lower blood pressure. PG also has a significant effect on the cardiovascular systems. Materials and methods The aqueous extract of Platycodon grandiflorum (JACQ.) A. DC. root was screened for inhibiting Ang II-induced IGF-IIR activation and apoptosis pathway in H9c2 cardiomyocytes. The effects were also studied in spontaneously hypertensive rats (five groups, n=5) using low and high doses of PG for 50 days. The Ang II-induced IGF-IIR activation was analyzed by luciferase reporter, RT-PCR, western blot and surface IGF-IIR expression assay. Furthermore, the major active constituent of PG was carried out by high performance liquid chromatography-mass spectrometry (HPLC-MS). Results Our results indicate that a crude extract of PG significantly suppresses the Ang II-induced IGF-IIR signaling pathway to prevent cardiomyocyte apoptosis. PG extract inhibits Ang II-mediated JNK activation and SIRT1 degradation to reduce IGF-IIR activity. Moreover, PG maintains SIRT1 stability to enhance HSF1-mediated IGF-IIR suppression, which prevents cardiomyocyte apoptosis. In animal models, the administration of PG markedly reduced this apoptotic pathway in the heart of SHRs. Conclusion Taken together, PG may be considered as an effective treatment for cardiac diseases in hypertensive patients.

Doxorubicin (DOX) is an anthracycline antibiotic commonly employed for the treatment of various cancers. However, its therapeutic uses are hampered by side effects associated with cumulative doses during the course of treatment. Whereas deregulation of autophagy in the myocardium has been involved in a variety of cardiovascular diseases, the role of autophagy in DOX-induced cardiomyopathy remains debated. Our earlier studies have shown that DOX treatment in a rat animal model leads to increased expression of the novel stress-inducible protein insulin-like growth factor II receptor-α (IGF-IIRα) in cardiac tissues, which exacerbated the cardiac injury by enhancing oxidative stress and p53-mediated mitochondria-dependent cardiac apoptosis. Through this study, we investigated the contribution of IGF-IIRα to dysregulation of autophagy in heart using both in vitro H9c2 cells (DOX treated, 1 µM) and in vivo transgenic rat models (DOX treated, 5 mg/kg ip for 6 wk) overexpressing IGF-IIRα specifically in the heart. We found that IGF-IIRα primarily localized to mitochondria, causing increased mitochondrial oxidative stress that was severely aggravated by DOX treatment. This was accompanied by a significant perturbation in mitochondrial membrane potential and increased leakage of cytochrome c, causing increased cleaved caspase-3 activity. There were significant alterations in phosphorylated AMP-activated protein kinase (p-AMPK), phosphorylated Unc-51 like kinase-1 (p-ULK1), PARKIN, PTEN-induced kinase 1 (PINK1), microtubule-associated protein 1 light chain 3 (LC3), and p62 proteins, which were more severely disrupted under the combined effect of IGF-IIRα overexpression plus DOX. Finally, LysoTracker Red staining showed that IGF-IIRα overexpression causes lysosomal impairment, which was rescued by rapamycin treatment. Taken together, we found that IGF-IIRα leads to mitochondrial oxidative stress, decreased antioxidant levels, disrupted mitochondrial membrane potential, and perturbed mitochondrial autophagy contributing to DOX-induced cardiomyopathy.

Cardiac hypertrophy is a common phenomenon observed in progressive heart disease associated with heart failure. Insulin-like growth factor receptor II (IGF-IIR) has been much implicated in myocardial hypertrophy. Our previous studies have found that increased activities of signaling mediators, such as calcium/calmodulin-dependent protein kinase II (CaMKII) and calcineurin induces pathological hypertrophy. Given the critical roles played by CaMKII and calcineurin signaling in the progression of maladaptive hypertrophy, we anticipated that inhibition of CaMKII and calcineurin signaling may attenuate IGF-IIR-induced cardiac hypertrophy. The current study, therefore, investigated the effects of IGF-IIR activation on the CaMKII and calcineurin signaling and whether the combinatorial inhibition of the CaMKIIδ and calcineurin signaling could ameliorate IGF-IIR-induced pathological hypertrophy. In the present study, we induced IGF-IIR through the cardiomyocyte-specific transduction of IGFIIY27L via adeno-associated virus 2 (AAV2) to evaluate its effects on cardiac hypertrophy. Interestingly, it was observed that the activation of IGF-IIR signaling through IGFIIY27L induces significant hypertrophy of the myocardium and increased cardiac apoptosis and fibrosis. Moreover, we found that Leu27 IGF-II significantly induced calcineurin and CaMKII expression. Furthermore and importantly, the combinatorial treatment with CaMKII and calcineurin inhibitors significantly alleviates IGF-IIR-induced hypertrophic responses. Thus, it could be envisaged that the inhibition of IGF-IIR may serve as a promising candidate for attenuating maladaptive hypertrophy. Both calcineurin and CaMKII could be valuable targets for developing treatment strategies against hypertension-induced cardiomyopathies.

// Wei-Jen Ting 1,2 , Jaw-Ji Yang 3 , Chia-Hua Kuo 4 , Zi-Jun Xiao 5 , Xin-Ze Lu 5 , Yu-Lan Yeh 6,7 , Cecilia-Hsuan Day 8 , Su-Ying Wen 9,10 , Vijaya PadmaViswanadha 11 , Chong-He Jiang 1 , Wei-Wen Kuo 12,* and Chih-Yang Huang 2,13,14,* 1 The Sixth Affiliated Hospital of Guangzhou Medical University, Guangdong, China 2 Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan 3 Institute of Medicine, School of Dentistry, Chung-Shan Medical University, Taichung, Taiwan 4 Department of Sports Sciences, University of Taipei, Taipei, Taiwan 5 National Taichung First Senior High School, Taichung, Taiwan 6 Department of Pathology, Changhua Christian Hospital, Changhua, Taiwan 7 Jen-Teh Junior College of Medicine, Nursing and Management, Miaoli, Taiwan 8 Department of Nursing, Mei Ho University, Pingtung, Taiwan 9 Graduate Institute of Clinical Medical Science, China Medical University, Taichung, Taiwan 10 Department of Dermatology, Taipei City Hospital, Renai Branch, Taipei, Taiwan 11 Department of Biotechnology, Bharathiar University, Coimbatore, India 12 Department of Biological Science and Technology, China Medical University, Taichung, Taiwan 13 Graduate Institute of Chinese Medical Science, China Medical University, Taichung, Taiwan 14 Department of Health and Nutrition Biotechnology, Asia University, Taichung, Taiwan * These authors have contributed equally to this paper Correspondence to: Chih-Yang Huang, email: // Keywords : environmental tobacco smoke (ETS); Sirt-1; autophagy; Gerotarget Received : November 02, 2015 Accepted : April 16, 2016 Published : May 04, 2016 Abstract Recently, a survey by the Centers for Disease Control and Prevention (CDC) reported that nearly 90% of U.S. adult smokers began smoking at the age of 18. This demonstrates that the exposure to environmental tobacco smoke (ETS) of youngsters today is changing from passive smoking to active smoking (direct inhalation of tobacco). In the current study, an investigation of ETS exposure in young C57BL mice was conducted. After 6 weeks of ETS exposure, the Sirt-1 protein level was decreased and cardiac autophagy was increased in C57BL mice. Furthermore, the IGF2R cardiac hypertrophy signaling pathway was also triggered, although cardiac apoptosis and hypertrophy were not induced. Youngsters’ desire to look more mature is one of the psychological factors that impacts smoking amongst young people. Our results suggest that though ETS exposure might cause cardiac autophagy amongst youngsters, the loss of the longevity Sirt-1 protein and the increase in IGF2R cardiac hypertrophy signaling could still promote heart diseases that are age-specific.

Angiotensin II (Ang II) is a very important cardiovascular disease inducer and may cause cardiac pathological hypertrophy and remodeling. We evaluated a Chinese traditional medicine, alpinate oxyphyllae fructus (AOF), for therapeutic efficacy for treating Ang II-induced cardiac hypertrophy. AOF has been used to treat patients with various symptoms accompanying hypertension and cerebrovascular disorders in Korea. We investigated its protective effect against Ang II-induced cytoskeletal change and hypertrophy in H9c2 cells. The results showed that treating cells with Ang II resulted in pathological hypertrophy, such as increased expression of transcription factors NFAT-3/p-NFAT-3, hypertrophic response genes (atrial natriuretic peptide [ANP] and b-type natriuretic peptide [BNP]), and Gαq down-stream effectors (PLCβ3 and calcineurin). Pretreatment with AOF (60-100 μg/mL) led to significantly reduced hypertrophy. We also found that AOF pretreatment significantly suppressed the cardiac remodeling proteins, metalloproteinase (MMP9 and MMP2), and tissue plasminogen activator (tPA), induced by Ang II challenge. In conclusion, we provide evidence that AOF protects against Ang II-induced pathological hypertrophy by specifically inhibiting the insulin-like growth factor (IGF) II/IIR-related signaling pathway in H9c2 cells. AOF might be a candidate for cardiac hypertrophy and ventricular remodeling prevention in chronic cardiovascular diseases.

Extreme hypoxia often leads to myocardial apoptosis and causes heart failure. Angiotensin-(1-7)Ang-(1-7) is well known for its cardio-protective effects. However, the effects of Ang-(1-7) on long-term hypoxia (LTH)-induced apoptosis remain unknown. In this study, we found that Ang-(1-7) reduced myocardial apoptosis caused by hypoxia through the Mas receptor. Activation of the Ang-(1-7)/Mas axis down-regulated the hypoxia pro-apoptotic signaling cascade by decreasing the protein levels of hypoxia-inducible factor 1α (HIF-1α) and insulin-like growth factor binding protein-3 (IGFBP3). Moreover, the Ang-(1-7)/Mas axis further inhibited HIF-1α nuclear translocation. On the other hand, Ang-(1-7) activated the IGF1R/PI3K/Akt signaling pathways, which mediate cell survival. However, the above effects were abolished by A779 treatment or silencing of Mas expression. Taken together, our findings indicate that the Ang-(1-7)/Mas axis protects cardiomyocytes from LTH-stimulated apoptosis. The protective effect of Ang-(1-7) is associated with the inhibition of HIF-1α nuclear translocation and the induction of IGF1R and Akt phosphorylation.

This study investigates the cardio-protective effect of Nos. 1 and 5 extracts from Taiwan Mingjian Oolong Tea on H9c2 cardiomyoblast cells treated with isoproterenol (ISO). Treatment with Nos. 1 and 5 extracts increased cell viability and blocked apoptosis in ISO exposed H9c2 cells. Moreover, Nos. 1 and 5 extracts blocked hypertrophy markers like G[Formula: see text]s, calcineurin, NFATc3, and BNP, thereby increasing cell proliferation markers -PI3K and AKT in a dose dependent manner. In contrast, apoptotic proteins, such as caspase-3 and cytochrome c were decreased in H9c2 cells treated with Nos. 1 and 5 extracts. We confirmed that the protective effect of No. 1 extract was partially mediated through the expression of ERK and p38, however, the No. 5 extract showed a protective effect via the ERK, JNK, and p38 pathways. This evidence provides new insights into the pharmacological role and therapeutic mechanism of Taiwan Mingjian Oolong Tea in heart diseases.

Pancreatic damage is the major causative agent in type 1 diabetes mellitus (DM). Several strategies have been suggested to regenerate pancreatic functions, such as stem cell transplantation and administration of active components isolating from natural herbals. This study aims to investigate if the synergistically protective effect on damaged pancreatic tissues can be observed in STZ-induced DM rats with autologous transplantation of adipose-derived stem cells (ADSC) coupling with oral administration of resveratrol. Pathological conditions can be recognized in DM rats with pancreatic damage, including reduction of islet size, suppression of survival markers, downregulation of AMPK/Sirt1 axis, and activation of apoptotic signaling. Autologous transplantation of ADSC slightly improves pancreatic functions, whereas autologous transplantation of ADSC coupling with oral administration of resveratrol significantly improves pancreatic functions in DM rats. We suggest that oral administration of resveratrol may enhance the therapeutic effect on DM patients receiving autologous transplantation of ADSC.

Consumption of high fat diet (HFD) is associated with increased cardiovascular risk factors among elderly people. Aging and obesity induced-cardiac remodeling includes hypertrophy and fibrosis. Gelsolin (GSN) induces cardiac hypertrophy and TGF-β, a key cytokine, which induces fibrosis. The relationship between TGF-β and GSN in aging induced cardiac remodeling is still unknown. We evaluated the expressions of TGF-β and GSN in HFD fed 22 months old aging SD rats, followed by the administration of either probucol or alcalase potato protein hydrolysate (APPH). Western blotting and Masson trichrome staining showed that APPH (45 and 75 mg/kg/day) and probucol (500 mg/kg/day) treatments significantly reduced the aging and HFD-induced hypertrophy and fibrosis. Echocardiograph showed that the performance of the hearts was improved in APPH, and probucol treated HFD aging rats. Serum from all rats was collected and H9c2 cells were cultured with collected serums separately. The GSN dependent hypertrophy was inhibited with an exogenous TGF-β in H9c2 cells cultured in HFD+ APPH treated serum. Thus, we propose that along with its role in cardiac fibrosis, TGF-β also acts as an upstream activator of GSN dependent hypertrophy. Hence, TGF-β in serum could be a promising therapeutic target for cardiac remodeling in aging and/or obese subjects.

Obesity is one of the major risk factors for nonalcoholic fatty liver disease (NAFLD), and NAFLD is highly associated with an increased risk of cardiovascular disease (CVD). Scholars have suggested that certain probiotics may significantly impact cardiovascular health, particularly certain Lactobacillus species, such as Lactobacillus reuteri GMNL-263 (Lr263) probiotics, which have been shown to reduce obesity and arteriosclerosis in vivo. In the present study, we examined the potential of heat-killed bacteria to attenuate high fat diet (HFD)-induced hepatic and cardiac damages and the possible underlying mechanism of the positive effects of heat-killed Lr263 oral supplements. Heat-killed Lr263 treatments (625 and 3125 mg/kg-hamster/day) were provided as a daily supplement by oral gavage to HFD-fed hamsters for eight weeks. The results show that heat-killed Lr263 treatments reduce fatty liver syndrome. Moreover, heat-killed Lactobacillus reuteri GMNL-263 supplementation in HFD hamsters also reduced fibrosis in the liver and heart by reducing transforming growth factor β (TGF-β) expression levels. In conclusion, heat-killed Lr263 can reduce lipid metabolic stress in HFD hamsters and decrease the risk of fatty liver and cardiovascular disease.

Myocardial infarction (MI) usually results in myocardial ischemia, remodeling and hypoxia that lead to cell death. To date, the insulin-like growth factor binding protein-3 (IGFBP3) is known to play an important role in insulin growth factor (IGF) bioavailability. Previous studies have found that hypoxia results in cell apoptosis. However, the detailed mechanism and roles of IGFBP3 in long-term hypoxia (LTH) regulated heart cell apoptosis remains unknown. In this study H9c2 cardiomyoblast cells were treated with investigated long-term hypoxic exposure with the possible mechanisms involved. The results showed that LTH enhanced IGFBP3 protein synthesis and induced its secretion. The accumulated IGFBP3 sequestered Insulin growth factor 1 (IGF-1) away from the type I IGF receptor (IGF-1 R), which blocked the IGF1R/PI3K/Akt survival signaling pathway, resulting in cell apoptosis. According to our findings, IGFBP3 could be a valuable target for developing treatments for cardiac diseases in long-term hypoxia exposure patients.

The prevalence of obesity is high in older adults. Alcalase potato protein hydrolysate (APPH), a nutraceutical food, might have greater benefits and be more economical than hypolipidemic drugs. In this study, serum lipid profiles and heart protective effects were evaluated in high fat diet (HFD) induced hyperlipidemia in aging rats treated with APPH (15, 45 and 75 mg/kg/day) and probucol (500 mg/kg/day). APPH treatments reduced serum triacylglycerol (TG), total cholesterol (TC), and low density lipoprotein (LDL) levels to the normal levels expressed in the control group. Additionally, the IGF1R-PI3K-Akt survival pathway was reactivated, and Fas-FADD (Fas-associated death domain) induced apoptosis was inhibited by APPH treatments (15 and 45 mg/kg/day) in HFD aging rat hearts. APPH (75 mg/kg/day) rather than probucol (500 mg/kg/day) treatment could reduce serum lipids without affecting HDL expression. The heart protective effect of APPH in aging rats with hyperlipidemia was through lowering serum lipids and enhancing the activation of the compensatory IGF1R-PI3K-Akt survival pathway.

Background/Aims: The risk of heart disease is higher in males than in females. However, this advantage of females declines with increasing age, presumably a consequence of decreased estrogen secretion and malfunctioning of the estrogen receptor. We previously demonstrated that 17β-estradiol (E2) prevents cardiomyocyte hypertrophy, autophagy and apoptosis via estrogen receptor α (ERα), but the effects of ERβ on myocardial injury remained elusive. The present paper thus, investigated the cardioprotective effects of estrogen (E2) and ERβ against hypoxia-induced cell death. Methods: Transient transfection of Tet-On ERβ gene construct was used to overexpress ERβ in hypoxia-treated H9c2 cardiomyoblast cells. Results: Our data revealed that IGF1R, Akt phosphorylation and Bcl-2 expression are enhanced by ERβ in H9c2 cells. Moreover, ERβ overexpression reduced accumulation of hypoxia-related proteins, autophagy-related proteins and mitochondria-apoptotic proteins and enhanced the protein levels of Bcl-2, pAkt and Bad under hypoxic condition. In neonatal rat ventricular myocytes (NRVMs), we observed that hypoxia induced cell apoptosis as measured by TUNEL staining, and E2 and/or ERβ could totally abolish hypoxia-induced apoptosis. The suppressive effects of E2 and/or ERβ in hypoxia-treated NRVMs were totally reversed by ER antagonist, ICI. Taken together, E2 and/or ERβ exert the protective effect through repressed hypoxia-inducible HIF-1α, BNIP3 and IGFBP-3 levels to restrain the hypoxia-induced autophagy and apoptosis effects in H9c2 cardiomyoblast cells. Conclusion: The results suggest that females probably could tolerate better prolonged hypoxia condition than males, and E2/ERβ treatment could be a potential therapy to prevent hypoxia-induced heart damage.”

Background The goal of this study is to determine if salidroside has protective effects on hypoxia-induced cardiac widely dispersed apoptosis in mice with severe sleep apnea model. Methods Sixty-four C57BL/6J mice 5–6months of age were divided into four groups, i.e. Control group (21% O 2 , 24h per day, 8weeks, n =16); Hypoxia group (Hypoxia: 7% O 2 60s, 20% O 2 alternating 60s, 8h per day, 8weeks, n =16); and Hypoxia+S10 and Hypoxia+S30 groups (Hypoxia for 1st 4weeks, hypoxia pretreated 10mg/kg and 30mg/kg salidroside by oral gavage per day for 2nd 4weeks, n =16 and 16). The excised hearts from four groups were measured by the heart weight index, H&E staining, TUNEL-positive assays and Western blotting. Results TUNEL-positive apoptotic cells in mice heart were less in Hypoxia+S10 and Hypoxia+S30 than those in the Hypoxia group. Compared with Hypoxia, the protein levels of Fas ligand, Fas death receptors, Fas-Associated Death Domain (FADD), activated caspase 8, and activated caspase 3 (Fas pathways) were decreased in Hypoxia+S10 and Hypoxia+S30. In the mitochondria pathway, the protein levels of BcLx, Bcl2, and Bid (anti-apoptotic Bcl2 family) in Hypoxia+S10 and Hypoxia+S30 were more than those in Hypoxia. The protein levels of Bax, t-Bid, activated caspase 9, and activated caspase 3 were less in Hypoxia+S10 and Hypoxia+S30 than those in hypoxia. Conclusions Our findings suggest that salidroside has protective effects on chronic intermittent hypoxia-induced Fas-dependent and mitochondria-dependent apoptotic pathways in mice hearts.

Apoptosis is recognized as a predictor of adverse outcomes in subjects with cardiac diseases. The aim of this study was to explore the effects of probiotic-fermented purple sweet potato yogurt (PSPY) with high γ-aminobutyric acid (GABA) content on cardiac apoptosis in spontaneously hypertensive rat (SHR) hearts. The rats were orally adminsitered with 2 different concentrations of PSPY (10 and 100%) or captopril, 15.6 mg/kg, body weight (BW)/day. The control group was administered distilled water. DAPI and TUNEL staining were used to detect the numbers of apoptotic cells. A decrease in the number of TUNEL-positive cardiac myocytes was observed in the SHR-PSPY (10 and 100%) groups. In addition, the levels of key components of the Fas receptor- and mitochondrial-dependent apoptotic pathways were determined by western blot analysis. The results revealed that the levels of the key components of the Fas receptor- and mitochondrial-dependent apoptotic pathway were significantly decreased in the SHR-captopril, and 10 and 100% PSPY groups. Additionally, the levels of phosphorylated insulin-like growth factor‑I receptor (p-IGF‑IR) were increased in SHR hearts from the SHR-control group; however, no recovery in the levels of downstream signaling components was observed. In addition, the levels of components of the compensatory IGF-IR-dependent survival pathway (p-PI3K and p-Akt) were all highly enhanced in the left ventricles in the hearts form the SHR-10 and 100% PSPY groups. Therefore, the oral administration of PSPY may attenuate cardiomyocyte apoptosis in SHR hearts by activating IGF‑IR-dependent survival signaling pathways.

Increased serum norepinephrine level is one of pathological processes relating to heart disease (HD). Estrogens are considered as potential therapeutics for the treatment of HD; however, estrogen supplementation shows some side-effects, such as increasing the risk of developing breast, endometrial and ovarian cancers. This study investigated the cardio-protective effects of daidzein (Dai), a selective estrogen receptor modulator (SERM) from soy bean extract, in H9c2 cardiomyoblast cells treated with isoproterenol (ISO), a norepinephrine analog. In this in vitro model, H9c2 cells treated with Dai at different concentrations showed no statistical difference in cell viability. TdT-mediated digoxigenin-dUTP nick-end labeling (TUNEL) data and western blotting results indicated that Dai treated-H9c2 cells recovered from the damage induced by ISO. The recovery effects of Dai on ISO-induced damage were blocked by inhibition of Akt activation through adding Akt inhibitor. On the other hand, the fold changes of phosphorylated Akt (p-Akt)/Akt normalized with the control for con, 0.25, 0.5, 1, 3 and 24 h of treatment were 1, 2, 5, 13, 11 and 10, respectively. In conclusion, Dai ameliorates apoptosis of cardiomyoblasts induced by ISO through Akt signaling pathway.

Background San Huang Shel Shin Tang (SHSST) is a traditional herbal decoction used as a hepato-protective agent and is composed of Rheum officinale Baill, Scutellaria baicalnsis Geprgi and Coptis chinensis Franch (2:1:1 w/w). Beta-cyclodextrin (β-CD) modification may potentially increase the solubility and spectral properties of SHSST. Methods In this research, the hepato-protective effects of unmodified SHSST, β-CD modified SHSST complex (SHSSTc) and silymarin were evaluated in carbon tetrachloride (CCl4) induced acute hepatotoxicity in rats. Results SHHSTc (40 mg/kg/day) and silymarin (100 mg/kg/day) both decreased the CCl4-induced cirrhosis pathway-related transforming growth factor beta (TGF-β) and apoptosis pathway-related caspase-8 protein expressions, but SHSST (40 mg/kg/day) did not reduce TGF-β and caspase-8 significantly . Moreover, SHHSTc (40 mg/kg/day) enhanced the activation of insulin-like growth factor 1 receptor (IGF1R) mediated survival pathway than the silymarin (100 mg/kg/day) to protect the liver from damage induced by CCl4. Conclusions β-CD modification promotes hepato-protective effects of SHSST and reduces the required-dosage of the SHSST.

The present study tests a hypothesis that cardioprotective effects mediated by autologous adipose-derived stem cells (ADSC) in rats afflicted with insulin-dependent diabetes mellitus (IDDM) may be synergistically enhanced by oral treatment with green tea epigallocatechin gallate (EGCG). Wistar rats were divided into sham, DM, DM+ADSC (autologous transplanted 1 × 106 cells per rat), and DM+ADSC+E (E, green tea oral administration EGCG). Heart tissues were isolated from all rats, and investigations were performed after 2-mo treatment. In the sham, DM, and DM+ADSC groups, we found that DM induced cardiac dysfunction (sham and DM) and autologous ADSC transplantation could partially recover cardiac functions (DM and DM+ADSC) in DM rats. Compared with DM+ADSC, significant improvement in cardiac functions can be observed in DM+ADSC+E in echocardiographic data, histological observations, and even cellular protein expression. Oral green tea EGCG administration and autologous ADSC transplantation show synergistically beneficial effects on diabetic cardiac myopathy in DM rats. NEW & NOTEWORTHY Cardiomyopathy can be induced in rats with diabetes mellitus (DM). Heart function can be restored in DM rats with adipose-derived stem cell treatment. Oral epigallocatechin gallate (EGCG) administration synergistically enhances cardiac function in DM rats with stem cell treatment. The EGCG and stem cell treatment cross-effect occurs via survival protein expression.

High-calorie diet-induced obesity leads to cardiomyocyte dysfunction and apoptosis. Impaired regulation of epididymal fat content in obese patients has been known to increase the risk of cardiac injury. In our study, a lactic acid bacteria, Lactobacillus reuteri GMNL-263, was evaluated for its potential to reduce body weight and body fat ratio and to prevent heart injury in rats with high-fat diet-induced obesity. Lactic acid bacteria supplementation restored the cardiac function and decreased the physiological changes in the heart of the obese rats. In addition, the Fas/Fas-associated protein pathway-induced caspase 3/e Poly polymerase mediated apoptosis in the cardiomyocytes of the obese rats was reversed in the Lr263-treated rats. These results reveal that fed with Lr-263 reduces body fat ratio, inhibits caspase 3-mediated apoptosis and restores cardiac function in obese rats through recovery of ejection fraction and fractional shortening. Our results indicated that the administration of Lr263 lactic acid bacteria can significantly down-regulate body fat and prevent cardiomyocyte injury in obese rats.

The aim of this study was to establish the effective hepatoprotective properties of traditional Chinese medicines (TCMs) in fibrotic rat liver regeneration after partial hepatectomy (PHx). Fibrosis was induced in rats by ethanol (EtOH, 5 ml/kg) administration for 6, 24, 72, and 168 h. The rats were then fed four TCMs (1 g/kg/day, Codonopsis pilosula (CP), Salvia miltorrhiza Bunge (SMB), Bupleurum kasi (BK), and Elephantopus scaber L (ESL)) to Spraque-Dawley rats for 6, 24, 72 and 168 h, respectively. Surgical 70% cirrhotic fibrosis PHx was then conducted at 6, 24, 72, and 168 h. The effects on liver regeneration were examined to estimate and measure hepatocyte growth factor (HGF), focal adhesion kinase (FAK), Cyclin D1, Cyclin E, and retinoblastoma protein (pRb) protein expression using Western blotting analysis. Cyclin D1, matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitors of metalloproteinase (TIMP)-1, TIMP-2 and TIMP-3 mRNA by Reverse transcription polymerase chain reaction (RT-PCR) were analyzed in cirrhotic fibrosis rats. Transforming growth factor-β1 (TGF-β1), Cyclin D1, Cyclin E, pRb and E2F mRNA expression levels were determined in fibrotic rats following PHx using RT-PCR. We found elevated glutamyl oxaloacetic transaminase (GOT), glutamyl pyrubic transaminase (GPT), alkaline phosphatase (ALP), gammaglutamyl transpeptidase (γ-GT), glutathione (GSH), nonprotein sulfhydryl (NPSH) and total bilirubin in serum after 6 h EtOH administration. These levels were progressively decreased over 168 h. Total protein and albumin were reduced in serum after 6 h administration and then progressively increased. In contrast, tissues disorder histology and morphology were determined in liver sections. After rats were fed TCMs we found that SMB extraction not only induced HGF, FAK, Cyclin D1, and pRb protein expression and Cyclin D1 mRNA increases, but also reduced MMP-2 and MMP-9 after 24 and 72 h post injury. In the cell cycle S phase the Cyclin E protein expression was increased by ESL. CP induced TIMP-1, TIMP-2 and TIMP-3 mRNA increases in fibrotic rats. We detected liver regeneration in fibrotic rats. We also found that the liver regeneration index increased from 6 to 168 h post PHx. After 168 h fibrotic liver regeneration rats exhibited reduced TGF-β1 mRNA expression and enhanced Cyclin D1, Cyclin E, pRb and E2F mRNA expression. TCMs play a crucial role in the early mediating process in fibrotic rat liver regeneration after PHx.

Severe and potentially fatal hypotension and cardiac contractile dysfunction are common symptoms in patients with sepsis. LPS was previously found to dramatically upregulate expression of fibrosis-related factors FGF-2, uPA, MMP-2, and MMP-9 in primary cardiac fibroblasts. MMPs are capable of denaturing and degrading fibrillar collagens and other components of the extracellular matrix (ECM). Studies have shown that dysregulation of expression of MMPs is associated with development of myocardial extracellular matrix remodeling and cardiac fibrosis, which contribute to progression of heart failure. In this study, H9c2 cells and cardiac fibroblasts were divided into five treatment groups: control, LPS (1 μg/mL) and three concentrations of FC

Metabolic regulation is inextricably linked with cardiac function. Fatty acid metabolism is a significant mechanism for creating energy for the heart. However, cardiomyocytes are able to switch the fatty acids or glucose, depending on different situations, such as ischemia or anoxia. Lipotoxicity in obesity causes impairments in energy metabolism and apoptosis in cardiomyocytes. We utilized the treatment of H9c2 cardiomyoblast cells palmitic acid (PA) as a model for hyperlipidemia to investigate the signaling mechanisms involved in these processes. Our results show PA induces time- and dose-dependent lipotoxicity in H9c2 cells. Moreover, PA enhances cluster of differentiation 36 (CD36) and reduces glucose transporter type 4 (GLUT4) pathway protein levels following a short period of treatment, but cells switch from CD36 back to the GLUT4 pathway after during long-term exposure to PA. As sirtuin 1 (SIRT1) and protein kinase Cζ (PKCζ) play important roles in CD36 and GLUT4 translocation, we used the SIRT1 activator resveratrol and si-PKCζ to identify the switches in metabolism. Although PA reduced CD36 and increased GLUT4 metabolic pathway proteins, when we pretreated cells with resveratrol to activate SIRT1 or transfected si-PKCζ, both were able to significantly increase CD36 metabolic pathway proteins and reduce GLUT4 pathway proteins. High-fat diets affect energy metabolism pathways in both normal and aging rats and involve switching the energy source from the CD36 pathway to GLUT4. In conclusion, PA and high-fat diets cause lipotoxicity in vivo and in vitro and adversely switch the energy source from the CD36 pathway to the GLUT4 pathway.

Cytoskeleton filaments play an important role in cellular functions such as maintaining cell shape, cell motility, intracellular transport, and cell division. Actin-binding proteins (ABPs) have numerous functions including regulation of actin filament nucleation, elongation, severing, capping, cross linking, and actin monomer sequestration. Gelsolin (GSN) is one of the actin-binding proteins. Gelsolin (GSN) is one of the actin-binding proteins that regulate cell morphology, differentiation, movement, and apoptosis. GSN also regulates cell morphology, differentiation, movement, and apoptosis. In this study, we have used H9c2 cardiomyoblast cell and H9c2-GSN stable clones to understand the roles and mechanisms of GSN overexpression in hypoxia-induced cardiomyoblast cell death. The data show that hypoxia or GSN overexpression induces HIF-1α expression and reduces the expression of survival markers p-Akt and Bcl-2 in H9c2 cardiomyoblast cells. Under hypoxic conditions, GSN overexpression further reduces p-Akt expression and elevates total as well as cleaved GSN levels and HIF-1α levels. In addition, GSN overexpression enhances apoptosis in cardiomyoblasts under hypoxia. Hypoxic challenge further induced activated caspase-3 and cell death that was attenuated after GSN knock down, which implies that GSN is a critical therapeutic target against hypoxia-induced cardiomyoblast cell death.

Obesity and hyperlipidaemia increase the risk of CVD. Some strains of probiotics have been suggested to have potential applications in cardiovascular health by lowering serum LDL-cholesterol. In this work, high-fat diet-induced hyperlipidaemia in hamsters was treated with different doses (5×108 and 2·5×109 cells/kg per d) of heat-killed Lactobacillus reuteri GMNL-263 (Lr263) by oral gavage for 8 weeks. The serum lipid profile analysis showed that LDL-cholesterol and plasma malondialdehyde (P-MDA) were reduced in the GMNL-263 5×108 cells/kg per d treatment group. Total cholesterol and P-MDA were reduced in the GMNL-263 2·5×109 cells/kg per d treatment group. In terms of heart function, the GMNL-263 2·5×109 cells/kg per d treatments improved the ejection fraction from 85·71 to 91·81 % and fractional shortening from 46·93 to 57·92 % in the high-fat diet-fed hamster hearts. Moreover, the GMNL-263-treated, high-fat diet-fed hamster hearts exhibited reduced Fas-induced myocardial apoptosis and a reactivated IGF1R/PI3K/Akt cell survival pathway. Interestingly, the GMNL-263 treatments also enhanced the heat-shock protein 27 expression in a dose-dependent manner, but the mechanism for this increase remains unclear. In conclusion, supplementary heat-killed L. reuteri GMNL-263 can slightly reduce serum cholesterol. The anti-hyperlipidaemia effects of GMNL-263 may reactivate the IGF1R/PI3K/Akt cell survival pathway and reduce Fas-induced myocardial apoptosis in high-fat diet-fed hamster hearts.

Exposure to second-hand tobacco smoke (SHS) has been epidemiologically linked to heart disease among non-smokers. However, the molecular mechanism behind SHS-induced cardiac disease is not well known. This study found that SD rats exposed to cigarette smoke at a dose of 10 cigarettes for 30 min twice a day for 1 month had a reduced left ventricle-to-tibia length ratio (mg/mm), increased cardiomyocyte apoptosis by TUNEL assay and a wider interstitial space by H&E staining. However, lumbrokinase and dilong both reversed the effects of SHS. Western blotting demonstrated significantly increased expression of the pro-apoptotic protein caspase-3 in the hearts of the rats exposed to SHS. Elevated protein expression levels of Fas, FADD and the apoptotic initiator activated caspase-8, a molecule in the death-receptor-dependent pathway, coupled with increased t-Bid and apoptotic initiator activated caspase-9 were found. Molecules in the mitochondria-dependent pathway, which disrupts mitochondrial membrane potential, were also found in rats exposed to SHS. These factors indicate myocardial apoptosis. However, treatment with lumbrokinase and dilong inhibited SHS-induced apoptosis. Regarding regulation of the survival pathway, we found in western blot analysis that cardiac protein expression of pAkt, Bcl2, and Bcl-xL was significantly down-regulated in rats exposed to SHS. These effects were reversed with lumbrokinase and dilong treatment. The effects of SHS on cardiomyocytes were also found to be mediated by the Fas death receptor-dependent apoptotic pathway, an unbalanced mitochondria membrane potential and decreased survival signaling. However, treatment with both lumbrokinase and dilong inhibited the effects of SHS. Our data suggest that lumbrokinase and dilong may prevent heart disease in SHS-exposed non-smokers.

Background: Secondhand smoke (SHS) exposure is associated with increased risk of cardiovascular disease. Aging is a physiological process that involves progressive impairment of normal heart functions due to increased vulnerability to damage. This study examines secondhand smoke exposure in aging rats to determine the age-related death-survival balance. Methods: Rats were placed into a SHS exposure chamber and exposed to smog. Old age male Sprague-Dawley rats were exposed to 10 cigarettes for 30 min, day and night, continuing for one week. After 4 weeks the rats underwent morphological and functional studies. Left ventricular sections were stained with hematoxylin-eosin for histopathological examination. TUNEL detected apoptosis cells and protein expression related death and survival pathway were analyzed using western blot. Results: Death receptor-dependent apoptosis upregulation pathways and the mitochondria apoptosis proteins were apparent in young SHS exposure and old age rats. These biological markers were enhanced in aging SHS-exposed rats. The survival pathway was found to exhibit compensation only in young SHS-exposed rats, but not in the aging rats. Further decrease in the activity of this pathway was observed in aging SHS-exposed rats. TUNEL apoptotic positive cells were increased in young SHS-exposed rats, and in aging rats with or without SHS-exposure. Conclusions: Aging reduces IGF-I compensated signaling with accelerated cardiac apoptotic effects from second-hand smoke.

Exposure to environmental tobacco smoke has been epidemiologically linked to heart disease among nonsmokers. However, the molecular mechanism behind the pathogenesis of cardiac disease is unknown. In this study, we found that Wistar rats, exposed to tobacco cigarette smoke at doses of 5, 10, or 15 cigarettes for 30 min twice a day for 1 month, had a dose-dependently reduced heart weight to body weight ratio and enhanced interstitial fibrosis as identified by histopathologic analysis. The mRNA and activity of matrix metalloprotease-2 (MMP-2), representing the progress of cardiac remodeling, were also elevated in the heart. In addition, we used reverse-transcriptase polymerase chain reaction and Western blotting to demonstrate significantly increased levels of the apoptotic effecter caspase-3 in treated animal hearts. Dose-dependently elevated mRNA and protein levels of Fas, and promoted apoptotic initiator caspase-8 (active form), a molecule of a death-receptor–dependent pathway, coupled with unaltered or decreased levels of cytosolic cytochrome c and the apoptotic initiator caspase-9 (active form), molecules of mitochondria-dependent pathways, may be indicative of cardiac apoptosis, which is Fas death-receptor apoptotic-signaling dependent, but not mitochondria pathway dependent in rats exposed to second-hand smoke (SHS). With regard to the regulation of survival pathway, using dot blotting, we found cardiac insulin-like growth factor-1 (IGF-1) and IGF-1 receptor mRNA levels to be significantly increased, indicating that compensative effects of IGF-1 survival signaling could occur. In conclusion, we found that the effects of SHS on cardiomyocyte are mediated by the Fas death-receptor–dependent apoptotic pathway and might be related to the epidemiologic incidence of cardiac disease of SHS-exposed non-smokers.

Systemic lupus erythematosus (SLE) is a disease that mostly affects women. Accelerated atherosclerosis is a high-risk factor associated with SLE patients. SLE associated with cardiovascular disease is one of the most important causes of death. In this study, we demonstrated that Lactobacillus paracasei GMNL-32 (GMNL-32), a probiotic species, exhibits anti-fibrosis and anti-apoptotic effects on the cardiac tissue of NZB/WF1 mice. Female NZB/W F1 mice, a well-known and commonly used lupus-prone mouse strain, were treated with or without GMNL-32 administration for 12 weeks. Oral administration of GMNL-32 to NZB/WF1 mice significantly increased the ventricular thickness when compared to that of NZB/WF1 mice. Administration of GMNL-32 significantly attenuated the cardiac cell apoptosis that was observed in exacerbate levels in the control NZB/WF1 mice. Further, the cellular morphology that was slightly distorted in the NZB/WF1 was effectively alleviated in the treatment group mice. In addition, GMNL-32 reduced the level of Fas death receptor-related pathway of apoptosis signaling and enhanced anti-apoptotic proteins. These results indicate that GMNL-32 exhibit an effective protective effect on cardiac cells of SLE mice. Thus, GMNL-32 may be a potential therapeutic strategy against SLE associated arthrosclerosis.

Cardiomyocyte hypertrophy is an adaptive response of heart to various stress conditions. During the period of stress accumulation, transition from physiological hypertrophy to pathological hypertrophy results in the promotion of heart failure. Our previous studies found that ZAK, a sterile alpha motif and leucine zipper containing kinase, was highly expressed in infarcted human hearts and demonstrated that overexpression of ZAK induced cardiac hypertrophy. This study evaluates, cellular events associated with the expression of two doxycycline (Dox) inducible Tet-on ZAK expression systems, a Tet-on ZAK WT (wild-type), and a Tet-on ZAK DN (mutant, Dominant-negative form) in H9c2 myoblast cells; Tet-on ZAK WT was found to increase cell size and hypertrophic marker BNP in a dose-dependent manner. To ascertain the mechanism of ZAK-mediated hypertrophy, expression analysis with various inhibitors of the related upstream and downstream proteins was performed. Tet-on ZAK WT expression triggered the p38 and JNK pathway and also activated the expression and nuclear translocation of p-GATA4 and p-c-Jun transcription factors, without the involvement of p-ERK or NFATc3. However, Tet-on ZAK DN showed no effect on the p38 and JNK signaling cascade. The results showed that the inhibitors of JNK1/2 and p38 significantly suppressed ZAK-induced BNP expression. The results show the role of ZAK and/or the ZAK downstream events such as JNK and p38 phosphorylation, c-Jun, and GATA-4 nuclear translocation in cardiac hypertrophy. ZAK and/or the ZAK downstream p38, and JNK pathway could therefore be potential targets to ameliorate cardiac hypertrophy symptoms in ZAK-overexpressed patients.

Through a high-fat diet, obesity leads to cardiomyocyte dysfunction and apoptosis. In addition, there is no evidence that probiotics have potential health effects associated with cardiac apoptosis in obese rats. The present study aimed to explore the effects of probiotics on obesity and cardiac apoptosis in rats fed a high-fat diet (HF). Eight‑week‑old male Wistar rats were separated randomly into five equally sized experimental groups: Normal diet (NC) and high-fat diet (HFC) groups, and high-fat diet supplemented with low (HFL), medium (HFM) or high (HFH) doses of multi‑strain probiotics groups. The rats were subsequently studied for 8 weeks. Food intake and body weights were recorded following sacrifice, and food utilization rates, body fat and serum cholesterol levels were analysed. The myocardial architecture of the left ventricle was evaluated by hematoxylin‑eosin staining, and key apoptotic‑related pathway molecules were analysed by western blotting. Rat weights and triglyceride levels were decreased with oral administration of high doses of probiotics (HFH) compared to the HFC group. Abnormal myocardial architecture and enlarged interstitial spaces were observed in HFC hearts, but were significantly decreased in groups that were provided multi‑strain probiotics compared with NC hearts. Western blot analysis demonstrated that key components of the Fas receptor‑ and mitochondrial‑dependent apoptotic pathways were significantly suppressed in multi‑strain probiotic treated groups compared to the HF group. Additionally, cardiac insulin, such as the insulin‑like growth factor I receptor (IGFIR)‑dependent survival signalling components, were highly induced in left ventricles from rats administered probiotics. Together, these findings strongly suggest that oral administration of probiotics may attenuate cardiomyocyte apoptosis by activation of the phosphatidylinositol‑3 kinase/AKT survival‑signalling pathway in obese rats.

Obesity is often associated with the development of cardiac hypertrophy but the hypertrophy-related pathways in obesity remain unknown. The purpose of this study was to evaluate cardiac hypertrophy-related markers, atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP), tumor necrosis factor-alpha (TNFα) and hypertrophy-related pathways, interleukin (IL)-6-STAT3, IL-6-MEK5-ERK5 and calcineurin-nuclear factor of activated T-cells (NFAT)3 in the excised hearts from obese rats. Twelve obese Zucker rats were studied at 5-6 months of age and twelve age-matched lean Zucker rats served as the control group. The cardiac characteristics, myocardial architecture, ANP, BNP, TNFα levels, IL-6, STAT3, p-STAT3, MEK5, ERK-5, p-ERK-5, calcineurin and NFAT3 in the left ventricle from the rats were measured by heart weight index, echocardiography, vertical cross section, histological analysis, reverse transcription polymerase chain reaction and western blotting. Compared with the lean control, the whole heart weight, the left ventricule weight, the ratio of the whole heart weight to tibia length, echocardiographic interventricular septum, left ventricular posterior wall thickness, myocardial morphological changes and systolic blood pressure were found to increase in the obese rats. The protein levels of ANP, BNP, TNFα, IL-6, STAT3, p-STAT3, MEK-5, ERK-5, p-ERK 5, calcineurin and NFAT3 were also significantly increased in the hearts of the obese rats. The results showed that the hypertrophy-related markers, ANP, BNP and TNFα, the hypertrophy-related pathways IL-6-STAT3 and IL-6-MEK5-ERK5, and the calcineurin-NFAT3 hypertrophy-related pathways were more active in obese Zucker rats, which may provide possible hypertrophic mechanisms for developing cardiac hypertrophy and pathological changes in obesity.

Patients with liver cirrhosis also have subtle cardiac structure or function abnormalities. This cardiac dysfunction commonly occurs in 56% of waiting orthotopic liver transplantation (OLT) patients and is defined as cirrhotic cardiomyopathy (CCM). Up to now, there is no standard treatment because CCM does not have a solidly established diagnosis and is based on high clinical suspicion. The liver function of CCM is particularly limited, making patients vulnerable to more drug treatments. Here, we use silymarin (100 mg/kg/day), baicalein (30 mg/kg/day), San Huang Shel Shin Tang (SHSST, 30 mg/kg/day) and β-cyclodextrin modified SHSST (SHSSTc, 30 and 300 mg/kg/day) treatments for a CCl4-induced CCM rat model. The results show that silymarin, baicalein and SHSST treatments can only slightly reduce the collagen accumulation in CCM rat hearts. However, SHSSTc treatment protects the heart in CCM and significantly inhibits collagen acumination and the fibrosis regulating transforming growth factor-β (TGF-β) pathway expression. SHSSTc treatments further reduced the heart weight and the ratio between left ventricular weight (LVW) and tibia length (TL). This experimental data show that water solubility improved β-cyclodextrin modified Chinese herbal medicine formula (SHSSTc) can provide an excellent heart protection effect through TGF-β pathway inhibition.

Statistical and clinical reports indicate that betel nut chewing is strongly associated with progression of oral cancer because some ingredients in betel nuts are potential cancer promoters, especially arecoline. Early diagnosis for cancer biomarkers is the best strategy for prevention of cancer progression. Several methods are suggested for investigating cancer biomarkers. Among these methods, gel-based proteomics approach is the most powerful and recommended tool for investigating biomarkers due to its high-throughput. However, this proteomics approach is not suitable for screening biomarkers with molecular weight under 10 KDa because of the characteristics of gel electrophoresis.This study investigated biomarkers with molecular weight under 10 KDa in rats with arecoline challenge.The centrifuging vials with membrane (10 KDa molecular weight cut-off) played a crucial role in this study. After centrifuging, the filtrate (containing compounds with molecular weight under 10 KDa) was collected and spotted on a sample plate for MALDI-TOF mass spectrometry analysis.Compared to control, three extra peaks (m/z values were 1553.1611, 1668.2097 and 1740.1832, respectively) were found in sera and two extra peaks were found in heart tissue samples (408.9719 and 524.9961, respectively). These small compounds should play important roles and may be potential biomarker candidates in rats with arecoline.This study successfully reports a mass-based method for investigating biomarker candidates with small molecular weight in different types of sample (including serum and tissue). In addition, this reported method is more time-efficient (1 working day) than gel-based proteomics approach (5~7 working days).

Inflammation plays an important role in triggering fibrosis of cardiovascular disease and hypertension. Gamma-aminobutyric acid (GABA) has hypotensive effect; GABA concentration could be enhanced in milk fermented with lactic acid bacteria (LAB). This study evaluated the effect of probiotic-fermented purple sweet potato yogurt (PSPY) on the toll-like receptor 4 (TLR-4)-related inflammatory components, and on fibrosis in the heart of spontaneously hypertensive rat (SHR). TLR4-related pathway and fibrosis-associated proteins TGFbeta and FGF2 were significantly increased in SHR hearts, but were highly suppressed in 10% PSPY-fed rats. Microscopic examination with Masson trichrome staining of left ventricle further demonstrated that 10% and 100% PSPY both significantly reduced interstitial fibrosis in SHR hearts. These findings indicated that oral administration of 10% probiotic-fermented PSPY was strong enough to lower cardiac fibrosis in SHR rats through the suppression of TLR-4-related inflammatory pathway. Therefore, PSPY may be included in diets to help prevent cardiac fibrosis in patients with hypertension.

Aging, a natural biological/physiological phenomenon, is accelerated by reactive oxygen species (ROS) accumulation and identified by a progressive decrease in physiological function. Several studies have shown a positive relationship between aging and chronic heart failure (HF). Cardiac apoptosis was found in age-related diseases. We used a traditional Chinese medicine, Alpinate Oxyphyllae Fructus (AOF), to evaluate its effect on cardiac anti-apoptosis and pro-survival. Male eight-week-old Sprague–Dawley (SD) rats were segregated into five groups: normal control group (NC), d-Galactose-Induced aging group (Aging), and AOF of 50 (AL (AOF low)), 100 (AM (AOF medium)), 150 (AH (AOF high)) mg/kg/day. After eight weeks, hearts were measured by an Hematoxylin–Eosin (H&E) stain, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-assays and Western blotting. The experimental results show that the cardiomyocyte apoptotic pathway protein expression increased in the d-Galactose-Induced aging groups, with dose-dependent inhibition in the AOF treatment group (AL, AM, and AH). Moreover, the expression of the pro-survival p-Akt (protein kinase B (Akt)), Bcl-2 (B-cell lymphoma 2), anti-apoptotic protein (Bcl-xL) protein decreased significantly in the d-Galactose-induced aging group, with increased performance in the AOF treatment group with levels of p-IGFIR and p-PI3K (Phosphatidylinositol-3′ kinase (PI3K)) to increase by dosage and compensatory performance. On the other hand, the protein of the Sirtuin 1 (SIRT1) pathway expression decreased in the aging groups and showed improvement in the AOF treatment group. Our results suggest that AOF strongly works against ROS-induced aging heart problems.

OBJECTIVE To clarify the efficacy of Taohe Chengqi Tang (THCQT), a compound traditional Chinese herbal medicine, in protecting liver damage induced by carbon tetrachloride (CCl(4)) in rats. METHODS Thirty male Wistar rats were divided into normal control group, untreated group, low-dose THCQT group (receiving 0.3 g/kg of THCQT), high-dose THCQT group (receiving 0.5 g/kg of THCQT), and positive control group (receiving silymarin 25 mg/kg). All testing substances were orally administered 1 hour before the intraperitoneal injection of CCl(4) (1.5 mL/kg). Twenty-four hours after CCl(4) injection, the rats were sacrificed to observe liver histopathological changes, and to evaluate activities of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and lipid peroxidation (LPO) and glutathione (GSH) levels in liver tissues. RESULTS CCl(4) injection elevated the serum AST and ALT activities, but THCQT significantly reversed this effect. The increase of hepatic LPO by CCl(4) was markedly reduced by THCQT. Also, this herbal mixture increased hepatic GSH in the rats. In histopathology analysis, THCQT decreased the fatty accumulation, necrosis and lymphocyte infiltration. The in vitro study in rat brain showed that LPO induced by Fe(2+)/ascorbic acid was dose-dependently reduced by THCQT. According to the biochemical and morphological data, THCQT could protect the liver from CCl(4-)induced injuries. CONCLUSION THCQT seems helpful for protection of liver damage induced by chemicals depending on its anti-oxidant-like function, and THCQT is more effective than silymarin.

It is unclear whether cardiac hypertrophy and hypertrophy-related pathways will be induced by long-term intermittent hypoxia. Thirty-six Sprague-Dawley rats were randomly assigned into three groups: normoxia, and long-term intermittent hypoxia (12% O(2), 8 h per day) for 4 weeks (4WLTIH) or for 8 weeks (8WLTIH). Myocardial morphology, trophic factors and signalling pathways in the three groups were determined by heart weight index, histological analysis, Western blotting and reverse transcriptase-polymerase chain reaction from the excised left ventricle. The ratio of whole heart weight to body weight, the ratio of left ventricular weight to body weight, the gross vertical cross-section of the heart and myocardial morphological changes were increased in the 4WLTIH group and were further augmented in the 8WLTIH group. In the 4WLTIH group, tumour necrosis factor-alpha(TNFalpha), insulin-like growth factor (IGF)-II, phosphorylated p38 mitogen-activated protein kinase (P38), signal transducers and activators of transcription (STAT)-1 and STAT-3 were significantly increased in the cardiac tissues. However, in the 8WLTIH group, in addition to the above factors, interleukin-6, mitogen-activated protein kinase (MEK)5 and extracellular signal-regulated kinase (ERK)5 were significantly increased compared with the normoxia group. We conclude that cardiac hypertrophy associated with TNFalpha and IGF-II was induced by intermittent hypoxia. The longer duration of intermittent hypoxia further activated the eccentric hypertrophy-related pathway, as well as the interleukin 6-related MEK5-ERK5 and STAT-3 pathways, which could result in the development of cardiac dilatation and pathology.

Background It is unclear whether the cardiac mitochondrial dependent apoptotic pathways and Fas death receptor dependent apoptotic pathways will be induced by long-term intermittent hypoxia. Methods Twenty-seven Sprague-Dawley rats were randomly assigned into three groups: normoxia, long-term intermittent hypoxia (12% O 2 , 8 h/day) for 4 weeks (4WLTIH) and for 8 weeks (8WLTIH). Histological analysis, Western blotting and RT-PCR in the three groups were performed on tissue from the excised left ventricle. Results Mitochondrial dependent pro-apoptotic pathway, BNIP3, caspase 9, and caspase 3, and the Fas death receptor dependent pro-apoptotic pathway, Fas, caspase 8, and caspase 3 were all significantly increased after 4WLTIH and even further enhanced after 8WLTIH. In addition, mitochondrial related anti-apoptotic proteins, Bcl2, its upstream phosphorylated protein kinase B (Akt), and the mitochondrial key oxidative enzyme, cytochrome c oxidase, were all decreased after 4WLTIH and further reduced after 8WLTIH. Conclusions The mitochondrial dependent apoptotic pathways and Fas death receptor dependent apoptotic pathways in rat hearts were both activated by long-term intermittent hypoxia.

Background: Aging is associated with physiological or pathological left ventricular hypertrophy (LVH) cardiac changes. Secondhand smoke (SHS) exposure is associated with pathological LVH. The action mechanism in cardiac concentric hypertrophy from SHS exposure is understood, but the transition contributed from SHS exposure is not. To determine whether exposure to SHS has an impact on age-induced LVH we examined young and old hamsters that underwent SHS exposure in a chamber for 30 mins. Methods: Morphological and histological studies were then conducted using hematoxylin and eosin (H&E) and Masson’s trichrome staining. Echocardiographic analysis was used to determine left ventricular wall thickness and function. LVH related protein expression levels were detected by western blot analysis. Results: The results showed that both young and aged hamsters exposed to SHS exhibited increased heart weights and left ventricular weights, left ventricular posterior wall thickness and intraventricular septum systolic and diastolic pressure also increased. However, left ventricular function systolic and diastolic pressure deteriorated. H&E and Masson’s trichrome staining results showed LV papillary muscles were ruptured, resulting in lower cardiac function at the myocardial level. LV muscle fiber arrangement was disordered and collagen accumulation occurred. Concentric LVH related protein molecular markers increased only in young hamsters exposed to SHS. However, this declined with hamster age. By contrast, eccentric LVH related proteins increased in aging hamsters exposed the SHS. Pro-inflammatory proteins, IL-6, TNF-α, JAK1, STAT3, and SIRTI expression increased in aging hamsters exposed to SHS. Conclusions: We suggest that SHS exposure induces a pro-inflammatory response that results in concentric transition to aging eccentric LVH.