Paeoniflorin protects myocardial cell from doxorubicin-induced apoptosis through inhibition of NADPH oxidase

Increased intracellular reactive oxygen species (ROS) are involved in doxorubicin (DOX)-induced myocardial cell apoptosis, and paeoniflorin (PEF) has been shown to exert an antioxidant effect. The aim of the present study was to explore the protective effect of PEF on DOX-induced myocardial cell apoptosis and the underlying mechanisms. In cultured H9c2 cells, different concentrations (1, 10, or 100 μmol/L) of PEF was added for 2 h prior to exposure to DOX (5 μmol/L) for 24 h. Cell apoptosis was evaluated by hoechst 33342 staining, and caspase-3 expression and activity. The mRNA and protein expression of NADPH oxidase (NOX) 2 and NOX4 was determined by real-time polymerase chain reaction and Western blot, respectively. Intracellular ROS and NOX activity were measured by assay kit. The results showed that DOX significantly increased myocardial cell apoptosis, increased caspase-3 expression and activity concomitantly with enhanced ROS production, and increased NOX2, NOX4 mRNA and protein expression, and NOX activity. These effects were remarkably inhibited by pretreatment of PEF. Our results suggested that PEF has a protective effect against DOX-induced myocardial cell apoptosis through a mechanism involving a decrease in ROS production by inhibition of NOX2, NOX4 expression, and NOX activity.

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Ultrasound-Targeted Microbubble Destruction-Mediated Inhibition of Livin Expression Accelerates Ovarian Cancer Cell Apoptosis

Genetics Research ◽

10.1155/2021/7624346 ◽

2021 ◽

Vol 2021 ◽

pp. 1-7

Author(s):

Xiaolin Xu ◽

Shuqin Yu ◽

Xiaoyuan Liu ◽

Ying Feng

Keyword(s):

Ovarian Cancer ◽

Protein Expression ◽

Cancer Cell ◽

Cell Apoptosis ◽

Caspase 3 ◽

Ovarian Cancer Cell ◽

Ovarian Cancer Cells ◽

Caspase 8 ◽

Expression Levels ◽

Mrna And Protein Expression

Objective. Ultrasound-targeted microbubble destruction (UTMD) technique has recently been developed as a nonviral delivery of gene therapy. This study aimed at investigating the survival and apoptosis of ovarian cancer cell line OVCA-433 by inhibiting Livin expression through ultrasound-targeted microbubble destruction. Methods. We synthesized a targeted microbubble agent for UTMD-mediated shRNA against Livin gene in human ovarian cancer OVCA-433 cells. Lipid microbubbles were conjugated with a luteinizing hormone-releasing hormone analog (LHRHa) by an avidin-biotin linkage to target the ovarian cancer OVCA-433 cells expressing LHRH receptors. The microbubbles were mixed with the recombinant plasmid harboring shRNA-Livin. shRNA-Livin was transfected into OVCA-433 cells upon exposure to 1 MHz pulsed ultrasound beam (0.5 W/cm2) for 8 s. Cell survival was measured by the MTT assay, cell apoptosis by flow cytometry using annexin V/PI double staining, and cell ultrastructure by using the transmission electron microscope. The mRNA and protein expression levels of caspase-3 and caspase-8 were detected by RT-qPCR and western blotting. Results. UTMD-mediated delivery of shRNA-Livin remarkably reduced the survival of OVCA-433 cells but promoted the apoptosis compared with shRNA-Livin alone, shRNA-Livin plus nontargeted microbubbles, and shRNA-Livin plus LHRHa-conjugated microbubbles containing shRNA-Livin with or without exposure to ultrasound pulses. It was also found that UTMD-mediated delivery of shRNA-Livin notably declined the mRNA and protein expression levels of caspase-3 and caspase-8 in OVCA-433 cells compared with shRNA-Livin alone, shRNA-Livin plus nontargeted microbubbles, and shRNA-Livin plus LHRHa-conjugated microbubbles containing shRNA-Livin with or without exposure to ultrasound pulses. Conclusion. Our experiment verifies the hypothesis that ultrasound mediation of targeted microbubbles can enhance the transfection efficiency of shRNA-Livin in ovarian cancer cells.

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Expression of apoptosis on rat liver by hepatic vagus hyperactivity after ventromedial hypothalamic lesioning

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.2001.280.5.g958 ◽

2001 ◽

Vol 280 (5) ◽

pp. G958-G967 ◽

Cited By ~ 11

Author(s):

Takayoshi Kiba ◽

Satoru Saito ◽

Kazushi Numata ◽

Yasuhiro Kon ◽

Tetsuya Mizutani ◽

...

Keyword(s):

Protein Expression ◽

Rat Liver ◽

Cholinergic System ◽

Caspase 3 ◽

Fas Ligand ◽

Ventromedial Hypothalamus ◽

Ligand System ◽

Fas Mrna ◽

Mrna And Protein Expression ◽

Induced Apoptosis

We examined whether the Fas (APO-1/CD95)/Fas ligand system mediates apoptosis in rats with ventromedial hypothalamus (VMH) lesions. Northern and Western blotting indicated that VMH lesions lead to a significant increase in Fas mRNA and protein expression from day 1 to day 7 and in Fas ligand mRNA and protein expression from day 2 to day 7. Immunohistochemistry indicated that the region of strongest Fas expression shifted from acinar zone 1 to zones 2 and 3 by day 7 after VMH lesioning and that at days 2–7Fas-ligand-positive hepatocyte cell membranes and cytoplasm were randomly distributed in acinar zones 1–3. We also analyzed activation of caspase 3-like proteases in hepatocytes, Kupffer cells, and sinusoidal endothelial cells. Spectrofluorometric assay demonstrated that caspase 3-like activity significantly increased only in hepatocytes after VMH lesioning. Moreover, electron microscopy and TUNEL assay showed that VMH lesions induced apoptosis. All of these effects were completely inhibited by hepatic vagotomy and administration of atropine. Vagal firing after VMH lesioning may stimulate Fas/Fas ligand system-mediated apoptosis through the cholinergic system in the rat liver.

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Resistance to apoptosis is a mechanism of adaptation of rat stomach to aspirin

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.2000.278.6.g839 ◽

2000 ◽

Vol 278 (6) ◽

pp. G839-G846 ◽

Cited By ~ 20

Author(s):

Barbara M. Alderman ◽

Gregory A. Cook ◽

Mary Familari ◽

Neville D. Yeomans ◽

Andrew S. Giraud

Keyword(s):

Protein Expression ◽

Caspase 3 ◽

Stress Protein ◽

Apoptotic Cells ◽

Rat Stomach ◽

Drug Induced ◽

Protein Levels ◽

Mrna And Protein Expression ◽

Mucosal Adaptation ◽

Induced Apoptosis

Adaptation of the gastric mucosa to nonsteroidal anti-inflammatory drug-induced injury is a well-documented phenomenon, but the mechanisms are not known. We investigated whether changes in stress protein expression and apoptosis play roles in adaptation of rat stomach to aspirin. RT-PCR and Western blotting techniques were used to analyze mRNA and protein expression of HSP72 and HSP90 and cleavage of caspase 3 protein. Apoptosis was detected by the TUNEL method and quantified. HSP72 mRNA and protein expression was unchanged in adapted mucosa, whereas HSP90 mRNA and protein levels decreased. Caspase 3 protein was activated, and the number of apoptotic cells increased in mucosa after one aspirin dose. However, in adapted mucosa after aspirin, activated caspase 3 and the number of apoptotic cells had returned to basal levels. Induction of the stress response was found not to be a mechanism of mucosal adaptation against multiple doses of aspirin. Our results lead us to propose instead that resistance to aspirin-induced apoptosis plays a role in the protective phenomenon of adaptation.

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Protective Effect of Two Alkaloids from Hippophae rhamnoides Linn. against Doxorubicin-Induced Toxicity in H9c2 Cardiomyoblasts

Molecules ◽

10.3390/molecules26071946 ◽

2021 ◽

Vol 26 (7) ◽

pp. 1946

Author(s):

Wenna Zhou ◽

Jian Ouyang ◽

Na Hu ◽

Gang Li ◽

Honglun Wang

Keyword(s):

Protein Expression ◽

Protective Effect ◽

Caspase 3 ◽

Cardiac Cells ◽

Potential Candidate ◽

H9c2 Cells ◽

Cardiac Muscle Cells ◽

Anti Cancer ◽

H9c2 Cardiomyoblasts ◽

Induced Apoptosis

Background: Doxorubicin (Dox) is one of the most frequently prescribed anti-cancer drugs. However, clinical application with Dox is limited due to its potentially fatal cumulative cardiotoxicity. N-p-coumaroyl-4-aminobutan-1-ol (alk-A), an organic amide alkaloid and hippophamide (alk-B), a rare pyridoindole alkaloid were successfully obtained by purification and separation of seabuckthorn seed residue in our previous research. This study was undertaken to investigate the protective effect of alk-A and alk-B against Dox-induced embryonic rat cardiac cells (H9c2 cells) apoptosis. Methods: H9c2 cells were treated with Dox (2.5 µM) in the presence of alk-A and alk-B (10, 20, and 40 µM) and incubated for 24 h. Results: It was shown that pretreatment of the H9c2 cells with alk-A and alk-B significantly reduced Dox-induced apoptosis. Alk-A and alk-B both inhibited reactive oxygen species (ROS) production and suppressed cleaved-caspase-3 protein expression and the activation of JNK (Jun N-terminal kinases), as well as increasing ATP levels, favoring mitochondrial mitofusin protein expression, and relieving damage to mitochondrial DNA. Conclusions: These results suggest that alk-A and alk-B can inhibit Dox-induced apoptosis in H9C2 cardiac muscle cells via inhibition of cell apoptosis and improvement of mitochondrial function, while alk-B showed more protection. Alk-B could be a potential candidate agent for protecting against cardiotoxicity in Dox-exposed patients.

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Ghrelin Protects against Dexamethasone-Induced INS-1 Cell Apoptosis via ERK and p38MAPK Signaling

International Journal of Endocrinology ◽

10.1155/2016/4513051 ◽

2016 ◽

Vol 2016 ◽

pp. 1-11 ◽

Cited By ~ 14

Author(s):

Chengshuo Zhang ◽

Le Li ◽

Bochao Zhao ◽

Ao Jiao ◽

Xin Li ◽

...

Keyword(s):

Protective Effect ◽

Cell Apoptosis ◽

Caspase 3 ◽

Islet Cells ◽

Dexamethasone Treatment ◽

Growth Hormone Secretagogue Receptor ◽

Growth Hormone Secretagogue ◽

Erk Activation ◽

P38mapk Signaling ◽

Induced Apoptosis

Glucocorticoid excess induces apoptosis of islet cells, which may result in diabetes. In this study, we investigated the protective effect of ghrelin on dexamethasone-induced INS-1 cell apoptosis. Our data showed that ghrelin (0.1 μM) inhibited dexamethasone-induced (0.1 μM) apoptosis of INS-1 cells and facilitated cell proliferation. Moreover, ghrelin upregulated Bcl-2 expression, downregulated Bax expression, and decreased caspase-3 activity. The protective effect of ghrelin against dexamethasone-induced INS-1 cell apoptosis was mediated via growth hormone secretagogue receptor 1a. Further studies revealed that ghrelin increased ERK activation and decreased p38MAPK expression after dexamethasone treatment. Ghrelin-mediated protection of dexamethasone-induced apoptosis of INS-1 cells was attenuated using the ERK inhibitor U0126 (10 μM), and cell viability increased using the p38MAPK inhibitor SB203580 (10 μM). In conclusion, ghrelin could protect against dexamethasone-induced INS-1 cell apoptosis, at least partially via GHS-R1a and the signaling pathway of ERK and p38MAPK.

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TPO exerts a protective effect on iron-overload induces apoptosis in cardiomyocytes via mitochondrial pathways

Blood ◽

10.1182/blood.v122.21.4668.4668 ◽

2013 ◽

Vol 122 (21) ◽

pp. 4668-4668 ◽

Cited By ~ 2

Author(s):

Mo Yang ◽

Shing Chan ◽

Jie yu Ye ◽

Godfrey ChiFung Chan

Keyword(s):

Oxidative Stress ◽

Iron Overload ◽

Protective Effect ◽

Mitochondrial Membrane ◽

Caspase 3 ◽

Ros Production ◽

H9c2 Cells ◽

Plot Analysis ◽

Iron Treatment ◽

Induced Apoptosis

Thalassaemia companied with iron-overload is common in Hong Kong. Iron overload induced cardiomyopathy is the commonest cause of morbidity and mortality in b-thalassaemia patients. One of the causes of cardiac failure is chronic iron overload of blood transfusion. Some studies showed that iron overload can cause toxic effect in heart cells. Iron-overload induced cardiomyopathy damages are the major complications in patients with beta-thalassaemia major. Iron-overload may induce apoptosis in cardiomyocytes. Our previous study showed TPO has cardiac protective effect (Li et al, Circulation, 2007). In this study, we demonstrated firstly that iron induced oxidative stress can cause apoptosis in cardiomyocytes. By using H9C2 cells, we showed that iron increased reactive oxygen species (ROS) production (n=3) and reduced cell viability in a dose-dependent manner (0-0.6 mM) (n=6). Apoptotic cells were found to be significantly increased under iron treatment (0.3 mM, 72 hrs) in the AnnexinV/PI assay (n=6). The expression of active caspase-3 significantly increased in iron-treated cells. Furthermore, iron treatment increased the proportion of cells containing JC-1 monomers, indicating a trend in the drop of mitochondrial membrane potential (n=6). Secondly, we found that TPO exerted cardio-protective effect on iron-induced apoptosis. H9C2 cells were cultured in the presence of iron (0.3 mM) with or without TPO (50 ng/mL). The ROS production was significantly decreased with the addition of TPO at 50 ng/mL (n=3). Dot-plot analysis of AnnexinV/PI staining demonstrated that TPO significantly reduced the population of apoptotic cells (n=6). Incubation with TPO also decreased the iron-induced caspase-3 expression (n=6). Flow cytometric dot-plot analysis also showed trends of amelioration of the increase in JC-1 monomers in the iron plus TPO group (n=6), indicating a trend in attenuation of the drop of mitochondrial membrane potential. Our findings suggest that iron-overload lead to generation of ROS which further induces apoptosis in cardiomyocytes via mitochondrial pathways and TPO might exert a protective effect on iron-overload induced apoptosis via inhibiting oxidative stress and mitochondrial pathway in cardiomyocytes. Disclosures: No relevant conflicts of interest to declare.

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Resistin is a survival factor for porcine ovarian follicular cells

Reproduction ◽

10.1530/rep-15-0255 ◽

2015 ◽

Vol 150 (4) ◽

pp. 343-355 ◽

Cited By ~ 17

Author(s):

Agnieszka Rak ◽

Eliza Drwal ◽

Anna Wróbel ◽

Ewa Łucja Gregoraszczuk

Keyword(s):

Cell Proliferation ◽

Protein Expression ◽

Western Blot ◽

Dna Fragmentation ◽

Cell Apoptosis ◽

Caspase 3 ◽

Inhibitory Effect ◽

Ovarian Cell ◽

Proliferation And Apoptosis ◽

Mrna And Protein Expression

Previously, we demonstrated the expression of resistin in the porcine ovary, the regulation of its expression and its direct effect on ovarian steroidogenesis. The objective of this study was to examine the effect of resistin on cell proliferation and apoptosis in a co-culture model of porcine granulosa and theca cells. First, we analysed the effect of resistin at 1 and 10 ng/ml alone or in combination with FSH- and IGF1 on ovarian cell proliferation with an alamarBlue assay and protein expression of cyclins A and B using western blot. Next, the mRNA and protein expression of selected pro-apoptotic and pro-survival regulators of cell apoptosis, caspase-9, -8 and -3 activity and DNA fragmentation using real time PCR, western blot, fluorescent assay and an ELISA kit, respectively, were analysed after resistin treatment. Furthermore, we determined the effect of resistin on the protein expression of ERK1/2, Stat and Akt kinase. Using specific inhibitors of these kinases, we also checked caspase-3 activity and protein expression. We found that resistin, at both doses, has no effect on cell proliferation. The results showed that resistin decreased pro-apoptotic genes, which was confirmed on protein expression of selected factors. We demonstrate an inhibitory effect of resistin on caspase activity and DNA fragmentation. Finally, resistin stimulated phosphorylation of the ERK1/2, Stat and Akt and kinases inhibitors reversed resistin action on caspase-3 activity and protein expression to control. All of these results showed that resistin has an inhibitory effect on porcine ovarian cell apoptosis by activation of the MAPK/ERK, JAK/Stat and Akt/PI3 kinase signalling pathways.

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NADPH Oxidase Hyperactivity Contributes to Cardiac Dysfunction and Apoptosis in Rats with Severe Experimental Pancreatitis through ROS-Mediated MAPK Signaling Pathway

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/4578175 ◽

2019 ◽

Vol 2019 ◽

pp. 1-18 ◽

Cited By ~ 7

Author(s):

Yi Wen ◽

Ruohong Liu ◽

Ning Lin ◽

Hao Luo ◽

Jiajia Tang ◽

...

Keyword(s):

Oxidative Stress ◽

Nadph Oxidase ◽

Cardiac Function ◽

Protein Expression ◽

Cell Apoptosis ◽

Cardiac Injury ◽

Sodium Taurocholate ◽

Mapk Signaling ◽

Signaling Molecules ◽

Ros Production

NADPH oxidase (Nox) is considered a major source of reactive oxygen species (ROS) in the heart in normal and pathological conditions. However, the role of Nox in severe acute pancreatitis- (SAP-) associated cardiac injury remains unclear. Therefore, we aim to investigate the contribution of Nox to SAP-associated cardiac injury and to explore the underlying molecular mechanisms. Apocynin, a Nox inhibitor, was given at 20 mg/kg for 30 min before SAP induction by a retrograde pancreatic duct injection of 5% sodium taurocholate. Histopathological staining, Nox activity and protein expression, oxidative stress markers, apoptosis and associated proteins, cardiac-related enzyme indexes, and cardiac function were assessed in the myocardium in SAP rats. The redox-sensitive MAPK signaling molecules were also examined by western blotting. SAP rats exhibited significant cardiac impairment along with increased Nox activity and protein expression, ROS production, cell apoptosis, and proapoptotic Bax and cleaved caspase-3 protein levels. Notably, Nox inhibition with apocynin prevented SAP-associated cardiac injury evidenced by a decreased histopathologic score, cardiac-related enzymes, and cardiac function through the reduction of ROS production and cell apoptosis. This protective role was further confirmed by a simulation experiment in vitro. Moreover, we found that SAP-induced activation in MAPK signaling molecules in cardiomyocytes was significantly attenuated by Nox inhibition. Our data provide the first evidence that Nox hyperactivation acts as the main source of ROS production in the myocardium, increases oxidative stress, and promotes cell apoptosis via activating the MAPK pathway, which ultimately results in cardiac injury in SAP.

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Artemisinin attenuated oxidative stress and apoptosis by inhibiting autophagy in MPP+-treated SH-SY5Y cells

Journal of Biological Research-Thessaloniki ◽

10.1186/s40709-021-00137-6 ◽

2021 ◽

Vol 28 (1) ◽

Author(s):

Junqiang Yan ◽

Hongxia Ma ◽

Xiaoyi Lai ◽

Jiannan Wu ◽

Anran Liu ◽

...

Keyword(s):

Oxidative Stress ◽

Membrane Potential ◽

Protein Expression ◽

Mitochondrial Membrane Potential ◽

Western Blotting ◽

Cell Apoptosis ◽

Mitochondrial Membrane ◽

Caspase 3 ◽

Neuroprotective Effect ◽

Neuroprotective Effects

Abstract Background Parkinson’s disease (PD) is the second most common neurodegenerative disease after Alzheimer's disease. The oxidative stress is an important component of the pathogenesis of PD. Artemisinin (ART) has antioxidant and neuroprotective effects. The purpose of this study is to explore the neuroprotective effect of ART on 1-methyl-4-phenyliodine iodide (MPP +)-treated SH-SY5Y cells and underlying mechanism. Methods We used MPP+-treated SH-SY5Y cells to study the neuroprotective effect of ART. Cell viability was measured by MTT assay after incubating the cells with MPP+ and/or ART for 24 h. DCFH-DA was used to detect the level of intracellular reactive oxygen species (ROS), and WST-8 was used to detect the level of superoxide dismutase (SOD). The level of intracellular reduced glutathione (GSH) was detected with 5,5΄-dithiobis-(2-nitrobenzoic acid), and the level of malondialdehyde (MDA) was assessed based on the reaction of MDA and thiobarbituric acid. A mitochondrial membrane potential detection kit (JC-1) was used to detect changes in the mitochondrial membrane potential (MMP), and an Annexin V-FITC cell apoptosis kit was used to detect cell apoptosis. The expression levels of caspase-3, cleaved caspase-3 and the autophagy-related proteins LC3, beclin-1, and p62 were detected by Western blotting. In addition, to verify the change in autophagy, we used immunofluorescence to detect the expression of LC3 and p62. Results No significant cytotoxicity was observed at ART concentrations up to 40 μM. ART could significantly increase the viability of SH-SY5Y cells treated with MPP+ and reduce oxidative stress damage and apoptosis. In addition, the Western blotting and immunofluorescence results showed that MPP+ treatment could increase the protein expression of beclin1 and LC3II/LC3I and decrease the protein expression of p62, indicating that MPP+ treatment could induce autophagy. Simultaneous treatment with ART and MPP+ could decrease the protein expression of beclin1 and LC3II/LC3I and increase the protein expression of p62, indicating that ART could decrease the level of autophagy induced by MPP+. Conclusion Our results indicate that ART has a protective effect on MPP+-treated SH-SY5Y cells by the antioxidant, antiapoptotic activities and inhibition of autophagy. Our findings may provide new hope for the prevention and treatment of PD.

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Neuroprotective Effects of Alpha-Mangostin on MPP+-Induced Apoptotic Cell Death in Neuroblastoma SH-SY5Y Cells

Journal of Toxicology ◽

10.1155/2015/919058 ◽

2015 ◽

Vol 2015 ◽

pp. 1-11 ◽

Cited By ~ 21

Author(s):

Prachya Janhom ◽

Permphan Dharmasaroja

Keyword(s):

Cell Death ◽

Cell Viability ◽

Caspase 3 ◽

Nuclear Dna ◽

Apoptotic Cell ◽

Apoptotic Cell Death ◽

Concomitant Treatment ◽

Ros Production ◽

Cellular Models ◽

Induced Apoptosis

In vitrostudies have shown that extracts from mangosteen (Garcinia mangostanaLinn.) act as antioxidants and cytoprotective agents against oxidative damage. The protective effect of alpha-mangostin, the major xanthone found in the pericarp of the mangosteen, in cellular models of Parkinson’s disease (PD), has not been investigated. This study aims to investigate whether alpha-mangostin could protect SH-SY5Y neuroblastoma cells from MPP+-induced apoptosis. The effects of alpha-mangostin on MPP+-induced cell death were evaluated with a cell viability assay, staining for nuclear DNA morphology, flow cytometry for apoptotic cells and reactive oxygen species (ROS) production, quantitative real-time PCR for the expression of p53, Bax, and Bcl-2, and western blot analysis for cleaved caspase-3. Concomitant treatment with alpha-mangostin attenuated the effect of MPP+on cell viability and apoptotic cell death. Alpha-mangostin reduced ROS formation induced by MPP+. Bax/Bcl-2 expression ratio and expression of p53 were significantly lower in cells cocultured with alpha-mangostin and MPP+. The cotreated cells showed a significant decrease in activated caspase-3 compared with MPP+treatment alone. Our data suggest that cytoprotection of alpha-mangostin against MPP+-induced apoptosis may be associated with the reduction of ROS production, modulating the balance of pro- and antiapoptotic genes, and suppression of caspase-3 activation.

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