Brucea javanica Oil Induces Apoptosis in T24 Bladder Cancer Cells via Upregulation of Caspase-3, Caspase-9, and Inhibition of NF-κB and COX-2 Expressions

2010 ◽  
Vol 38 (03) ◽  
pp. 613-624 ◽  
Author(s):  
Guo-Guang Lou ◽  
Hang-Ping Yao ◽  
Li-Ping Xie
Keyword(s):  
Caspase 3 ◽  
Flow Cytometric Analysis ◽  
Morphological Characteristics ◽  
Control Group ◽  
Caspase 9 ◽  
Brucea Javanica ◽  
Flow Cytometric ◽  
T24 Cells ◽  
Bladder Cancer Cells ◽  
Cox 2

The potential molecular mechanism of Brucea javanica oil in the induction of apoptosis of T24 bladder cancer cells was investigated in vitro. T24 cells were divided into two groups: one, treated with B. javanica oil and the other, untreated. The cells were maintained in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal calf serum (FCS) and 4 mM glutamine. The morphological characteristics of T24 cells were examined microscopically at the 2nd and 5th day of the culture. The drug toxicity spectrum ( IC 50) was estimated by the MTT assay, and viability of T24 cells was assessed on the basis of the percentage of T24 apoptotic cells, as determined by Annexin/PI staining and flow cytometric analysis. The expression of caspase-3, capase-9, NF-κB p65, and COX-2 was analyzed by Western blotting. Morphological characteristics of the cells on the 2nd day showed apoptosis of the treated T24 cells; it was more apparent in the cells on the 5th day. B. javanica oil decreased the cell viability at the testing concentrations spectrum (5–0.156 mg/ml), and this viability was significantly higher as compared to the control group. In this concentration spectrum, B. javanica oil also induced apoptosis of T24 cells, which was analyzed by annexin/PI staining and flow cytometric analysis. These results were also statistically significant as compared to those of the control group. The expressions of caspase-3 and caspase-9 were low in the control T24 cells, while the expressions of NF-κB and COX-2 were high in normal T24 cells. Treatment with B. javanica oil significantly induced the expressions of caspase-3 and caspase-9 proteins in T24 cells, whereas the expressions of NF-κB and COX-2 proteins were inhibited. B. javanica oil significantly reduced the viability of T24 cells and induced T24 cell apoptosis. The molecular mechanism underlying these effects may be the activation of caspase apoptotic pathway by upregulation of the expression of caspase-3 and caspase-9 proteins and inhibition of the expression of NF-κB and COX-2 proteins.

scholarly journals Achyranthes asperaRoot Extracts Induce Human Colon Cancer Cell (COLO-205) Death by Triggering the Mitochondrial Apoptosis Pathway and S Phase Cell Cycle Arrest

2014 ◽  
Vol 2014 ◽  
pp. 1-15 ◽  
Author(s):  
Shagun Arora ◽  
Simran Tandon
Keyword(s):  
Cell Cycle ◽  
Colon Cancer ◽  
Cell Cycle Arrest ◽  
Caspase 3 ◽  
Flow Cytometric Analysis ◽  
S Phase ◽  
Caspase 9 ◽  
Root Extracts ◽  
Cycle Arrest ◽  
Flow Cytometric

Achyranthes aspera(AA) has been used traditionally for the cure of various disorders. However, the action of root extracts of AA as anticancer agent and its cellular mechanism remain unclear. The aim was to screen the antitumor effect of ethanolic (EAA) and aqueous (AAA) root extracts on the growth of colon cancer COLO-205 cells by testing their cytotoxicity, followed by their effect on clonogenicity, migration, and induction of apoptosis. Mechanisms leading to apoptosis and cell cycle arrest were also investigated by expression studies of caspase-9, caspase-3, Bax, Bcl-2, p16, p21, and p27 genes, followed by flow cytometric analysis for cell cycle distribution. Cytotoxicity screening of AA extracts indicated greater cytotoxic activity of AAA extract against COLO-205 cells. A series of events marked by apoptosis revealed loss of cell viability, chromatin condensation, and DNA fragmentation in AAA treated cells to a greater extent. The mRNA expression levels of caspase-9, caspase-3, Bax, p16, p21, and p27 were markedly increased in the AAA treated cells, along with decreased Bcl-2 expression. The cell cycle arrest at S phase was detected by flow cytometric analysis after treatment with AAA. Overall the study signifies the aqueous extracts as a promising therapeutic candidate against cancer.

Apoptotic Effects of Melittin on 4T1 Breast Cancer Cell Line is associated with Up Regulation of Mfn1 and Drp1 mRNA Expression

2020 ◽  
Vol 20 (7) ◽  
pp. 790-799 ◽  
Author(s):  
Farnaz D. Moghaddam ◽  
Pejman Mortazavi ◽  
Somayeh Hamedi ◽  
Mohammad Nabiuni ◽  
Nasim H. Roodbari
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Hemolytic Activity ◽  
Breast Cancer Cells ◽  
Flow Cytometric Analysis ◽  
Control Group ◽  
Flow Cytometric ◽  
4T1 Cells ◽  
4T1 Breast Cancer ◽  
Apoptotic Effects

Background and Purpose: Melittin, as the main ingredient of honeybee venom, that has shown anticancer properties. The present study aimed at investigating the cytotoxic impacts of melittin on 4T1 breast cancer cells. Methods: Hemolytic activity of different concentrations (0.125, 0.25, 0.5, 1, 2, 4, 8μg/ml) of melittin was assayed and then cytotoxicity of selected concentrations of melittin (2, 4, 8, 16, 32, and 64μg/ml), 2 and 4μg/ml of cisplatin and 0.513, 0.295 and 0.123μg/ml of doxorubicin was evaluated on 4T1 cells using MTT assay. We used Morphological evaluation and flow cytometric analysis was used. Real time PCR was also used to determine mRNA expression of Mfn1 and Drp1 genes. Results: All compounds showed anti-proliferative effects on the tumor cell line with different potencies. Melittin had higher cytotoxicity against 4T1 breast cancer cells (IC50= 32μg/ml-72h) and higher hemolytic activity (HD50= 1μg/ml), as compared to cisplatin and doxorubicin. Mellitin at 16 and 32μg/ml showed apoptotic effects on 4T1 cells according to the flow cytometric analysis. The Real time PCR analysis of Drp1 and Mfn1 expression in cells treated with 16μg/ml of melittin revealed an up-regulation in Drp1 and Mfn1 genes mRNA expression in comparison with control group. Treatment with 32μg/ml of melittin was also associated with a rise in mRNA expression of Drp1 and Mfn1 as compared to the control group. Conclusion: The results of this study showed that melittin has anticancer effects on 4T1 cell lines in a dose and time dependent manner and can be a good candidate for further research on breast cancer treatment.

scholarly journals Silica nanoparticles inducing the apoptosis of spermatocyte cell through microRNA-450b-3p targeting MTCH2-mediating mitochondrial signaling pathway

2020 ◽  
Author(s):  
Guiqing Zhou ◽  
Jianhui Liu ◽  
Xiangyang Li ◽  
Yujian Sang ◽  
Yue Zhang ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Silica Nanoparticles ◽  
Caspase 3 ◽  
Reproductive Toxicity ◽  
Control Group ◽  
Caspase 9 ◽  
Protein Expressions ◽  
Underlying Mechanisms

Abstract Background: Silica nanoparticles (SiNPs) are found in environmental particulate matter and are proven to have adverse effects on fertility. The relationship and underlying mechanisms between miRNAs and apoptosis induced by SiNPs during spermatogenesis is currently ambiguous. Experimental design: The present study was designed to investigate the role of miRNA-450b-3p in the reproductive toxicity caused by SiNPs. In vivo, 40 male mice were randomly divided into control and SiNPs groups, 20 per group. The mice in the SiNPs group were administrated 20 mg/kg SiNPs by tracheal perfusion once every 5 days, for 35 days, and the control group were given the equivalent of a normal luminal saline. In vitro, spermatocyte cells were divided into 0 and 5 μg/mL SiNPs groups, after passaged for 30 generations, the GC-2spd cells in 5 μg/mL SiNPs groups were transfected with miRNA-450b-3p and its mimic and inhibitor. Results: In vivo, the results showed that SiNPs damaged tissue structures of testis, decreased the quantity and quality of the sperm, reduced the expression of miR-450b-3p, and increased the protein expressions of the MTCH2, BID, BAX, Cytochrome C, Caspase-9, and Caspase-3 in the testis. In vitro, SiNPs obviously repressed the viability and increased the LDH level and apoptosis rate, decreased the levels of the miR-450b-3p, significantly enhanced the protein expressions of the MTCH2, BID, BAX, Cytochrome C, Caspase-9, Caspase-3; while the mimic of miR-450b-3p reversed the changes induced by SiNPs, but inhibitor further promoted the effects induced by SiNPs.Conclusion: The result suggested that SiNPs could induce the spermatocyte apoptosis by inhibiting the miR-450b-3p expression to target promoting the MTCH2 resulting in activating mitochondrial apoptotic signaling pathways in the spermatocyte cells.

Flow cytometric analysis of DNA content in laryngeal cancer

1990 ◽  
Vol 104 (6) ◽  
pp. 485-487 ◽  
Author(s):  
Raphael Feinmesser ◽  
Jeremy L. Freeman ◽  
Arnold Noyek
Keyword(s):  
Squamous Cell Carcinoma ◽  
Dna Content ◽  
Laryngeal Squamous Cell Carcinoma ◽  
Flow Cytometric Analysis ◽  
Morphological Characteristics ◽  
Flow Cytometric ◽  
Dna Values ◽  
Laryngeal Tumours ◽  
N Stage ◽  
N Staging

AbstractDNA content was measured by flow-cytometric analysis in 30 paraffin embedded sections from patients with laryngeal squamous cell carcinoma. The morphological characteristics and N staging of the tumours as registered in their clinical charts were correlated with their DNA content. Eighty per cent of the tumours were found to have a predominantly aneuploid distribution of DNA values. There was no correlation between the N stage of the tumour or degree of cell differentiation and DNA content. A multiploid pattern correlates with non-metastatic laryngeal tumours and we suggest that this pattern may be a good indicator for biological activity.

scholarly journals Downregulation of CD4+LAP+ and CD4+CD25+ Regulatory T Cells in Acute Coronary Syndromes

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
Ying-zhong Lin ◽  
Shan-he Lu ◽  
Zheng-de Lu ◽  
Ying Huang ◽  
Ying Shi ◽  
...  
Keyword(s):  
Acute Coronary Syndromes ◽  
Flow Cytometric Analysis ◽  
Treg Cells ◽  
Protective Role ◽  
Control Group ◽  
Flow Cytometric ◽  
Coronary Syndromes ◽  
St Elevation ◽  
And Control ◽  
Elevation Acute Myocardial Infarction

Background. Regulatory T (Treg) cells play a protective role in atherosclerosis prone models and are related to the onset of acute coronary syndromes (ACS, including non-ST-elevation ACS (NSTEACS) and ST-elevation acute myocardial infarction (STEAMI)). CD4+LAP+ Treg cells are a novel subset of Tregs that have been found to ameliorate atherosclerosis inApoE-/-mice, and these cells also exist in humans. The present study was designed to investigate whether CD4+LAP+ Treg cells are involved in the onset of ACS.Methods. The frequencies of CD4+LAP+ and CD4+CD25+ Treg cells were detected using flow cytometric analysis, and the plasma IL-10 and TGF-β1 levels were measured using an ELISA in 29 stable angina (SA) patients, 30 NSTEACS patients, 27 STEAMI patients, and a control group (30 cases).Results. The results revealed a significant decrease in the frequencies of CD4+LAP+ and CD4+CD25+ Treg cells and in the levels of IL-10 and TGF-β1 in patients with ACS compared with those in the SA and control groups.Conclusions. The decrease in the frequencies of CD4+LAP+ and CD4+CD25+ Treg cells may play a role in the onset of ACS.

scholarly journals Molecular docking data of piperine with Bax, Caspase 3, Cox 2 and Caspase 9

2020 ◽  
Vol 16 (6) ◽  
pp. 458-461
Author(s):  
Rajagopal Ponnulakshmi ◽  
Keyword(s):  
Molecular Docking ◽  
Caspase 3 ◽  
Caspase 9 ◽  
Cox 2

scholarly journals The mitochondrial calcium uniporter induces apoptosis in cardiomyocytes cultured with high-glucose medium by affecting mitochondrial function

2020 ◽  
Author(s):  
Xia Chen ◽  
Wenyun Guo ◽  
Zhe Jing ◽  
Tao Zhang ◽  
Zhaoqi Wu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
High Glucose ◽  
Caspase 3 ◽  
Normal Group ◽  
Control Group ◽  
Caspase 9 ◽  
Mitochondrial Ros ◽  
H9c2 Cardiomyocytes ◽  
Experimental Group

Abstract Background As the number of diabetics worldwide continues to increase, diabetic cardiomyopathy has become one of the main causes of cardiovascular disease risk in diabetic patients. Currently, the pathophysiological mechanism of DCM has not been fully elucidated. In the present study, relevant pathological changes of cardiomyocytes in the high glucose environment were simulated by in vitro culture of rat H9C2 cardiomyocytes, to explore the mechanism by which MCU induces apoptosis in cardiomyocytes. Method: Cultured rat myocardium H9C2 cells in vitro and divided into high glucose group (glucose concentration 33 mmol/L), normal group (glucose concentration 5.5 mmol/L), experimental group (5.5 mmol/L glucose and transfected with MCU siRNA) and control group (5.5 mmol/L glucose and transfected negative control siRNA). Comparative analysis of MCU expression, Ca2+ uptake, mitochondrial function, oxidative stress and apoptosis of two groups of cells. Results (1) Compared with normal group, in the high glucose group the MCU expression of myocardial cells in H9C2 rats decreased, The Ca2+ levels, membrane potential and mitochondrial ATP levels decreased, mitochondrial ROS levels increased, NADH+/NADPH ratio in cardiomyocytes increased, GSH/GSSG ratio decreased, the expression levels of cleaved caspase-3 and cleaved caspase-9 increased, bcl-2 expression decreased, the number of cardiomyocytes apoptotic cells increases. (2) Compared with the normal group and the control group, the experimental group MCU expression of myocardial cells in H9C2 rats decreased, The Ca2+ levels, membrane potential and mitochondrial ATP levels decreased, mitochondrial ROS levels increased, NADH+/NADPH ratio in cardiomyocytes increased, GSH/GSSG ratio decreased, the expression levels of cleaved caspase-3 and cleaved caspase-9 increased, bcl-2 expression decreased, the number of cardiomyocytes apoptotic cells increases. Discussion This study suggested that MCU expression in rat H9C2 cardiomyocytes was decreased in the high glucose environment, causing abnormal mitochondrial calcium uptake and imbalanced calcium homeostasis, which may further contribute to mitochondrial dysfunction and enhanced oxidative stress in cardiomyocytes. Mitochondrial dysfunction and enhanced oxidative stress ultimately led to apoptosis in cardiomyocytes.

scholarly journals Neuroinflammation trajectories precede cognitive impairment after experimental meningitis—evidence from an in vivo PET study

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Vijayasree V. Giridharan ◽  
Allan Collodel ◽  
Jaqueline S. Generoso ◽  
Giselli Scaini ◽  
Rico Wassather ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cognitive Impairment ◽  
Microglial Activation ◽  
Caspase 3 ◽  
Cns Infection ◽  
Control Group ◽  
Caspase 9 ◽  
Activation Markers

Abstract Background Bacterial meningitis is a devastating central nervous system (CNS) infection with acute and long-term neurological consequences, including cognitive impairment. The aim of this study was to understand the association between activated microglia-induced neuroinflammation and post-meningitis cognitive impairment. Method Meningitis was induced in male Wistar rats by injecting Streptococcus pneumoniae into the brain through the cisterna magna, and rats were then treated with ceftriaxone. Twenty-four hours and 10 days after meningitis induction, rats were imaged with positron emission tomography (PET) using [11C]PBR28, a specific translocator protein (TSPO) radiotracer, to determine in vivo microglial activation. Following imaging, the expression of TSPO, cardiolipin, and cytochrome c, inflammatory mediators, oxidative stress markers, and glial activation markers were evaluated in the prefrontal cortex and hippocampus. Ten days after meningitis induction, animals were subjected to behavioral tests, such as the open-field, step-down inhibitory avoidance, and novel object recognition tests. Results Both 24-h (acute) and 10-day (long-term) groups of rats demonstrated increased [11C]PBR28 uptake and microglial activation in the whole brain compared to levels in the control group. Although free from infection, 10-day group rats exhibited increased expression levels of cytokines and markers of oxidative stress, microglial activation (IBA-1), and astrocyte activation (GFAP) similar to those seen in the 24-h group. Acute meningitis induction also elevated TSPO, cytochrome c, and caspase-3 levels with no change in caspase-9 levels. Furthermore, upregulated levels of TSPO, cytochrome c, and caspase-3 and caspase-9 were observed in the rat hippocampus 10 days after meningitis induction with a simultaneous reduction in cardiolipin levels. Animals showed a cognitive decline in all tasks compared with the control group, and this impairment may be at least partially mediated by activating a glia-mediated immune response and upregulating TSPO. Conclusions TSPO-PET could potentially be used as an imaging biomarker for microglial activation and long-term cognitive impairment post-meningitis. Additionally, this study opens a new avenue for the potential use of TSPO ligands after infection-induced neurological sequelae.

scholarly journals JNK pathway promotes hepatocyte apoptosis by inhibiting Bcl-2 and upregulating expressions of Bim, caspase-3 and caspase-9 after cardiopulmonary bypass

2020 ◽  
Vol 19 (3) ◽  
pp. 519-524
Author(s):  
Haibin Yu ◽  
Haojie Zhang ◽  
Yan Cheng ◽  
Xian’en Fa ◽  
Fangtao Zhu ◽  
...  
Keyword(s):  
Cardiopulmonary Bypass ◽  
Protein Expression ◽  
Caspase 3 ◽  
Control Group ◽  
Hepatocyte Apoptosis ◽  
Caspase 9 ◽  
Jnk Pathway ◽  
Expression Levels ◽  
Over Expression ◽  
Mrna Expression Levels

Purpose: To study the effect of Jun N-terminal kinase (JNK) signaling pathway on hepatocyte apoptosis in vivo and in vitro, and to elucidate the mechanism of action. Methods: TdT-mediated dUTP Nick-End Labeling (TUNEL) method was used to determine apoptosis in control and cardiopulmonary bypass (CPB) groups at 0, 3 and 6 hours after rat surgery. The expressions of JNK and p-c-Jun in liver tissues at 0, 3 and 6 h after surgery, and the levels of p-c-Jun, Bcl-2 and Bim following overexpression of JNK, were determined using Western blot assay. Human liver cell line HL-7702 was cultured and transfected with over-expressed JNK plasmid and empty plasmid. Proliferation of HL-7702 cells after JNK over-expression was assessed by Cell Counting Kit-8 (CCK-8), while quantitative real-time polymerase chain reaction (RT-qPCR) was employed to evaluate mRNA expression levels of caspase-3 and caspase-9 mRNA after JNK over-expression. Apoptosis of the cells was determined by flow cytometry (FC) after JNK over-expression. Results: FC results showed that the number of apoptotic hepatocytes increased after JNK overexpression in hepatocytes while TUNEL assay results demonstrated that hepatocyte apoptosis increased in CPB group, when compared to control group; furthermore, the number of apoptotic cells gradually increased within 6 h after surgery. The expressions of JNK and p-c-Jun were higher in CPB group than in control group, and increased gradually in both groups within 6 h after surgery. Overexpression of JNK decreased the proliferation of hepatocytes, and also lowered protein expression levels of p-c-Jun and Bim; on the other hand, the protein expression levels of Bcl-2 fell, while mRNA expression levels of caspase-3 and caspase-9 mRNA increased. Conclusion: JNK pathway promotes hepatocyte apoptosis after cardiopulmonary bypass by inhibiting Bcl-2 pathway and promoting the expressions of Bim caspase-3 and caspase-9. Keywords: Cardiopulmonary bypass, Apoptosis, JNK pathway, Bim, caspase-3 and caspase-9

Effects of subchronic exposure to carbendazim on spermatogenesis and fertility in male rats

2009 ◽  
Vol 25 (1) ◽  
pp. 41-47 ◽  
Author(s):  
G Yu ◽  
Q Guo ◽  
L Xie ◽  
Y Liu ◽  
X Wang
Keyword(s):  
Germ Cells ◽  
Flow Cytometric Analysis ◽  
Testicular Tissue ◽  
Male Rats ◽  
Seminiferous Tubules ◽  
Control Group ◽  
Testis Weight ◽  
Subchronic Exposure ◽  
Flow Cytometric ◽  
Sperm Counts

The aim of this study is to investigate the effects of subchronic exposure to carbendazim on spermatogenesis and fertility in male rats. Ninety-eight healthy male rats were divided into four groups: three exposure groups and a control group. Carbendazim was administered orally to male rats at 0, 20, 100 and 200 mg/kg for 80 days prior to mating. Each male was cohabited with an unexposed female for a maximum of 5 days. In 100 and 200 mg/kg groups, the mating index was relatively increased, the fertility index was decreased, and the testis weight, the sperm counts and motility were also decreased. The levels of luteinizing hormone (LH) showed a decreasing tendency and there was a statistical difference between the 200 mg/kg group and the control group. There were no obvious effects on the levels of follicle stimulating hormone (FSH) and testosterone (T). Histopathological evaluation showed atrophic seminiferous tubules, decreased germ cells, and increased sloughing of germ cells. Flow cytometric analysis of the testicular tissue revealed that carbendazim inhibited meiotic transformation and interfered with the spermatogenic process. These results suggest that carbendazim has adverse effects on spermatogenesis, resulting in reduced fertility in male rats.

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