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Benavides RAS, Leiro-Vidal JM, Rodriguez-Gonzalez JA, Ares-Pena FJ, López-Martín E. The HL-60 human promyelocytic cell line constitutes an effective in vitro model for evaluating toxicity, oxidative stress and necrosis/apoptosis after exposure to black carbon particles and 2.45 GHz radio frequency. THE SCIENCE OF THE TOTAL ENVIRONMENT 2023; 867:161475. [PMID: 36632900 DOI: 10.1016/j.scitotenv.2023.161475] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/12/2022] [Revised: 12/21/2022] [Accepted: 01/04/2023] [Indexed: 06/17/2023]
Abstract
The cellular and molecular mechanisms by which atmospheric pollution from particulate matter and/or electromagnetic fields (EMFs) may prove harmful to human health have not been extensively researched. We analyzed whether the combined action of EMFs and black carbon (BC) particles induced cell damage and a pro-apoptotic response in the HL-60 promyelocytic cell line when exposed to 2.45 GHz radio frequency (RF) radiation in a gigahertz transverse electromagnetic (GTEM) chamber at sub-thermal specific absorption rate (SAR) levels. RF and BC induced moderately significant levels of cell damage in the first 8 or 24 h for all exposure times/doses and much greater damage after 48 h irradiation and the higher dose of BC. We observed a clear antiproliferative effect that increased with RF exposure time and BC dose. Oxidative stress or ROS production increased with time (24 or 48 h of radiation), BC dose and the combination of both. Significant differences between the proportion of damaged and healthy cells were observed in all groups. Both radiation and BC participated separately and jointly in triggering necrosis and apoptosis in a programmed way. Oxidative-antioxidant action activated mitochondrial anti-apoptotic BCL2a gene expression after 24 h irradiation and exposure to BC. After irradiation of the cells for 48 h, expression of FASR cell death receptors was activated, precipitating the onset of pro-apoptotic phenomena and expression and intracellular activity of caspase-3 in the mitochondrial pathways, all of which can lead to cell death. Our results indicate that the interaction between BC and RF modifies the immune response in the human promyelocytic cell line and that these cells had two fates mediated by different pathways: necrosis and mitochondria-caspase dependent apoptosis. The findings may be important in regard to antimicrobial, inflammatory and autoimmune responses in humans.
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Affiliation(s)
- Rosa Ana Sueiro Benavides
- Institute of Research in Biological and Chemical Analysis, IAQBUS, University of Santiago de Compostela, Santiago de Compostela, Spain.
| | - José Manuel Leiro-Vidal
- Institute of Research in Biological and Chemical Analysis, IAQBUS, University of Santiago de Compostela, Santiago de Compostela, Spain.
| | - J Antonio Rodriguez-Gonzalez
- Department of Applied Physics, Santiago de Compostela School of Physics, University of Santiago de Compostela, Santiago de Compostela, Spain.
| | - Francisco J Ares-Pena
- Department of Applied Physics, Santiago de Compostela School of Physics, University of Santiago de Compostela, Santiago de Compostela, Spain.
| | - Elena López-Martín
- Department of Morphological Sciences, Santiago de Compostela School of Medicine, University of Santiago de Compostela, Santiago de Compostela, Spain.
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Shi C, Han W, Zhang M, Zang R, Du K, Li L, Xu X, Li C, Wang S, Qiu P, Guan H, Yang J, Xiao S, Wang X. Sulfated polymannuroguluronate TGC161 ameliorates leukopenia by inhibiting CD4 + T cell apoptosis. Carbohydr Polym 2020; 247:116728. [PMID: 32829850 PMCID: PMC7336955 DOI: 10.1016/j.carbpol.2020.116728] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2020] [Revised: 06/23/2020] [Accepted: 07/02/2020] [Indexed: 02/07/2023]
Abstract
Polysaccharides have aroused considerable interest due to their diverse biological activities and low toxicity. In this study, we evaluated the effect of marine polysaccharide sulfated polymannuroguluronate (TGC161) on the leukopenia induced by chemotherapy. It is found that TGC161 ameliorates the leukopenia. Besides, TGC161 would promote CD4+ T cell differentiation and maturation in the thymus, but does not have a significant effect on precursor cells in bone marrow. Furthermore, TGC161 inhibits CD4+ T cell apoptosis in vitro. Collectively, our findings offer a natural and harmless polysaccharide to ameliorate leukopenia.
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Affiliation(s)
- Chuanqin Shi
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China
| | - Wenwei Han
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China
| | - Meifang Zhang
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China
| | - Ruochen Zang
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China
| | - Kaixin Du
- Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Li Li
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Ximing Xu
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Chunxia Li
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Shixin Wang
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Peiju Qiu
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Huashi Guan
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Jinbo Yang
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China
| | - Shuai Xiao
- Department of Gastrointestinal Surgery and Institute of Clinical Medicine, the First Affiliated Hospital, University of South China, Hengyang, 421001, China.
| | - Xin Wang
- Key Laboratory of Marine Drugs of Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China; Center for Innovation Marine Drug Screening & Evaluation, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China; Marine Biomedical Research Institute of Qingdao, Qingdao, 266071, China.
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Lv C, Hao Y, Tu G. MicroRNA-21 promotes proliferation, invasion and suppresses apoptosis in human osteosarcoma line MG63 through PTEN/Akt pathway. Tumour Biol 2016; 37:9333-42. [PMID: 26779632 DOI: 10.1007/s13277-016-4807-6] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2015] [Accepted: 01/06/2016] [Indexed: 01/04/2023] Open
Abstract
Osteosarcoma, which accounts for 5 % of pediatric tumor, remains the major cause of death among orthopedic malignancies. However, the factors associated with its malignant biological behavior are still poorly understood. MicroRNAs are a class of small noncoding RNAs, which have been considered to associate with malignant progression including cell differentiation, proliferation, apoptosis, invasion, and distant metastasis. In our research, we found that microRNA-21 (miR-21) was significantly overexpressed in human osteosarcoma cell line MG63 compared to human fetal osteoblastic cell line hFOB1.19 by using quantitative real-time polymerase chain reaction (qRT-PCR). Moreover, miR-21 overexpression in MG63 caused a significant raise in cell proliferation and invasion and a significant reduction in cell apoptosis. However, miR-21 underexpression in MG63 caused an opposite result. Western blotting displayed that proteins related with proliferation, apoptosis, and invasion were significantly changed in different groups, respectively. Furthermore, we demonstrated that PTEN may be a potential target of miR-21 in MG63 cells and miR-21 could activate PI3K/Akt pathway by suppressing PTEN expression. In summary, our findings suggested that miR-21 played an active role in osteosarcoma and it could predict the occurrence and development of osteosarcoma.
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Affiliation(s)
- Chen Lv
- Department of Orthopedics, The First Affiliated Hospital of China Medical University, No. 155 Nanjingbei Street, Heping District, Shenyang, 110001, China
| | - Yuehan Hao
- Department of Neurology, The First Affiliated Hospital of China Medical University, Shenyang, 110001, Liaoning, China
| | - Guanjun Tu
- Department of Orthopedics, The First Affiliated Hospital of China Medical University, No. 155 Nanjingbei Street, Heping District, Shenyang, 110001, China.
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Xu P, Jiang EJ, Wen SY, Lu DD. Amentoflavone acts as a radioprotector for irradiated v79 cells by regulating reactive oxygen species (ROS), cell cycle and mitochondrial mass. Asian Pac J Cancer Prev 2015; 15:7521-6. [PMID: 25292022 DOI: 10.7314/apjcp.2014.15.18.7521] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022] Open
Abstract
Radioprotective effects of amentoflavone were investigated by examining cell viability, apoptosis, cell cycling concentrations of intracellular ROS (reactive oxygen species), and relative mitochondrial mass by flow cytometry after 60Co irradiation. Pretreatment with amentoflavone 24 hours prior to 8 Gy 60Co γ-ray irradiation significantly inhibited apoptosis, promoted the G2 phase, decreased the concentration of ROS and mitochondrial mass. These results collectively indicate that amentoflavone is an effective radioprotective agent.
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Affiliation(s)
- Ping Xu
- Department of Pharmacy, Xinxiang Medical University, Xinxiang, Henan, China E-mail :
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Senthamizhselvan O, Manivannan J, Silambarasan T, Raja B. Diosmin pretreatment improves cardiac function and suppresses oxidative stress in rat heart after ischemia/reperfusion. Eur J Pharmacol 2014; 736:131-7. [PMID: 24769512 DOI: 10.1016/j.ejphar.2014.04.026] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2014] [Revised: 04/10/2014] [Accepted: 04/16/2014] [Indexed: 12/31/2022]
Abstract
Reperfusion of ischemic tissue leads to the generation of oxygen derived free radicals which plays an important role in cellular damage. Objective of the current study is to evaluate the cardio-protective and antioxidant effect of diosmin on ischemia-reperfusion related cardiac dysfunction, oxidative stress and apoptosis. Diosmin (50 and 100 mg/kg body weight (bw)) was given every day to the rats orally throughout the experimental period. Ischemia/reperfusion protocol was carried out ex vivo using langendorff perfusion method and the cardiac functional recovery was assessed in terms of percentage rate pressure product. Coronary effluents of LDH and CK-MB activities, antioxidant enzyme activities, lipid peroxidation products, activity of TCA cycle enzymes were evaluated. Moreover, in vitro superoxide anion and hydroxyl radical scavenging potential of diosmin was also quantified. Finally, quantitative real-time PCR was used for assessing Bcl-2 mRNA expression in heart. Cardiac functional recovery was impaired after reperfusion compared with continuously perfused heart. It was significantly prevented by diosmin treatment. Impaired antioxidant enzyme activities and elevated lipid peroxidation products level were also significantly suppressed. The activity of TCA cycle enzymes was protected against reperfusion stress. Down regulated Bcl-2 was also significantly increased. This study concluded that diosmin pretreatment prevents all the impaired patterns including cardiac function, oxidative stress and apoptosis associated with reperfusion in control heart by its antioxidant role.
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Affiliation(s)
- Oomaidurai Senthamizhselvan
- Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalai Nagar 608 002, Tamil Nadu, India
| | - Jeganathan Manivannan
- Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalai Nagar 608 002, Tamil Nadu, India
| | - Thangarasu Silambarasan
- Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalai Nagar 608 002, Tamil Nadu, India
| | - Boobalan Raja
- Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalai Nagar 608 002, Tamil Nadu, India.
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Bavithra S, Selvakumar K, Krishnamoorthy G, Venkataraman P, Arunakaran J. Melatonin attenuates polychlorinated biphenyls induced apoptosis in the neuronal cells of cerebral cortex and cerebellum of adult male rats--in vivo. ENVIRONMENTAL TOXICOLOGY AND PHARMACOLOGY 2013; 36:152-163. [PMID: 23619521 DOI: 10.1016/j.etap.2013.03.005] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/05/2012] [Revised: 03/01/2013] [Accepted: 03/08/2013] [Indexed: 06/02/2023]
Abstract
Polychlorinated biphenyls (PCBs) are widespread persistent environmental contaminants that display a complex spectrum of toxicological properties, including neurotoxicity. Studies have shown that PCBs increase oxidative stress in brain, leading to apoptosis. The progressive loss of neurons in cerebral cortex and cerebellum, leads to various neurodegenerative diseases. Hence the present study is designed to determine PCBs toxicity toward neuronal cells and whether it could be inhibited by potent antioxidant melatonin. Four groups of adult male Wistar rats were treated for 30 days with corn oil, PCBs, PCBs+Mel and Melatonin, respectively. After treatment period the rats were euthanized and the brain was dissected to isolate cerebral cortex and cerebellum. The neuronal cells alone were then separated from the isolated brain regions, to detect the mRNA levels of apoptotic and neurofilament gene, a neuronal specific marker. Our results suggests that PCBs induces apoptosis in neuronal cells which is subsided by the anti apoptotic effect of melatonin.
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Affiliation(s)
- S Bavithra
- Department of Endocrinology, Dr. ALM Post Graduate Institute of Basic Medical Sciences, University of Madras, Chennai 600 113, India
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Wang YZ, Wang SW, Zhang YC, Sun ZJ. Protective effect of exogenous IGF-I on the intestinalmucosal barrier in rats with severe acute pancreatitis. World J Emerg Med 2012; 3:213-20. [PMID: 25215066 PMCID: PMC4129782 DOI: 10.5847/wjem.j.issn.1920-8642.2012.03.010] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2012] [Accepted: 07/20/2012] [Indexed: 12/18/2022] Open
Abstract
BACKGROUND Severe acute pancreatitis (SAP) can result in intestinal mucosal barrier (IMB) dysfunction. This study was undertaken to demonstrate the effect of IGF-I on the intestinal mucosal barrier in rats with SAP and its possible mechanisms. METHODS Seventy-two male Wistar rats were randomly divided into three groups: a sham operation (SO group, n=24), a SAP group not treated with IGF-I (SAP group, n=24), and a SAP group treated with IGF-I (IGF-I group, n=24). SAP was induced in the rats by injecting 5.0% sodium taurocholate into the biliary-pancreatic duct. The SO rats were given an infusion of normal saline instead. The rats in the IGF-I group underwent the SAP procedure and were given a subcutaneous injection of IGF-I at 30 minutes before the operation and at 3 hours after the operation. Eight rats in each group were sacrificed at 6, 12 and 24 hours after operation. Apoptosis of mucosal cells in the small intestine was determined by TUNEL. The levels of endotoxin and DAO and serum amylase were also measured. Pathologic changes in the small intestine were monitored. Changes of bax and bcl-2 mRNA expression in the small intestine were determined by reverse transcription polymerase chain reaction (RT-PCR). RESULTS The levels of serum amylase were lower in the IGF-I group than in the SAP group at all three time points (P<0.05). The levels of endotoxin in the IGF-I group were higher than those in the SAP group at 6 hours, but lower in the IGF-I group than in the SAP group at 12 and 24 hours (P<0.05). The levels of diamine oxidase were higher in the IGF-I group at 6 hours but lower than those in the SAP group at 12 and 24 hours. The pathological score of the small intestine was lower in the IGF-I group than in the SAP group, and the difference was statistically significant at 12 and 24 hours. The pathologic changes observed under electron microscopy were better in the IGF-I group than those in the SAP group. The apoptosis index of intestinal epithelial cells was significantly decreased in the IGF-I group compared with the SAP group. Compared with the SO group, the mRNA expression levels of bax were increased at each time point in the SAP group, and were significantly decreased in the IGF-I group as compared with the SAP group at each time point (P<0.05). The expression levels of bcl-2 were weak and not different between the SO group and the SAP group (P>0.05). They were significantly increased in the IGF-I group versus the SO and SAP groups (P<0.05). The ratio of bax and bcl-2 mRNA expression levels at each time point in the SAP group were significantly higher than those in the SO group, but they were obviously decreased in the IGF-I group. CONCLUSIONS Exogenous IGF-I seems to protect mucosal cells in the small intestine against SAP-induced apoptosis and could alleviate SAP-induced injury of the intestinal mucosa. The underlying mechanisms include enhanced mRNA expression of bcl-2 and inhibition of bax mRNA expression.
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Affiliation(s)
- Ying-zhen Wang
- Intensive Care Unit, Second Hospital of Gansu Province, Lanzhou 730000, China
- Department of General Surgery, Lanzhou University Second Hospital, Lanzhou 730030, China
| | - Shi-wen Wang
- Intensive Care Unit, Second Hospital of Gansu Province, Lanzhou 730000, China
| | - You-cheng Zhang
- Department of General Surgery, Lanzhou University Second Hospital, Lanzhou 730030, China
| | - Zhi-jiang Sun
- Intensive Care Unit, General Hospital of Lanzhou Petrochemical, Lanzhou 730060, China
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Dong L, Fan Y, Shao X, Chen Z. Vitexin protects against myocardial ischemia/reperfusion injury in Langendorff-perfused rat hearts by attenuating inflammatory response and apoptosis. Food Chem Toxicol 2011; 49:3211-6. [PMID: 22001368 DOI: 10.1016/j.fct.2011.09.040] [Citation(s) in RCA: 58] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2011] [Revised: 09/05/2011] [Accepted: 09/26/2011] [Indexed: 01/04/2023]
Abstract
The aim of the present study is to investigate the effects and its possible underlying mechanisms of vitexin on myocardial ischemia/reperfusion (I/R) injury in isolated rat hearts. Isolated rat hearts were perfused with Langendorff apparatus, which subjected to 30 min ischemia and then followed by 60 min reperfusion. In the isolated rat heart subjected to I/R injury, treatment of vitexin (50, 100, 200 μmol/L) significantly enhanced coronary flow, and decreased the pathological scores of myocardium. 50, 100, 200 μmol/L vitexin significantly attenuated I/R-induced increases of myocardial TNF-α and IL-1β, and 25, 50, 100, 200 μmol/L vitexin significantly reduced apoptosis index of cardiac muscle cell of rat isolated heart subjected to I/R injury. Vitexin significantly inhibited I/R-induced increase of myocardial Bax protein expression; however, 100, 200 μmol/L vitexin markedly increased myocardial Bcl-2 protein expression. Furthermore, vitexin at concentrations of 50, 100, 200 μmol/L significantly reduced expression of myocardial NF-κBp65 protein. Therefore, these results demonstrate that vitexin exhibits significant protective effect against myocardial I/R injury in isolated rat heart, which is related to inhibition of the release of inflammatory cytokines and the apoptosis of cardiac muscle cell via up-regulating protein expression of Bcl-2 as well as down-regulating Bax and NF-κBp65.
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Affiliation(s)
- Liuyi Dong
- Department of Pharmacology, Anhui Medical University, Hefei, Anhui 230032, PR China
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Atapattu DN, Czuprynski CJ. Mannheimia haemolytica leukotoxin induces apoptosis of bovine lymphoblastoid cells (BL-3) via a caspase-9-dependent mitochondrial pathway. Infect Immun 2005; 73:5504-13. [PMID: 16113266 PMCID: PMC1231077 DOI: 10.1128/iai.73.9.5504-5513.2005] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2005] [Revised: 04/08/2005] [Accepted: 05/03/2005] [Indexed: 12/21/2022] Open
Abstract
Mannheimia haemolytica is a key pathogen in the bovine respiratory disease complex. It produces a leukotoxin (LKT) that is an important virulence factor, causing cell death in bovine leukocytes. The LKT binds to the beta(2) integrin CD11a/CD18, which usually activates signaling pathways that facilitate cell survival. In this study, we investigated mechanisms by which LKT induces death in bovine lymphoblastoid cells (BL-3). Incubation of BL-3 cells with a low concentration of LKT results in the activation of caspase-3 and caspase-9 but not caspase-8. Similarly, the proapoptotic proteins Bax and BAD were significantly elevated, while the antiapoptotic proteins Bcl-2, Bcl(XL) and Akt-1 were downregulated. Following exposure to LKT, we also observed a reduction in mitochondrial cytochrome c and corresponding elevation of cytosolic cytochrome c, suggesting translocation from the mitochondrial compartment to the cytosol. Consistent with this observation, tetramethylrhodamine ethyl ester perchlorate staining revealed that mitochondrial membrane potential was significantly reduced. These data suggest that LKT induces apoptosis of BL-3 cells via a caspase-9-dependent mitochondrial pathway. Furthermore, scanning electron micrographs of mitochondria from LKT-treated BL-3 cells revealed lesions in the outer mitochondrial membrane, which are larger than previous reports of the permeability transition pore through which cytochrome c is usually released.
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Affiliation(s)
- Dhammika N Atapattu
- Department of Pathobiological Sciences, University of Wisconsin, School of Veterinary Medicine, 2015 Linden Dr. West, Madison, WI 53706, USA
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Chen W, Fu XB, Ge SL, Sun TZ, Zhou G, Han B, Du YR, Li HH, Sheng ZY. Intravenous acid fibroblast growth factor protects intestinal mucosal cells against ischemia-reperfusion injury via regulating Bcl-2/Bax expression. World J Gastroenterol 2005; 11:3419-25. [PMID: 15948248 PMCID: PMC4315997 DOI: 10.3748/wjg.v11.i22.3419] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To detect the effect of acid fibroblast growth factor (aFGF) on apoptosis and gene expression of bax and bcl-2 gene in rat intestine after ischemia/reperfusion (I/R) injury, and to explore the protective mechanisms of aFGF.
METHODS: One hundred and eight Wistar rats were randomly divided into sham-operated control group (C) (n = 6), intestinal ischemia group (I) (n = 6), aFGF treatment group (A) (n = 48) and intestinal ischemia-reperfusion group (R) (n = 48). In group I, the animals were killed after 45 min of superior mesenteric artery (SMA) occlusion, while in groups R and A, the rats sustained 45 min of SMA occlusion and were then treated with normal saline and aFGF, respectively, sustained 15 min, 30 min, 1, 2, 6, 12, 24, or 48 h of reperfusion, respectively. In group C, SMA was separated, but without occlusion. Apoptosis in intestinal villus was determined with terminal deoxynucleotidyl transferase mediated dUTP-biotin nick-end labeling technique (TUNEL). Intestinal tissue samples were taken not only for detection of bax and bcl-2 gene expression by RT-PCR, but also for detection of bax and bcl-2 protein expression and distribution by immunohistochemical analysis.
RESULTS: The rat survival rates in aFGF treated group were higher than group R (P<0.05) and the improvement of intestinal histological structures was observed at 2, 6, and 12 h after the reperfusion in group A compared with group R. The apoptotic rates were (41.17±3.49)%, (42.83±5.23)% and (53.33±6.92)% at 2, 6 and 12 h after reperfusion, respectively in group A, apparently less than those of group R at matched time points (50.67±6.95, 54.17±7.86, 64.33±6.47, respectively) (P<0.05). The bax gene transcription and translation were significantly decreased in group A vs group R, while mRNA and protein contents of Bcl-2 in group A were obviously higher than those in group R during 2-12 h period after reperfusion.
CONCLUSION: The changes in histological structure and the increment of apoptotic rate indicated that the intestinal barrier was damaged after intestinal I/R injury, whilst intravenous aFGF could alleviate apoptosis induced by ischemia and reperfusion in rat intestinal tissues, in which genes of bax and bcl-2 might play important roles.
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Affiliation(s)
- Wei Chen
- Wound Healing and Cell Biology Laboratory, 304th Hospital, Burns Institute, Trauma Center of Postgraduate Medical College, 51 Fu Cheng Road, Beijing 100037, China
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Chen W, Fu XB, Ge SL, Sun TZ, Zhao JY, Du YR, Sheng ZY. Effects of extrogenous aFGF on bax and bcl-2 expression in intestinal cells after ischemia/reperfusion. Shijie Huaren Xiaohua Zazhi 2004; 12:2599-2604. [DOI: 10.11569/wcjd.v12.i11.2599] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To detect the effects of acid fibroblast growth factor (aFGF) on apoptosis and Bax and bcl-2 expression in rat intestine after I/R injury, and to explore the protective mechanisms of aFGF on intestinal villus.
METHODS: One hundred and eight Wistar rats were randomly divided into 4 groups, namely intestinal ischemia/reperfusion group (R, n = 48), intestinal ischemia group (I, n = 6), aFGF treatment group (A, n = 48) and sham-operated group (C, n = 6). The rats sustained 45 min of arteria mesenterica (SMA) occlusion to establish the ischemia model. At the beginning of reperfusion, rats in group R and A were treated with normal saline (0.15 mL) and aFGF (20 μg/kg, 0.15 mL) respectively. Then each six rats as a sub-group were reperfused for a duration of 0.25, 0.5, 1, 2, 6, 12, 24, 48 h respectively. Cell apoptotic rates in intestinal villus were determined with terminal deoxynucl-eotidy transferase mediated dUTP-biotin nick-end-labeling technique (TUNEL). RT-PCR was used to detect the expressions of bax and bcl-2 gene in intestinal villus. Immunohistochemical methods were adopted to detect bax and bcl-2 protein expressions and distributions.
RESULTS: The improvement of intestinal histological structures was observed at 2 h, 6 h and 12 h after the reperfusion in group A, compared with group R. The apoptotic rates were (41.17±3.49 %), (42.83±5.23 %) and (53.33±6.92 %) at 2, 6, 12 h after reperfusion respectively in group A, and these rates were significantly lower than those in group R (P < 0.05). The expressions of bax gene and bax protein in intestinal villus were gradually increased after ischemia/reperfusion, while the transcription of bcl-2 gene and expression of bcl-2 protein were decreased. During the 2-12 h of reperfusion, the transcription of bcl-2 gene and expression of bcl-2 protein were significantly increased in group A compared with those in group R (P < 0.05). However, the expressions of bax gene and bax protein were significantly higher than those in group R (P < 0.05).
CONCLUSION: Intravenous aFGF could alleviate I/R-induced injury, in which its effects on the facilitation of bcl-2 transcription and inhibition of bax expression may play an important role.
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Affiliation(s)
- Wei Chen
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
| | - Xiao-Bing Fu
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
| | - Shi-Li Ge
- Institute of Radiation Medicine, Academy of Military Medicine Sciences, Beijing 100850, China
| | - Tong-Zhu Sun
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
| | - Jing-Yu Zhao
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
| | - Yi-Ri Du
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
| | - Zhi-Yong Sheng
- Key Research Laboratory of Wound Repair, 304th Hospital of PLA, Beijing 100037, China
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Dong JW, Zhu HF, Zhu WZ, Ding HL, Ma TM, Zhou ZN. Intermittent hypoxia attenuates ischemia/reperfusion induced apoptosis in cardiac myocytes via regulating Bcl-2/Bax expression. Cell Res 2004; 13:385-91. [PMID: 14672562 DOI: 10.1038/sj.cr.7290184] [Citation(s) in RCA: 104] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
Intermittent hypoxia has been shown to provide myocardial protection against ishemia/reperfusion-induced injury. Cardiac myocyte loss through apoptosis has been reported in ischemia/reperfusion injury. Our aim was to investigate whether intermittent hypoxia could attenuate ischemia/reperfusion-induced apoptosis in cardiac myocytes and its potential mechanisms. Adult male Sprague-Dawley rats were exposed to hypoxia simulated 5000 m in a hypobaric chamber for 6 h/day, lasting 42 days. Normoxia group rats were kept under normoxic conditions. Isolated perfused hearts from both groups were subjected to 30 min of global ischemia followed by 60 min reperfusion. Incidence of apoptosis in cardiac myocytes was determined by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) and DNA agarose gel electrophoresis. Expressions of apoptosis related proteins, Bax and Bcl-2, in cytosolic and membrane fraction were detected by Western Blotting. After ischemia/reperfusion, enhanced recovery of cardiac function was observed in intermittent hypoxia hearts compared with normoxia group. Ischemia/reperfusion-induced apoptosis, as evidenced by TUNEL-positive nuclei and DNA fragmentation, was significantly reduced in intermittent hypoxia group compared with normoxia group. After ischemia/reperfusion, expression of Bax in both cytosolic and membrane fractions was decreased in intermittent hypoxia hearts compared with normoxia group. Although ischemia/reperfusion did not induce changes in the level of Bcl-2 expression in cytosolic fraction between intermittent hypoxia and normoxia groups, the expression of Bcl-2 in membrane fraction was upregulated in intermittent hypoxia group compared with normoxia group. These results indicated that the cardioprotection of intermittent hypoxia against ischemia/reperfusion injury appears to be in part due to reduce myocardial apoptosis. Intermittent hypoxia attenuated ischemia/reperfusion-induced apoptosis via increasing the ratio of Bcl-2/Bax, especially in membrane fraction.
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Affiliation(s)
- Jian Wen Dong
- Laboratory of Hypoxic Cardiovascular Physiology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China
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Tang J, Xue H, Pan C, Chen J, Gu L, Zhao H. A homoharringtonine-based regimen for childhood acute myelogenous leukemia. MEDICAL AND PEDIATRIC ONCOLOGY 2003; 41:70-72. [PMID: 12764750 DOI: 10.1002/mpo.10264] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Affiliation(s)
- JingYan Tang
- Department of Hematology/Oncology, Xin Hua Hospital/Shanghai Children's Medical Center, Shanghai Second Medical University.
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Yuan JP, Zhao W, Wang HT, Wu KY, Li T, Guo XK, Tong SQ. Coxsackievirus B3-induced apoptosis and caspase-3. Cell Res 2003; 13:203-9. [PMID: 12862321 DOI: 10.1038/sj.cr.7290165] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023] Open
Abstract
Cell death can be classified into two categories: apoptosis and necrosis. Apoptotic pathway can be either caspase-dependent or caspase-independent. Caspase-independent cytopathic effect (CPE) has been described. In order to evaluate the pattern of HeLa cell death induced by Coxsackievirus B3 (CVB3) and whether apoptosis involves caspase activation, we co-cultivated HeLa cells with CVB3 and detected the cytopathic changes, the alteration of mRNA and protein expression of caspase-3 gene plus caspase-3 activity, as well as analyzing DNA fragmentation before and after caspase-3 activity inhibition. According to the results, we propose that CVB3 may induce apoptosis and necrosis in HeLa cells, the latter appearing much earlier. Caspase-3 is activated at the levels of both transcription and translation, and procaspase-3 is proteolytically cleaved, thus leading to the continuous increasing of both caspase-3 precursor protein and its subunit. However, besides CPE, apoptosis induced by CVB3 is not a direct consequence of the activation of caspase-3, or caspase-3 is not the only effector molecule in apoptotic cell death, for caspase-3 inhibitor can not decrease DNA fragmentation. Some other biochemical mechanisms may participate in the process, whose role weakens the effect of inhibiting caspase-3 activity.
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Affiliation(s)
- Jian Ping Yuan
- Department of Microbiology and Parasitology, Shanghai Second Medical University, Shanghai 200025, China
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Lai D, Fu L, Weng S, Qi L, Yu C, Yu T, Wang H, Chen W. Blocking caspase-3 activity with a U6 SnRNA promoter-driven ribozyme enhances survivability of CHO Cells Cultured in Low Serum Medium and Production of Interferon-? Biotechnol Bioeng 2003; 85:20-8. [PMID: 14705008 DOI: 10.1002/bit.10769] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
Apoptosis responding to various insults in bioreactors was observed to reduce cell viability and prevent cells from growing to high density. Inhibition of apoptosis in different ways has proved to be effective in keeping cells viable in high density and result in higher recombinant protein production. In apoptosis, death signals activate a family of proteinases, namely caspases, in a cascade and ultimately activate the final effector proteinase, caspase-3, which cleaves various substrates and drives the cells irreversibly to death. Caspase-3 is the executioner of an apoptotic cell and thus plays a central role in apoptosis. Therefore inhibition of caspase-3 may provide an effective way for apoptosis prevention. In this study, we designed a ribozyme targeted at the 451 nt of hamster caspase-3's open reading frame (ORF) and the ribozyme was proved to be effective in cleaving caspase-3 mRNA in vitro. Then, the ribozyme was cloned into a vector under the control of U6 snRNA promoter, an RNA polymerase III promoter, for high rate of transcription in vivo. The vector was transfected into an interferon-beta producing recombinant CHO cell line, and some clones were screened out that exhibited low caspase-3 production by Western blot analysis. One such clone was then further analyzed and it showed good anti-apoptosis property with respect to cell viability, cell density, and interferon-beta production.
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Affiliation(s)
- Dazhi Lai
- Department of Applied Molecular Biology, Beijing Institute of Microbiology and Epidemiology, 20 Dongdajie, Fengtai, Beijing 100071, China
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Li HL, Chen DD, Li XH, Zhang HW, Lu YQ, Ye CL, Ren XD. Changes of NF-κB, p53, Bcl-2 and caspase in apoptosis induced by JTE-522 in human gastric adenocarcinoma cell line AGS cells: role of reactive oxygen species. World J Gastroenterol 2002; 8:431-5. [PMID: 12046064 PMCID: PMC4656415 DOI: 10.3748/wjg.v8.i3.431] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To identify whether JTE-522 can induce apoptosis in AGS cells and ROS also involved in the process, and to investigate the changes in NF-κB, p53, bcl-2 and caspase in the apoptosis process.
METHODS: Cell culture, MTT, Electromicroscopy, agarose gel electrophoresis, lucigenin, Western blot and electrophoretic mobility shift assay (EMSA) analysis were employed to investigate the effect of JTE-522 on cell proliferation and apoptosis in AGS cells and related molecular mechanisms.
RESULTS: JTE-522 inhibited the growth of AGS cells and induced the apoptosis. Lucigenin assay showed the generation of ROS in cells under incubation with JTE-522. The increased ROS generation might contribute to the induction of AGS cells to apoptosis. EMSA and Western blot revealed that NF-κB activity was almost completely inhibited by preventing the degradation of IkBα. Additionally, by using Western blot we confirmed that the level of bcl-2 was decreased, whereas p53 showed a great increase following JTE-522 treatment. Their changes were in a dose-dependent manner.
CONCLUSION: These findings suggest that reactive oxygen species, NF-κB, p53, bcl-2 and caspase-3 may play an important role in the induction of apoptosis in AGS cells after treatment with JTE-522.
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Affiliation(s)
- Hong-Liang Li
- Department of Pharmacology, Jinan University Pharmacy College, Guangzhou 510632, Guangdong, China
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