| Loss of sphingosine kinase-1 in carcinoma cells increases formation of reactive oxygen species and sensitivity to doxorubicin-induced DNA damage. | |
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MedLine Citation:
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PMID: 20883472 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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BACKGROUND AND PURPOSE: Sphingosine kinases (SK) catalyse the formation of sphingosine 1-phosphate, which is a key lipid mediator regulating cell responses such as proliferation, survival and migration. Here we have investigated the effect of targeted inhibition of SK-1 on cell damage and elucidated the mechanisms involved. EXPERIMENTAL APPROACH: Three human carcinoma cell lines (colon HCT-116, breast MDA-MB-231, lung NCI-H358) were used, which were either transduced with shRNA constructs to deplete SK-1, or treated with a SK-1 inhibitor. Cell growth and viability were assayed by [(3) H]thymidine incorporation and colony formation. Reactive oxygen species (ROS) were measured by fluorescence and apoptosis by annexin V with flow cytometry. Proteins were analysed by Western blotting. DNA damage was induced by doxorubicin. KEY RESULTS: Knock-down of SK-1 by shRNA strongly inhibited DNA synthesis and colony formation of carcinoma cells. SK-1 knock-down (SK-1kd) cells revealed dysfunctional extracellular signal-regulated protein kinase and PKB/Akt cascades, and contained increased levels of ROS. After SK-1kd, treatment with doxorubicin increased DNA damage, measured by histone-2AX phosphorylation. Similar effects were found in cells with a SK-1 inhibitor and doxorubicin. The increased damage response in SK-1kd cells was accompanied by greater reduction of DNA synthesis and colony formation, and by more pronounced apoptosis. Addition of a NADPH oxidase inhibitor reduced the increased apoptosis in doxorubicin-treated SK-1kd cells. CONCLUSIONS AND IMPLICATIONS: SK-1kd in carcinoma cells triggered oxidative stress by increasing intracellular Ros production. Targeted inhibition of SK-1 represents a promising approach to sensitize cells to DNA damage and facilitate apoptosis upon doxorubicin treatment. |
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Authors:
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Andrea Huwiler; Nataliya Kotelevets; Cuiyan Xin; Oleksandr Pastukhov; Josef Pfeilschifter; Uwe Zangemeister-Wittke |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: British journal of pharmacology Volume: 162 ISSN: 1476-5381 ISO Abbreviation: Br. J. Pharmacol. Publication Date: 2011 Jan |
Date Detail:
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Created Date: 2010-12-15 Completed Date: 2011-06-29 Revised Date: 2012-01-02 |
Medline Journal Info:
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Nlm Unique ID: 7502536 Medline TA: Br J Pharmacol Country: England |
Other Details:
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Languages: eng Pagination: 532-43 Citation Subset: IM |
Copyright Information:
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© 2010 The Authors. British Journal of Pharmacology © 2010 The British Pharmacological Society. |
Affiliation:
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Institute of Pharmacology, University of Bern, Switzerland. huwiler@pki.unibe.ch |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Antibiotics, Antineoplastic
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pharmacology* Apoptosis / drug effects Cell Line, Tumor Cell Movement / drug effects Cell Proliferation / drug effects DNA Damage / drug effects* Down-Regulation Doxorubicin / pharmacology* Female HCT116 Cells Humans Molecular Targeted Therapy Phosphotransferases (Alcohol Group Acceptor) / antagonists & inhibitors*, genetics, metabolism RNA, Small Interfering / genetics Reactive Oxygen Species / metabolism* |
| Chemical | |
Reg. No./Substance:
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0/Antibiotics, Antineoplastic; 0/RNA, Small Interfering; 0/Reactive Oxygen Species; 23214-92-8/Doxorubicin; EC 2.7.1.-/Phosphotransferases (Alcohol Group Acceptor); EC 2.7.1.-/sphingosine kinase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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