Antioxidant defenses in TNF-treated MCF-7 cells: selective increase in MnSOD

Siemankowski, Linda; Morreale, Jeanne; Briehl, Margaret M.

doi:10.1016/s0891-5849(98)00273-1

Cited by 49 publications

(41 citation statements)

References 22 publications

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“…To the best of our knowledge, our study is the first to show protective effect of NAC against Fas-induced apoptosis in HeLa cells. Our results suggest that the cellular oxidative state plays an important role in Fas-induced apoptosis in HeLa cells either by directly transmitting the signals or by maintaining a permissive environment for signaling as previously reported in other systems (Cross et al, 1999;Siemankowski et al, 2000). Thus, while our data show that Nrf2 inhibits Fas-induced apoptosis, further studies would determine whether this inhibition is mediated by regulation of the sensitivity to oxidative stress or by direct regulation of the components in the Fas signaling pathway.…”

Section: Discussionsupporting

confidence: 80%

“…This result is in agreement with earlier findings concerning the effect of dicumarol on apoptosis: it strongly potentiates TNF-a-induced apoptosis in HeLa cells (Cross et al, 1999); it abrogates the suppression of apoptosis by IGF-I in FL5.12 cells overexpressing wide-type (wt) insulin-like growth factor I receptor (IGF-IR) and it induces apoptosis in MCF-7 cells (Krause et al, 2001). Contradictory are the reports that overexpression of NQOI in MCF-7 sensitizes cells to TNF-a (Siemankowski et al, 2000) and that dicumarol blocks wt p53-mediated apoptosis in girradiated normal thymocytes and in Ml myeloid leukemic cells that overexpress wt p53 (Asher et al, 2001). Thus, the effect of dicumarol on apoptosis appears to be specific to cell type as well as to the apoptotic stimuli.…”

Section: Discussionsupporting

confidence: 61%

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Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

et al. 2003

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Here we describe the Achilles' Heel Method (AHM), a new function-based approach for identification of inhibitors of signaling pathways, optimized for human cells. The principle of AHM is the identification of 'sensitizing' cDNAs based on their decreased abundance following selection. As a proof of principle, we have employed AHM for the identification of Fas/CD95/APO-1 pathway inhibitors. HeLa cells were transfected with an antisense cDNA expression library in an episomal vector followed by selection with a suboptimal dose of the apoptotic inducer. Antisense inactivation of Fas inhibitors rendered the cells more sensitive to apoptosis resulting in their preferential death and consequent loss of their sensitizing episomes that were identified by subtraction. We show that the resulting products were enriched for sensitizing cDNAs as seven out of eight candidates tested were confirmed as inhibitors of Fas-induced killing either by transfection or by pharmacological inhibition. Furthermore, we demonstrate by multiple approaches that one candidate, NF-E2 related factor 2 (Nrf2), is an inhibitor of Fas-induced apoptosis. Inactivation of Nrf2 by antisense or by a membrane permeable dominantnegative polypeptide sensitized cells while overexpression of Nrf2 protected cells from Fas-induced apoptosis. In addition, dicumarol, an inhibitor of the phase II detoxifying enzyme NQO1, a downstream target of Nrf2, sensitized cells. Nrf2 induces the production of Glutathione (GSH) and we demonstrated that N-acetyl L-cysteine (NAC), a precursor to GSH, protected cells from Fas-mediated killing. Taken together, AHM is a powerful approach for the identification of inhibitors of a signaling pathway with a low rate of false positives that opens new avenues for function profiling of human genes and discovery of new drug targets.

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Section: Discussionsupporting

confidence: 80%

Section: Discussionsupporting

confidence: 61%

Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

et al. 2003

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“…70 RNA was attached to the membranes by UV cross-linking (Stratalinker, Stratagene, La Jolla, CA, USA). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) cDNA was obtained as previously described by Siemankowski et al 71 cDNAs for the three human thioredoxin peroxidase isoforms, AOE372, 32 NKEFA and NKEFB, 33 were a kind gift from Margareta Berggren (Dr. Garth Powis' laboratory, University of Arizona, Tucson, AZ, USA). These sequences were obtained by using RT ± PCR to clone them from Jurkat cell mRNA.…”

Section: Northern Blotsmentioning

confidence: 99%

“…Target cDNA sequences were excised, purified and 32 P-labeled probes made as previously described. 71 Blots were probed using standard protocols. 70 Transcript levels were quantitated by exposure of the membrane to a phosphorimager screen (Molecular Dynamics, Sunnyvale, CA, USA).…”

Section: Northern Blotsmentioning

confidence: 99%

Thymocytes selected for resistance to hydrogen peroxide show altered antioxidant enzyme profiles and resistance to dexamethasone-induced apoptosis

Tome

Briehl

2001

Cell Death Differ

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Treatment of WEHI7.2 cells, a mouse thymoma-derived cell line, with dexamethasone, a synthetic glucocorticoid, causes the cells to undergo apoptosis. Previous work has shown that treatment of WEHI7.2 cells with dexamethasone results in a downregulation of antioxidant defense enzymes, suggesting that increased oxidative stress may play a role in glucocorticoid-induced apoptosis. To test whether resistance to oxidative stress causes resistance to dexamethasoneinduced apoptosis, WEHI7.2 cell variants selected for resistance to 50, 100 and 200 mM H 2 O 2 were developed. Resistance to H 2 O 2 is accompanied by increased antioxidant enzyme activity, resistance to other oxidants and a delayed loss of viable cells after dexamethasone treatment. In the 200 mM H 2 O 2 -resistant cell variant the delay in cell loss is correlated with delayed release of cytochrome c from the mitochondria into the cytosol. This suggests that reactive oxygen species play a role in a signaling event during steroidmediated apoptosis in lymphocytes. Cell Death and Differentiation (2001) 8, 953 ± 961.

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“…Ionizing radiation, anthracyclines, and cytokines such as TNF-a, produce free radicáis that aid in their anticancer activity (Das, 2002). TNF-a induces apoptosis in the MCF-7 human breast adenocarcinoma-derived cell line but increases MnSOD expression by northem blot hybridization analyses (Siemankowski et al, 1999). A correlation was found between elevated serum levels of MnSOD and recurrence of disease in human epithelial ovarían carcinoma patients and decreased levels after chemotherapy with combinations of cyclophosphamide, doxorubicin and cisplatin (Ishikawa et al, 1990).…”

Section: Loss O F Membrane Integrity or Percentage O F Necrotic Cellsmentioning

confidence: 98%

Cytotoxicity and drug potentiating activity of phenylheptatriyne

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Antioxidant defenses in TNF-treated MCF-7 cells: selective increase in MnSOD

Cited by 49 publications

References 22 publications

Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

Nrf2 is an inhibitor of the Fas pathway as identified by Achilles' Heel Method, a new function-based approach to gene identification in human cells

Thymocytes selected for resistance to hydrogen peroxide show altered antioxidant enzyme profiles and resistance to dexamethasone-induced apoptosis

Cytotoxicity and drug potentiating activity of phenylheptatriyne

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