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Marques-Fernandez F.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | Marques-Fernandez F.,Institute Of Neurociencies | Marques-Fernandez F.,Research Center Biome Dica en Red Sobre Enfermedades Neurodegenerativas | Planells-Ferrer L.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | And 21 more authors.
Cell Death and Disease | Year: 2013

Activation of tumor necrosis factor receptor-1 can trigger survival or apoptosis pathways. In many cellular models, including the neuronal cell model PC12, it has been demonstrated that inhibition of protein synthesis is sufficient to render cells sensitive to apoptosis induced by TNFα. The survival effect is linked to the translocation of the transcription factor nuclear factor-kappa B (NF-κB) to the nucleus and activation of survival-related genes such as FLICE-like inhibitory protein long form (FLIP-L) or IAPs. Nonetheless, we previously reported an NF-κB-independent contribution of Bcl-xL to cell survival after TNFα treatment. Here, we demonstrate that NF-κB-induced increase in FLIP-L expression levels is essential for mitogen-activated protein kinases/ extracellular signal-regulated kinases (MAPK/ERK) activation. We demonstrate that FLIP-L behaves as a Raf-1 activator through both proteinprotein interaction and Raf-1 kinase activation, without the requirement of the classical Ras activation. Importantly, prevention of FLIP-L increase by NF-κB inhibition or knockdown of endogenous FLIP-L blocks MAPK/ERK activation after TNFα treatment. From a functional point of view, we show that inhibition of the MAPK/ERK pathway and the NF-κB pathway are equally relevant to render PC12 cells sensitive to cell death induced by TNFα. Apoptosis induced by TNFα under these conditions is dependent on jun nuclear kinase1/2 JNK1/2-dependent Bim upregulation. Therefore, we report a previously undescribed and essential role for MAPK/ERK activation by FLIP-L in the decision between cell survival and apoptosis upon TNFα stimulation. © 2013 Macmillan Publishers Limited All rights reserved.


Galenkamp K.M.O.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | Carriba P.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | Urresti J.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | Planells-Ferrer L.,Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron | And 6 more authors.
Molecular Cancer | Year: 2015

Background: Patients with high-risk neuroblastoma (NBL) tumors have a high mortality rate. Consequently, there is an urgent need for the development of new treatments for this condition. Targeting death receptor signaling has been proposed as an alternative to standard chemo- and radio-therapies in various tumors. In NBL, this therapeutic strategy has been largely disregarded, possibly because ~50-70% of all human NBLs are characterized by caspase-8 silencing. However, the expression of caspase-8 is detected in a significant group of NBL patients, and they could therefore benefit from treatments that induce cell death through death receptor activation. Given that cytokines, such as TNFα, are able to upregulate Fas expression, we sought to address the therapeutic relevance of co-treatment with TNFα and FasL in NBL. Methods: For the purpose of the study we used a set of eight NBL cell lines. Here we explore the cell death induced by TNFα, FasL, cisplatin, and etoposide, or a combination thereof by Hoechst staining and calcein viability assay. Further assessment of the signaling pathways involved was performed by caspase activity assays and Western blot experiments. Characterization of Fas expression levels was achieved by qRT-PCR, cell surface biotinylation assays, and cytometry. Results: We have found that TNFα is able to increase FasL-induced cell death by a mechanism that involves the NF-κB-mediated induction of the Fas receptor. Moreover, TNFα sensitized NBL cells to DNA-damaging agents (i.e. cisplatin and etoposide) that induce the expression of FasL. Priming to FasL-, cisplatin-, and etoposide-induced cell death could only be achieved in NBLs that display TNFα-induced upregulation of Fas. Further analysis denotes that the high degree of heterogeneity between NBLs is also manifested in Fas expression and modulation thereof by TNFα. Conclusions: In summary, our findings reveal that TNFα sensitizes NBL cells to FasL-induced cell death by NF-κB-mediated upregulation of Fas and unveil a new mechanism through which TNFα enhances the efficacy of currently used NBL treatments, cisplatin and etoposide. © Galenkamp et al.;licensee BioMed Central.


PubMed | Fundacio Institute Of Recerca Of Lhospital Universitari Of La Vall Dhebron
Type: | Journal: Molecular cancer | Year: 2015

Patients with high-risk neuroblastoma (NBL) tumors have a high mortality rate. Consequently, there is an urgent need for the development of new treatments for this condition. Targeting death receptor signaling has been proposed as an alternative to standard chemo- and radio-therapies in various tumors. In NBL, this therapeutic strategy has been largely disregarded, possibly because ~50-70% of all human NBLs are characterized by caspase-8 silencing. However, the expression of caspase-8 is detected in a significant group of NBL patients, and they could therefore benefit from treatments that induce cell death through death receptor activation. Given that cytokines, such as TNF, are able to upregulate Fas expression, we sought to address the therapeutic relevance of co-treatment with TNF and FasL in NBL.For the purpose of the study we used a set of eight NBL cell lines. Here we explore the cell death induced by TNF, FasL, cisplatin, and etoposide, or a combination thereof by Hoechst staining and calcein viability assay. Further assessment of the signaling pathways involved was performed by caspase activity assays and Western blot experiments. Characterization of Fas expression levels was achieved by qRT-PCR, cell surface biotinylation assays, and cytometry.We have found that TNF is able to increase FasL-induced cell death by a mechanism that involves the NF-B-mediated induction of the Fas receptor. Moreover, TNF sensitized NBL cells to DNA-damaging agents (i.e. cisplatin and etoposide) that induce the expression of FasL. Priming to FasL-, cisplatin-, and etoposide-induced cell death could only be achieved in NBLs that display TNF-induced upregulation of Fas. Further analysis denotes that the high degree of heterogeneity between NBLs is also manifested in Fas expression and modulation thereof by TNF.In summary, our findings reveal that TNF sensitizes NBL cells to FasL-induced cell death by NF-B-mediated upregulation of Fas and unveil a new mechanism through which TNF enhances the efficacy of currently used NBL treatments, cisplatin and etoposide.

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