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San Sebastián de los Reyes, Spain

Vazquez-Martin A.,Catalan Institute of Oncology ICO | Vazquez-Martin A.,Girona Biomedical Research Institute IDIBGI | Vellon L.,Cell Reprogramming Unit | Quiros P.M.,University of Oviedo | And 10 more authors.
Cell Cycle | Year: 2012

The ability of somatic cells to reprogram their ATP-generating machinery into a Warburg-like glycolytic metabotype while overexpressing stemness genes facilitates their conversion into either induced pluripotent stem cells (iPSCs) or tumor-propagating cells. AMP-activated protein kinase (AMPK) is a metabolic master switch that senses and decodes intracellular changes in energy status; thus, we have evaluated the impact of AMPK activation in regulating the generation of iPSCs from non-stem cells of somatic origin. The indirect and direct activation of AMPK with the antidiabetic biguanide metformin and the thienopyridone A-769662, respectively, impeded the reprogramming of mouse embryonic and human diploid fibroblasts into iPSCs. The AMPK activators established a metabolic barrier to reprogramming that could not be bypassed, even through p53 deficiency, a fundamental mechanism to greatly improve the efficiency of stem-cell production. Treatment with metformin or A-769662 before the generation of iPSC colonies was sufficient to drastically decrease iPSC generation, suggesting that AMPK activation impedes early stem cell genetic reprogramming. Monitoring the transcriptional activation status of each individual reprogramming factor (i.e., Oct4, Sox2, Klf4 and c-Myc) revealed that AMPK activation notably prevented the transcriptional activation of Oct4, the master regulator of the pluripotent state. AMPK activation appears to impose a normalized metabolic flow away from the required pro-immortalizing glycolysis that fuels the induction of stemness and pluripotency, endowing somatic cells with an energetic infrastructure that is protected against reprogramming. AMPK-activating anti-reprogramming strategies may provide a roadmap for the generation of novel cancer therapies that metabolically target tumor-propagating cells. © 2012 Landes Bioscience.

de Eguino G.R.,Hospital Universitario Cruces | Infante A.,Hospital Universitario Cruces | Schlangen K.,CIC Biomagune | Aransay A.M.,CIC Biomagune | And 5 more authors.
Stem Cells Translational Medicine | Year: 2012

Lamin A (LMNA)-linked lipodystrophies may be either genetic (associated with LMNA mutations) or acquired (associated with the use of human immunodeficiency virus protease inhibitors [PIs]), and in both cases they share clinical features such as anomalous distribution of body fat or generalized loss of adipose tissue, metabolic alterations, and early cardiovascular complications. Both LMNA-linked lipodystrophies are characterized by the accumulation of the lamin A precursor prelamin A. The pathological mechanism by which prelamin A accumulation induces the lipodystrophy associated phenotypes remains unclear. Since the affected tissues in these disorders are of mesenchymal origin, we have generated an LMNA-linked experimental model using human mesenchymal stem cells treated with a PI, which recapitulates the phenotypes observed in patient biopsies. This model has been demonstrated to be a useful tool to unravel the pathological mechanism of the LMNA-linked lipodystrophies, providing an ideal system to identify potential targets to generate new therapies for drug discovery screening. We report for the first time that impaired adipogenesis is a consequence of the interaction between accumulated prelamin A and Sp1 transcription factor, sequestration of which results in altered extracellular matrix gene expression. In fact, our study shows a novel, essential, and finely tuned role for Sp1 in adipose lineage differentiation in human mesenchymal stem cells. These findings define a new physiological experimental model to elucidate the pathological mechanisms LMNA-linked lipodystrophies, creating new opportunities for research and treatment not only of LMNA-linked lipodystrophies but also of other adipogenesis-associated metabolic diseases. © AlphaMed Press.

Nam-Cha S.H.,University General Hospital | Serrano-Vargas R.,University General Hospital | Escario E.,University General Hospital | Azana J.M.,University General Hospital | And 3 more authors.
BioMed Research International | Year: 2013

Background. Expression of human CD133 (human prominin-1) in cancer cells has been postulated to be a marker of stemness and is considered as a putative marker of cancer stem cells (CSCs). We designed a study to describe the expression pattern of CD133 in normal skin and in epithelial cutaneous neoplasms. Methods. The CD133 immunohistochemical expression of forty-three eccrine and apocrine tumors was compared to that observed in other epithelial tumors of the skin. In addition, flow cytometry was used to detect the CD133 expression of four epithelial skin neoplasms, including one porocarcinoma. Results. CD133 immunoreactivity at the apical or at the apicolateral surface of cells forming glandular structures was observed. Cells from solid areas of benign or malignant tumors were not stained. The porocarcinoma derived culture cells showed a 22% of CD133 positive cells using flow cytometry, while squamous cell carcinoma cultures contained less than 0.1%. Conclusions. These observations indicate that CD133 is a specific marker of glandular differentiation that could be included in the diagnostic panel of cutaneous tumors with possible eccrine or apocrine differentiation. However, the use of CD133 expression as a marker of CSCs should be interpreted with caution in experiments of skin. © 2013 S. H. Nam-Cha et al.

Iglesias J.M.,Regulation of Cell Growth Laboratory | Iglesias J.M.,Synpromics Ltd | Leis O.,Regulation of Cell Growth Laboratory | Ruiz E.P.,Regulation of Cell Growth Laboratory | And 14 more authors.
Frontiers in Oncology | Year: 2014

The striking similarity displayed at the mechanistic level between tumorigenesis and the generation of induced pluripotent stem cells and the fact that genes and pathways relevant for embryonic development are reactivated during tumor progression highlights the link between pluripotency and cancer. Based on these observations, we tested whether it is possible to use a pluripotency-associated transcriptional reporter, whose activation is driven by the SRR2 enhancer from the Sox2 gene promoter (named S4+ reporter), to isolate cancer stem cells (CSCs) from breast cancer cell lines. The S4+ pluripotency transcriptional reporter allows the isolation of cells with enhanced tumorigenic potential and its activation was switched on and offin the cell lines studied, reflecting a plastic cellular process. Microarray analysis comparing the populations in which the reporter construct is active versus inactive showed that positive cells expressed higher mRNA levels of cytokines (IL-8, IL-6, TNF) and genes (such as ATF3, SNAI2, and KLF6) previously related with the CSC phenotype in breast cancer. © 2014 Iglesias, Leis, Pérez Ruiz, Gumuzio Barrie, Garcia-Garcia, Aduriz, Beloqui, Hernandez-Garcia, Lopez-Mato, Dopazo, Pandiella, Menendez and Martin.

Manuel Iglesias J.,Regulation of Cell Growth Laboratory | Beloqui I.,Regulation of Cell Growth Laboratory | Garcia-Garcia F.,Computational Genomics Institute | Leis O.,Regulation of Cell Growth Laboratory | And 11 more authors.
PLoS ONE | Year: 2013

Tumors are heterogeneous at the cellular level where the ability to maintain tumor growth resides in discrete cell populations. Floating sphere-forming assays are broadly used to test stem cell activity in tissues, tumors and cell lines. Spheroids are originated from a small population of cells with stem cell features able to grow in suspension culture and behaving as tumorigenic in mice. We tested the ability of eleven common breast cancer cell lines representing the major breast cancer subtypes to grow as mammospheres, measuring the ability to maintain cell viability upon serial non-adherent passage. Only MCF7, T47D, BT474, MDA-MB-436 and JIMT1 were successfully propagated as long-term mammosphere cultures, measured as the increase in the number of viable cells upon serial non-adherent passages. Other cell lines tested (SKBR3, MDA-MB-231, MDA-MB-468 and MDA-MB-435) formed cell clumps that can be disaggregated mechanically, but cell viability drops dramatically on their second passage. HCC1937 and HCC1569 cells formed typical mammospheres, although they could not be propagated as long-term mammosphere cultures. All the sphere forming lines but MDA-MB-436 express E-cadherin on their surface. Knock down of E-cadherin expression in MCF-7 cells abrogated its ability to grow as mammospheres, while re-expression of E-cadherin in SKBR3 cells allow them to form mammospheres. Therefore, the mammosphere assay is suitable to reveal stem like features in breast cancer cell lines that express E-cadherin. © 2013 Iglesias et al.

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