Científicos asturianos hallan un nuevo método para detectar el cáncer de páncreas

Artículo periodístico de divulgación científica escrito por Marcos Palicio.Peer Reviewe

DNA methylation patterns in newborns exposed to tobacco in utero

[Background] Maternal smoking during pregnancy is a major risk factor for adverse health outcomes. The main objective of the study was to assess the impact of in utero tobacco exposure on DNA methylation in children born at term with appropriate weight at birth.[Methods] Twenty mother-newborn dyads, after uncomplicated pregnancies, in the absence of perinatal illness were included. All mothers were healthy with no cardiovascular risk factors, except for the associated risks among those mothers who smoked. Umbilical cord blood and maternal peripheral venous blood were collected and an epigenome-wide association study was performed using a 450 K epigenome-wide scan (Illumina Infinium HumanMethylation 450BeadChip) with adjustment to normalize the DNA methylation for data cell variability in whole blood.[Results] The maternal plasmatic cotinine levels ranged from 10.70-115.40 ng/ml in the exposed group to 0-0.59 ng/ml in the non-exposed group. After adjusting for multiple comparisons in 427102 probes, statistically significant differences for 31 CpG sites, associated to 25 genes were observed. There was a greater than expected proportion of statistically-significant loci located in CpG islands (Fisher’s exact test, p = 0.029) and of those CpG islands, 90.3% exhibit higher methylation levels in the exposed group. The most striking and significant CpG site, cg05727225, is located in the chromosome 11p15.4, within the adrenomedullin gene.[Conclusions] In utero tobacco exposure, even in the absence of fetal growth restriction, may alter the epigenome, contributing to global DNA hypomethylation. Therefore, DNA status can be used as a biomarker of prenatal insults. Considering the possibility to reverse epigenetic modifications, a window of opportunity exists to change the programmed chronic disease.The study was partially funded by grant number PI11/00144, Instituto de Salud Carlos III, Spain and CIBER Fisiopatología Obesidad y Nutrición (CB06/03), Instituto de Salud Carlos III, Spain.Peer reviewe

The transcription factor Slug represses E-cadherin expression and induces epithelial to mesenchymal transitions: a comparison with Snail and E47 repressors

13 pages, 7 figures.Transcriptional repression mechanisms have emerged as one of the crucial processes for the downregulation of E-cadherin expression during development and tumour progression. Recently, several E-cadherin transcriptional repressors have been characterized (Snail, E12/E47, ZEB-1 and SIP-1) and shown to act through an interaction with proximal E-boxes of the E-cadherin promoter. We have analyzed the participation of another member of the Snail family, Slug, and observed that it also behaves as a repressor of E-cadherin expression. Stable expression of Slug in MDCK cells leads to the full repression of E-cadherin at transcriptional level and triggers a complete epithelial to mesenchymal transition. Slug-induced repression of E-cadherin is mediated by its binding to proximal E-boxes, particularly to the E-pal element of the mouse promoter. Detailed analysis of the binding affinity of different repressors to the E-pal element indicates that Slug binds with lower affinity than Snail and E47 proteins. These results, together with the known expression patterns of these factors in embryonic development and carcinoma cell lines, support the idea that the in vivo action of the different factors in E-cadherin repression can be modulated by their relative concentrations as well as by specific cellular or tumour contexts.This work was supported by the Spanish Ministry of Science and Technology (SAF2001-2819), Instituto de Salud Carlos III (FIS01/1174) and the Comunidad Autónoma de Madrid (08.1/0055./2000). V.B. has been funded by predoctoral fellowships from the Fundación Científica de la Asociación Española contra el Cáncer (AECC) and Instituto de Salud Carlos III. H.P. is a predoctoral fellow of the Spanish Ministry of Education, Culture and Sports.Peer reviewe

The affinity of different MBD proteins for a specific methylated locus depends on their intrinsic binding properties

The methyl-CpG binding domain (MBD) family of proteins was defined based on sequence similarity in their DNA binding domains. In light of their high degree of conservation, it is of inherent interest to determine the genomic distribution of these proteins, and their associated co-repressor complexes. One potential determinant of specificity resides in differences in the intrinsic DNA binding properties of the various MBD proteins. In this report, we use a capillary electrophoretic mobility shift assay (CEMSA) with laser-induced fluorescence (LIF) and neutral capillaries to calculate MBD-DNA binding affinities. MBD proteins were assayed on pairs of methylated and unmethylated duplex oligos corresponding to the promoter regions of the BRCA1, MLH1, GSTP1 and p16(INK4a) genes, and binding affinities for each case were calculated by Scatchard analyses. With the exception of mammalian MBD3 and Xenopus MBD3 LF, all the MBD proteins showed higher affinity for methylated DNA (in the nanomolar range) than for unmethylated DNA (in the micromolar range). Significant differences between MBD proteins in the affinity for methylated DNA were observed, ranging within two orders of magnitude. By mutational analysis of MBD3 and using CEMSA, we demonstrate the critical role of specific residues within the MBD in conferring selectivity for methylated DNA. Interestingly, the binding affinity of specific MBD proteins for methylated DNA fragments from naturally occurring sequences are affected by local methyl-CpG spacing

Generation of a human iPSC line from a patient with Leigh syndrome

Human iPSC line LND554SV.3 was generated from heteroplasmic fibroblasts of a patient with Leigh syndrome carrying a mutation in the MT-ND5 gene (m.13513G. >. A; p.D393N). Reprogramming factors Oct3/4, Sox2, Klf4, and cMyc were delivered using a non-integrative methodology that involves the use of Sendai virus.This work was supported by grants from the “Centro de Investigación Biomédica en Red en Enfermedades Raras” (CIBERER) (grant 13-717/132.05 to RG), the “Instituto de Salud Carlos III” [Fondo de Investigación Sanitaria and Regional Development Fund (ERDF/FEDER) funds PI10/0703 and PI13/00556 to RG and PI15/00484 to MEG], “Comunidad Autónoma de Madrid” (grant number S2010/BMD-2402 to RG); TG receives grant support from the Universidad Autónoma de Madrid (FPI-UAM) and FZD from the Ministerio de Educación, Cultura y Deporte (FPU13/00544). MEG is a staff scientist at the “Centro de Investigación Biomédica en Red en Enfermedades Raras” (CIBERER) at the “Centro de Investigación Biomédica en Red en Enfermedades Raras” (CIBERER)

Generation of a human control iPSC line with a European mitochondrial haplogroup U background

Human iPSC line N44SV.5 was generated from primary normal human dermal fibroblasts belonging to the European mitochondrial haplogroup U. For this purpose, reprogramming factors Oct3/4, Sox2, Klf4, and cMyc were delivered using a non-integrative methodology that involves the use of Sendai virus.This work was supported by grants from the “Centro de Investigación Biomédica en Red en enfermedades raras” (CIBERER) (grant 13-717/132.05 to RG), the “Instituto de Salud Carlos III” [Fondo de Investigación Sanitaria and Regional Development Fund (ERDF/FEDER) funds PI10/0703 and PI13/00556 to RG and PI15/00484 to MEG], “Comunidad Autónoma de Madrid” (grant number S2010/BMD-2402 to R.G); T.G. receives grant support from the Universidad Autónoma de Madrid, FPI-UAM and F.Z.D. from the Ministerio de Educación, Cultura y Deporte, grant number FPU13/00544. M.E.G. is staff scientist at the “Centro de Investigación Biomédica en Red en Enfermedades Raras” (CIBERER

Stability of Imprinting and Differentiation Capacity in Naïve Human Cells Induced by Chemical Inhibition of CDK8 and CDK19

Pluripotent stem cells can be stabilized in vitro at different developmental states by the use of specific chemicals and soluble factors. The naïve and primed states are the best characterized pluripotency states. Naïve pluripotent stem cells (PSCs) correspond to the early pre-implantation blastocyst and, in mice, constitute the optimal starting state for subsequent developmental applications. However, the stabilization of human naïve PSCs remains challenging because, after short-term culture, most current methods result in karyotypic abnormalities, aberrant DNA methylation patterns, loss of imprinting and severely compromised developmental potency. We have recently developed a novel method to induce and stabilize naïve human PSCs that consists in the simple addition of a chemical inhibitor for the closely related CDK8 and CDK19 kinases (CDK8/19i). Long-term cultured CDK8/19i-naïve human PSCs preserve their normal karyotype and do not show widespread DNA demethylation. Here, we investigate the long-term stability of allele-specific methylation at imprinted loci and the differentiation potency of CDK8/19i-naïve human PSCs. We report that long-term cultured CDK8/19i-naïve human PSCs retain the imprinting profile of their parental primed cells, and imprints are further retained upon differentiation in the context of teratoma formation. We have also tested the capacity of long-term cultured CDK8/19i-naïve human PSCs to differentiate into primordial germ cell (PGC)-like cells (PGCLCs) and trophoblast stem cells (TSCs), two cell types that are accessible from the naïve state. Interestingly, long-term cultured CDK8/19i-naïve human PSCs differentiated into PGCLCs with a similar efficiency to their primed counterparts. Also, long-term cultured CDK8/19i-naïve human PSCs were able to differentiate into TSCs, a transition that was not possible for primed PSCs. We conclude that inhibition of CDK8/19 stabilizes human PSCs in a functional naïve state that preserves imprinting and potency over long-term culture

Generation of a human iPSC line from a patient with an optic atrophy ‘plus’ phenotype due to a mutation in the OPA1 gene

AbstractHuman iPSC line Oex2054SV.4 was generated from fibroblasts of a patient with an optic atrophy ‘plus’ phenotype associated with a heterozygous mutation in the OPA1 gene. Reprogramming factors OCT3/4, SOX2, CMYC and KLF4 were delivered using a non-integrative methodology that involves the use of Sendai virus

Generation of a human iPSC line from a patient with a defect of intergenomic communication

Human iPSC line PG64SV.2 was generated from fibroblasts of a patient with a defect of intergenomic communication. This patient harbored a homozygous mutation (c.2243G>C; p.Trp748Ser) in the gene encoding the catalytic subunit of the mitochondrial DNA polymerase gamma gene (POLG). Reprogramming factors Oct3/4, Sox2, Klf4, and cMyc were delivered using a non integrative methodology that involves the use of Sendai virus.This work was supported by grants from the “Centro de Investigación Biomédica en Red en enfermedades raras” (CIBERER) (Grant 13-717/132.05 to RG), the “Instituto de Salud Carlos III” [Fondo de Investigación Sanitaria and Regional development fund (ERDF/FEDER) funds PI10/0703 and PI13/00556 to RG and PI15/00484 to MEG], “Comunidad Autónoma de Madrid” (Grant number S2010/BMD-2402 to RG); TG receives grant support from the Universidad Autónoma de Madrid (FPI-UAM) and FZD from the Ministerio de Educación, Cultura y Deporte (Grant FPU13/00544). MEG is staff scientist at the “Centro de Investigación Biomédica en Red en Enfermedades Raras” (CIBERER