Bases moleculars i estratègies terapèutiques contra les metàstasis del melanoma uveal

[cat] El melanoma uveal (UM) és el tumor intraocular més freqüent en adults. La cirurgia i la braquiteràpia han millorat les taxes de supervivència, però fins al 40% dels pacients desenvolupen metàstasis hepàtiques que són refractàries a les teràpies convencionals i causen la mort dels pacients. Així doncs, combatre el procés de metàstasi del càncer és imprescindible per obtenir un tractament eficaç que augmenti la supervivència global dels pacients. Recentment, s'ha proposat que els tumors contenen una subpoblació de cèl·lules amb un comportament similar a cèl·lules mare i una major resistència als fàrmacs, les quals són responsables de les metàstasis. És per això que per trobar noves dianes i ajustar les dosis terapèutiques recomanades, vam realitzar els nostres estudis farmacològics in vitro utilitzant tant les poblacions resistents a la mort cel·lular per anoikis com les cèl·lules no seleccionades de línies cel·lulars de melanoma uveal humà. Aquestes tenien morfologies diferents, eren derivades tant de metàstasis com del tumor primari, i presentaven mutacions en els oncogens GNAQ o GNA11. En primer lloc, vam veure que les poblacions resistents a l’anoikis eren capaces de créixer en condicions no adherents, com a esferoides i “melanosferes”, i mostraven característiques associades amb cèl·lules pluripotencials. També mostraven una major resistència a la quimioteràpia i expressaven marcadors propis de cèl·lules mare, com ara CD133, CD44, CD15 i els factors de transcripció OCT-4, SOX-2, NANOG i MITF, en comparació amb la població global que creixia en condicions 2D. A més, aquest fenotip característic de les cèl·lules mare tumorals revertia després de 5-6 divisions en condicions adherents de cultiu in vitro. Paral·lelament, vam observar que el tractament in vitro amb TGF-β de les cèl·lules cultivades en 2D desencadenava una regulació ascendent dels gens relacionats amb la pluripotència cel·lular i els factors de transcripció reguladors de la transició epiteli-mesènquima (EMT), SNAI1, SNAI2, TWIST1 i ZEB1, aconseguint nivells d'expressió similars als de les “melanosferes”. Significativament, només aquelles cèl·lules provinents de les “melanosferes” van mostrar la capacitat de migrar cap a un medi condicionat per fibroblasts que contenia IGF-1. Aquesta capacitat de migració, es va veure bloquejada a través de l’anticòs IMC-A12, contra el IGFR-1β. D'acord amb aquests resultats, vam determinar que les cèl·lules de les “melanosferes” sobreexpressaven el receptor IGFR-1β alhora que secretaven MMP-2 al medi extracel·luar. Els efectes antiproliferatius de diversos inhibidors de proteïna quinases sobre aquestes cèl·lules es van analitzar a través de la biblioteca d’inhibidors de quinases Screen-Well® (BML-2832-0100). En aquest sentit, vam seleccionar els inhibidors de MEK i IGFR-1β com a millors candidats per combatre les cèl·lules més agressives del melanoma uveal amb característiques de cèl·lules mare i un fenotip altament migratori. Finalment, resultats preliminars van indicar que la línia cel·lular OMM-2.5, derivada d’una metàstasi de melanoma uveal, produïa el major nombre de vesícules extracel·lulars in vitro. Aquestes vesícules, eren majoritàriament exosomes, transportaven biomarcadors, integrines i ARN exosomals com a càrrega, i podien ser captades pels hepatòcits.[eng] Uveal Melanoma (UM) is the most common intraocular tumour in adults. Surgery and brachytherapy have improved survival rates, but up to 40% of patients develop liver metastases that are refractory to conventional therapies and cause patients’ death. Thus, targeting cancer metastasis is imperative for effective treatment to increase patients’ overall survival. It has been proposed that tumours may contain a subpopulation of cells with stem cell-like behaviour and increased drug resistance, which are the seeds of metastases. To find new targets and refine appropriate therapeutic doses we performed our in vitro drug-screeening studies using both, anoikis-resistant populations and unselected cells from human uveal melanoma cell lines. These had different morphologies, metastatic or primary origin, and GNAQ or GNA11 mutations. First, we found that anoikis-resistant populations were able to grow in non- adherent conditions as multicellular spheroids and melanospheres which displayed traits associated with stemness. They showed increased chemoresistance and expression of stem-cell markers such as CD133, CD44, CD15, and the transcription factors OCT-4, SOX-2, NANOG and MITF, as compared to the global population growing in 2D conditions. Moreover, this stemness-like phenotype could be switched back after 5-6 passages under attached conditions. Furthermore, treatment of 2D cultures with TGF-β triggered an up-regulation of the stemness-related genes and transcription factors SNAI1, SNAI2, TWIST1 and ZEB1 among other EMT-related genes, reaching similar expression levels to those from melanospheres. Remarkably, only cells from melanospheres were able to migrate towards a conditioned-media from fibroblast containing IGF-1. This transwell-migration ability was impaired by IMC-A12 antibody against IGFR- 1β. In accordance with these results, cells from melanospheres overexpressed receptor IGFR-1β. In addition, these cells also produced MMP-2. The antiproliferative effects of protein kinase inhibitors were stated using the Screen-Well® Kinase Inhibitor Library (BML-2832-0100). In this regard, we favor the MEK and IGFR-1β inhibitors to fight aggressive uveal melanoma cells with a stemness and migratory phenotype. Finally, preliminary results indicated that OMM-2.5 cell line, derived from metastasis, produced the largest number of extracellular vesicles. These vesicles were mostly exosomes, were uptaken by hepatocytes and carried exosomal biomarkers, integrins and RNAs as cargo

In utero and childhood exposure to tobacco smoke and multi-layer molecular signatures in children

Background The adverse health effects of early life exposure to tobacco smoking have been widely reported. In spite of this, the underlying molecular mechanisms of in utero and postnatal exposure to tobacco smoke are only partially understood. Here, we aimed to identify multi-layer molecular signatures associated with exposure to tobacco smoke in these two exposure windows. Methods We investigated the associations of maternal smoking during pregnancy and childhood secondhand smoke (SHS) exposure with molecular features measured in 1203 European children (mean age 8.1 years) from the Human Early Life Exposome (HELIX) project. Molecular features, covering 4 layers, included blood DNA methylation and gene and miRNA transcription, plasma proteins, and sera and urinary metabolites. Results Maternal smoking during pregnancy was associated with DNA methylation changes at 18 loci in child blood. DNA methylation at 5 of these loci was related to expression of the nearby genes. However, the expression of these genes themselves was only weakly associated with maternal smoking. Conversely, childhood SHS was not associated with blood DNA methylation or transcription patterns, but with reduced levels of several serum metabolites and with increased plasma PAI1 (plasminogen activator inhibitor-1), a protein that inhibits fibrinolysis. Some of the in utero and childhood smoking-related molecular marks showed dose-response trends, with stronger effects with higher dose or longer duration of the exposure. Conclusion In this first study covering multi-layer molecular features, pregnancy and childhood exposure to tobacco smoke were associated with distinct molecular phenotypes in children. The persistent and dose-dependent changes in the methylome make CpGs good candidates to develop biomarkers of past exposure. Moreover, compared to methylation, the weak association of maternal smoking in pregnancy with gene expression suggests different reversal rates and a methylation-based memory to past exposures. Finally, certain metabolites and protein markers evidenced potential early biological effects of postnatal SHS, such as fibrinolysis

Dissecting the Shared Genetic Architecture of Suicide Attempt, Psychiatric Disorders, and Known Risk Factors

Background Suicide is a leading cause of death worldwide, and nonfatal suicide attempts, which occur far more frequently, are a major source of disability and social and economic burden. Both have substantial genetic etiology, which is partially shared and partially distinct from that of related psychiatric disorders. Methods We conducted a genome-wide association study (GWAS) of 29,782 suicide attempt (SA) cases and 519,961 controls in the International Suicide Genetics Consortium (ISGC). The GWAS of SA was conditioned on psychiatric disorders using GWAS summary statistics via multitrait-based conditional and joint analysis, to remove genetic effects on SA mediated by psychiatric disorders. We investigated the shared and divergent genetic architectures of SA, psychiatric disorders, and other known risk factors. Results Two loci reached genome-wide significance for SA: the major histocompatibility complex and an intergenic locus on chromosome 7, the latter of which remained associated with SA after conditioning on psychiatric disorders and replicated in an independent cohort from the Million Veteran Program. This locus has been implicated in risk-taking behavior, smoking, and insomnia. SA showed strong genetic correlation with psychiatric disorders, particularly major depression, and also with smoking, pain, risk-taking behavior, sleep disturbances, lower educational attainment, reproductive traits, lower socioeconomic status, and poorer general health. After conditioning on psychiatric disorders, the genetic correlations between SA and psychiatric disorders decreased, whereas those with nonpsychiatric traits remained largely unchanged. Conclusions Our results identify a risk locus that contributes more strongly to SA than other phenotypes and suggest a shared underlying biology between SA and known risk factors that is not mediated by psychiatric disorders.Peer reviewe

In-fan-cia : educar de 0 a 6 años : revista de la Associació de Mestres Rosa Sensat

Analiza el fenómeno fisiológico del sueño. Desde la fase de sueño hasta el despertar. Esta ultima etapa de desarrolla con profundidad, analiza qué tipos existen, el despertar espontáneo y el provocado; cuáles son las formas de despertarse y los estados anímicos después de despertarse y los trastornos al despertar, como despertares confusionarios, sonambulismo y terrores nocturnos.CataluñaES

Causas de insônia nos primeiros anos de vida e repercussão nas mães: atualização Causes of insomnia in the first years of life and maternal consequences: an update

OBJETIVO: Revisar a literatura sobre insônia de crianças pequenas e seus aspectos relacionados às repercussões familiares, à abordagem pediátrica e ao seu tratamento. FONTES DE DADOS: Levantamento de publicações indexadas no Medline e Lilacs entre os anos de 1998 e 2008, rastreadas com a combinação dos descritores: "sleep", "insomnia", "child", "depression", "mother-child relationship", além de teses e capítulos de livros pertinentes ao assunto. SÍNTESE DOS DADOS: A insônia da criança pequena, definida como dificuldade repetida em iniciar e/ou manter o sono, é queixa frequente na clínica pediátrica e usualmente traz repercussões nos pais relativas à privação de sono. Aos dois a três meses de idade, há condições biológicas para consolidação das horas de sono à noite; estando a conduta parental associada às dificuldades neste processo. Estudos demonstram que há associação entre insônia do bebê e depressão nas mães, e a insônia pode ser manifestação de dificuldades no desenvolvimento psíquico da criança pequena, que acontece na relação com sua mãe. CONCLUSÕES: O ritmo de sono nos bebês pode e deve estabelecer-se precocemente; os hábitos para o sono devem basear-se nas medidas de higiene do sono. A depressão nas mães, que pode contribuir para a insônia do bebê, idealmente poderia ser detectada pelo pediatra, prevenindo esse comprometimento das mães e de seus bebês.<br>OBJECTIVE: To review the literature on insomnia in small children, its aspects related to effects on parents, pediatric approach and treatment. DATA SOURCES: A survey of publications indexed in Medline and Lilacs between the years 1998 and 2008, searched through the combination of the descriptors "sleep", "insomnia", "child", "depression", "mother-child relationship", besides thesis and chapters of books concerning the subject. DATA SYNTHESIS: The small child insomnia, defined as repeated difficulty in initiating and/or maintaining sleep, is a common complaint in the pediatric clinic, and usually entails repercussions on parents related to sleep deprivation. At about two to three months of age, there are already biological conditions for consolidating sleep hours during the night. Parental behavior is associated to difficulties to establish this process. Studies point out that baby's insomnia and mother's depression are closely related. Probably, the insomnia is a sign of the difficulties in the small child's psychic development in the context of mother-child relationship. CONCLUSIONS: Rhythmic sleep in babies can and should be established early in life; sleep habits should be built on rules of sleep hygiene. Mother's depression, which may contribute to baby's insomnia, should ideally be detected by pediatricians, thus preventing such suffering for mothers and babies