Evaluation of the expression of the microphthalmia-associated transcription factor (MITF) in peripheral blood of individuals with and without malignant melanoma and in cell lines

Introducción:La incidencia de melanoma maligno se ha incrementado más rápido que cualquier otro tipo de cáncer,intensificando así la búsqueda de herramientas que faciliten la identificación temprana del melanoma. El factor de transcripciónasociado con microftalmia (MITF) es conocido como el regulador maestro de melanocitos. En el presente estudio se analizala expresión del gen MITF en sangre periférica de un grupo de individuos con melanoma, comparándola con un grupo depersonas sin cáncer y en algunas líneas celulares.Materiales y métodos:Se extrajo ARN de 31 muestras de sangre periférica: 19 de pacientes con melanoma y 12 de personassin ningún tipo de cáncer. Se cuantificaron niveles de expresión tanto para el gen MITF como para los genes de expresiónconstitutiva (?2M y GAPDH) mediante PCR tiempo real. Asimismo se evaluó la expresión de los mismos genes en cinco líneascelulares.Resultados:En todos los individuos se observó expresión del gen MITF, aunque no hubo diferencias estadísticamentesignificativas entre los niveles de expresión en los grupos de estudio (p=0.09). Sin embargo, la expresión de MITF en el grupode pacientes con melanoma fue más variable que la observada en el grupo de personas sin cáncer. Asimismo, en la línea celularde adenocarcinoma gástrico se detectó expresión del gen MITF, no descrita hasta el momento.Conclusiones:Se encontraronniveles de expresión del gen MITF en sangre periférica tanto de personas con melanomacomo en personas sin cáncer. Sin embargo, la variabilidad en los niveles de expresión del gen MITF observados en personascon melanoma, sugiere la posible presencia de células tumorales en circulaciónBackground:The incidence of malign melanoma tumours has increased more rapidly than any other type of cancer; thishas intensified the search for tools that facilitate early identification of melanoma. Microphthalmia associated transcriptionfactor (MITF) is currently known as being a master melanocyte regulator; we analyse MITF gene expression in peripheral bloodfrom individuals suffering from melanoma, compared to people without any type of cancer and ones cell lines.Materials and methods:Thirty one samples of peripheral blood were used: 19 from patients having melanoma and 12 frompeople without any cancer. RNA was then extracted from these samples. MITF and housekeeping genes (?2M and GAPDH)expression levels were then quantified by real-time PCR. Five cell lines were also used to determine the MITF expressionResults:MITF gene expression could be observed in all individuals, though no statistically significant differences werefound among expression levels in the groups studied (p=0.09). Even so, MITF expression in the group of patients sufferingfrom melanoma was much more variable than that observed in the group of cancer-free people. Expression was detected inthe cell line AGS (gastric adenocarcinoma), not yet described. Conclusions: MITF gene expression levels were detectedin the peripheral blood of both people suffering from melanomaand people without any type of cancer. However, variabilityin the number of molecules in MITF gene expression wasobserve in people with melanoma, this suggest the presenceof tumour cells in circulatio

Integrative analysis of global gene expression identifies opposite patterns of reactive astrogliosis in aged human prefrontal cortex

The prefrontal cortex (PFC) is one of the brain regions with more prominent changes in human aging. The molecular processes related to the cognitive decline and mood changes during aging are not completely understood. To improve our knowledge, we integrated transcriptomic data of four studies of human PFC from elderly people (58–80 years old) compared with younger people (20–40 years old) using a meta-analytic approximation combined with molecular signature analysis. We identified 1817 differentially expressed genes, 561 up-regulated and 1256 down-regulated. Pathway analysis revealed down-regulation of synaptic genes with conservation of gene expression of other neuronal regions. Additionally, we identified up-regulation of markers of astrogliosis with transcriptomic signature compatible with A1 neurotoxic astrocytes and A2 neuroprotective astrocytes. Response to interferon is related to A1 astrocytes and the A2 phenotype is mediated in aging by activation of sonic hedgehog (SHH) pathway and up-regulation of metallothioneins I and genes of the family ERM (ezrin, radixin, and moesin). The main conclusions of our study are the confirmation of a global dysfunction of the synapses in the aged PFC and the evidence of opposite phenotypes of astrogliosis in the aging brain, which we report for the first time in the present article. © 2018 by the authors. Licensee MDPI, Basel, Switzerland

Expression of the major and pro-oncogenic H3K9 lysine methyltransferase SETDB1 in non-small cell lung cancer

SETDB1 is a key histone lysine methyltransferase involved in gene silencing. The SETDB1 gene is amplified in human lung cancer, where the protein plays a driver role. Here, we investigated the clinical significance of SETDB1 expression in the two major forms of human non-small cell lung carcinoma (NSCLC), i.e., adenocarcinoma (ADC) and squamous cell carcinoma (SCC), by combining a meta-analysis of transcriptomic datasets and a systematic review of the literature. A total of 1140 NSCLC patients and 952 controls were included in the association analyses. Our data revealed higher levels of SETDB1 mRNA in ADC (standardized mean difference, SMD: 0.88; 95% confidence interval, CI: 0.73–1.02; p less than 0.001) and SCC (SMD: 0.40; 95% CI: 0.13–0.66; p = 0.003) compared to non-cancerous tissues. For clinicopathological analyses, 2533 ADC and 903 SCC patients were included. Interestingly, SETDB1 mRNA level was increased in NSCLC patients who were current smokers compared to non-smokers (SMD: 0.26; 95% CI: 0.08–0.44; p = 0.004), and when comparing former smokers and non-smokers (p = 0.009). Furthermore, the area under the curve (AUC) given by the summary receiver operator characteristic curve (sROC) was 0.774 (Q = 0.713). Together, our findings suggest a strong foundation for further research to evaluate SETDB1 as a diagnostic biomarker and/or its potential use as a therapeutic target in NSCLC. © 2019 by the authors. Licensee MDPI, Basel, Switzerland

Quantification of cell-free DNA for evaluating genotoxic damage from occupational exposure to car paints

cfDNA concentrations and type of comet in the exposed individuals - grouped by car paint shops - and BTX concentrations in the indoor air. Table S2. Socio-demographic data of the exposed cohort. Table S3. Socio-demographic data of the non-exposed cohort. Table S4. Exposed cohort. cfDNA and total count and types of comet. Table S5. Non-exposed cohort. cfDNA and total count and types of comet. Table S6. Air borne solvents concentrations in workshops. Table S7. Exposed cohort. Comet score data. Table S8. Non-exposed cohort. Comet score data. (DOCX 73 kb

Quantification of cell-free DNA for evaluating genotoxic damage from occupational exposure to car paints

Background: For several years, cell-free DNA has been emerging as an important biomarker for non-invasive diagnostic in a wide range of clinical conditions and diseases. The limited information available on the genotoxic effects associated with occupational exposure to car paints, as well as the fact that up-to-date there are not reports about cell-free DNA measurements for assessing this condition, led us to evaluate the DNA damage caused by the occupational exposure to organic solvents contained in car paints, through the quantification of the cell-free DNA and the comet assay, in a sample of 33 individuals taken from 10 automobile paint shops located in Bogota DC, Colombia. Results: By applying the two methods, cell-free DNA and comet assay, we found a significant increase in the extent of DNA damage in the exposed individuals compared with the non-exposed ones within the control group. Conclusions: Our findings provide useful information about the cell-free DNA levels in this type of exposure and can be considered as a support tool that contributes to the diagnosis of genotoxic damage in individuals occupationally exposed to car paints. © 2016 The Author(s)

Differences and homologies of chromosomal alterations within and between breast cancer cell lines : A clustering analysis

Background: The MCF7 (ER+/HER2-), T47D (ER+/HER2-), BT474 (ER+/HER2+) and SKBR3 (ER-/HER2+) breast cancer cell lines are widely used in breast cancer research as paradigms of the luminal and HER2 phenotypes. Although they have been subjected to cytogenetic analysis, their chromosomal abnormalities have not been carefully characterized, and their differential cytogenetic profiles have not yet been established. In addition, techniques such as comparative genomic hybridization (CGH), microarray-based CGH and multiplex ligation-dependent probe amplification (MLPA) have described specific regions of gains, losses and amplifications of these cell lines; however, these techniques cannot detect balanced chromosomal rearrangements (e.g., translocations or inversions) or low frequency mosaicism. Results: A range of 19 to 26 metaphases of the MCF7, T47D, BT474 and SKBR3 cell lines was studied using conventional (G-banding) and molecular cytogenetic techniques (multi-color fluorescence in situ hybridization, M-FISH). We detected previously unreported chromosomal changes and determined the content and frequency of chromosomal markers. MCF7 and T47D (ER+/HER2-) cells showed a less complex chromosomal make up, with more numerical than structural alterations, compared to BT474 and SKBR3 (HER2+) cells, which harbored the highest frequency of numerical and structural aberrations. Karyotype heterogeneity and clonality were determined by comparing all metaphases within and between the four cell lines by hierarchical clustering. The latter analysis identified five main clusters. One of these clusters was characterized by numerical chromosomal abnormalities common to all cell lines, and the other four clusters encompassed cell-specific chromosomal abnormalities. T47D and BT474 cells shared the most chromosomal abnormalities, some of which were shared with SKBR3 cells. MCF7 cells showed a chromosomal pattern that was markedly different from those of the other cell lines. Conclusions: Our study provides a comprehensive and specific characterization of complex chromosomal aberrations of MCF7, T47D, BT474 and SKBR3 cell lines.The chromosomal pattern of ER+/HER2- cells is less complex than that of ER+/HER2+ and ER-/HER2+ cells. These chromosomal abnormalities could influence the biologic and pharmacologic response of cells. Finally, although gene expression profiling and aCGH studies have classified these four cell lines as luminal, our results suggest that they are heterogeneous at the cytogenetic level. © 2014Rondón-Lagos et al.; licensee BioMed Central Ltd

Study of the MTHFR C677T polymorphism in newborns with isolated congenital heart defects, in a Colombian population

Introducción: El estudio del papel de polimorfismos en genes de las vías metabólicas de la homocisteína-metionina y el ácido fólico en anomalías congénitas, es cada vez más importante debido a que sus efectos podrían ser modulados. Objetivo: Determinar si la presencia del polimorfismo C677T en el gen de la metilentetrahidrofolato reductasa (MTHFR) se asocia con el desarrollo de cardiopatías congénitas aisladas. Métodos: Se compararon las frecuencias alélicas y genotípicas del polimorfismo en 34 recién nacidos con cardiopatías congénitas aisladas y en 102 individuos sanos. La genotipificación se hizo mediante la reacción en cadena de la polimerasa (PCR) y se determinó el genotipo por medio de la técnica de polimorfismo de longitud de los fragmentos de restricción (RFLP). Resultados: No se encontraron diferencias estadísticamente significativas en las frecuencias alélicas ni genotípicas entre los grupos de casos y controles. Sin embargo, se observó una tendencia estadística para un posible efecto protector del genotipo TT.The research of the role of gene polymorphisms in the metabolic pathways of homocysteine-methionine and folic acid in congenital malformations is very important because its effect could be modulated. Objetive: The aim of this study was to determine whether the C677T polymorphism in the gene of the enzyme methylenetetrahydrofolate reductase (MTHFR) was associated with the development of isolated congenital heart disease. Methodology: We compared the allele and genotype frequencies of this polymorphism in 34 infants with isolated congenital heart defects and 102 healthy individuals. Genotyping was performed by Polymerase Chain Reaction (PCR) and with the technique Restriction Fragment Length Polymorphism (RFLP). Results: There were no statistically significant differences in allele or genotype frequencies between case and control groups. Although our results show no statistically significant differences between the groups assessed there was a statistical trend for a possible protective effect of TT genotype against the development of congenital heart disease

Study of MTHFR C677T polymorphism in neonates with isolated congenital heart disease in a Colombian population

Introducción: El estudio del papel de polimorfismos en genes de las vías metabólicas de la homocisteína-metionina y el ácido fólico en anomalías congénitas, es cada vez más importante debido a que sus efectos podrían ser modulados.Objetivo: Determinar si la presencia del polimorfismo C677T en el gen de la metilentetrahidrofolato reductasa (MTHFR) se asocia con el desarrollo de cardiopatías congénitas aisladas.Métodos: Se compararon las frecuencias alélicas y genotípicas del polimorfismo en 34 recién nacidos con cardiopatías congénitas aisladas y en 102 individuos sanos. La genotipificación se hizo mediante la reacción en cadena de la polimerasa (PCR) y se determinó el genotipo por medio de la técnica de polimorfismo de longitud de los fragmentos de restricción (RFLP).Resultados: No se encontraron diferencias estadísticamente significativas en las frecuencias alélicas ni genotípicas entre los grupos de casos y controles. Sin embargo, se observó una tendencia estadística para un posible efecto protector del genotipo TT.Artículo original269-277The research of the role of gene polymorphisms in the metabolic pathways of homocysteine-me-thionine and folic acid in congenital malformations is very important because its effect could be modulated. Objetive: The aim of this study was to determine whether the C677T polymorphism in the gene of the enzyme methylenetetrahydrofolate reductase (NÍTHFR) was associated with the development of isolated congenital heart disease. Methodology: We compared the allele and genotype frequencies of this polymorphism in 34 infants with isolated congenital heart defects and 102 healthy individuals. Genotyping was performed by Polymerase Chain Reaction (PCR) and with the technique Restriction Fragment Length Polymorphism (RFLP). Results: There were no statistically significant differences in allele or genotype frequencies between case and control groups. Although our results show no statistically significant differences between the groups assessed there was a statistical trend for a possible protective effect of TT genotype against the development of congenital heart disease

Cytogenetic study in peripheral blood of patients with melanoma

En Colombia el melanoma es la principal causa de muerte por enfermedades dermatológicas (40%) y representa el 1% del total de muertes por cáncer. El rápido incremento en la incidencia del melanoma hace necesaria la realización de estudios que permitan entender mejor los mecanismos implicados en su génesis y progresión. En este estudio se determinaron anomalías cromosómicas en sangre periférica de 30 pacientes con melanoma y en 23 individuos control mediante Citogenética Convencional (Bandeo G), observándose alta incidencia de anomalías numéricas y baja incidencia de rearreglos estructurales recurrentes, siendo las pérdidas cromosómicas las alteraciones prevalentes en todos los estadíos tumorales estudiados. El análisis citogenético de los pacientes mostró que, los cromosomas X, 9 y 17 fueron los más frecuentemente afectados. De las anomalías numéricas las monosomías de los cromosomas X y 17 y la trisomía formada por un cromosoma marcador fueron las más frecuentes, en estadíos tempranos y tardíos de la enfermedad. Deleciones y translocaciones se presentaron como anomalías únicas. En el grupo control ningún tipo de anomalía fue identificada, y se observó bajo porcentaje de fragilidades en comparación con el grupo de pacientes. En comparación con los controles se observó alta frecuencia de anomalías cromosómicas en los pacientes, lo que sugiere la existencia de heterogeneidad y predisposición genética en el desarrollo de la enfermedad, que con investigaciones adicionales deben ser analizadas y validadas como posibles fuentes de marcadores moleculares, útiles para el diagnóstico temprano, tratamiento y seguimiento de la enfermedad.mong all the skin diseases, melanoma is the main cause of death in Colombia (40 %) and it represents 1 % of all deaths by cancer. Due to the fast increase in the incidence of melanoma, it is necessary to carry out research on the mechanisms involved in its genesis and progression. This study determined chromosomal anomalies from peripheral blood samples on 30 patients with melanoma and on 23 control subjects using conventional cytogenetics (G Banded), where a high incidence in numerical anomalies and a low incidence in recurrent structural rearrangements were observed. Chromosomic losses were prevalent in all the tumor stages studied. The analysis showed that the chromosomes X, 9 and 17 were mainly affected. Among the numerical anomalies, monosomies in X and 17 chromosomes, as well as trisomies formed by a marker chromosome, were the most common in both early and late stages of the disease. Deletions and chromosomal crossovers appeared to be as isolated anomalies. In the control group no anomaly was identified, and a low percentage of fragility was observed when compared with the patients group. A high frequency in chromosomal anomalies was observed in patients, in contrast with the control subjects. This suggests the existence of heterogeneity and genetic predisposition during the illness development. To further research, these must be analyzed and validated as possible sources of molecular markers, which could be of use for the early diagnosis, treatment and follow up of the disease