Adherencia a la participación en un trabajo grupal voluntario en una asignatura de licenciatura Biología

En el plan Bolonia el trabajo grupal es una metodología de aprendizaje que está debidamente protocolizada; no así en los planes anteriores. Nuestro objetivo fue valorar la adherencia en los últimos cinco años a la participación en un trabajo grupal de carácter voluntario en la asignatura optativa Métodos y Técnicas en Biopatología Clínica de la licenciatura de Biología. Características del trabajo grupal: voluntario; grupos reducidos; a realizar fuera de las horas de clase; tema libre dentro del contexto de la asignatura; entregar un documento escrito; realizar una presentación pública. Medidas para estimular la participación: hasta un punto extra sobre la nota final de la asignatura. El número medio de alumnos matriculados en la asignatura fue de 32. No presentados 21%. La participación media en el trabajo grupal fue del 74% sobre el total de alumnos y 91% considerando solamente los alumnos que se presentaban al examen de la asignatura. El nivel de calidad los trabajos fue evaluado entre bueno y excelente. Nuestros resultados ponen de manifiesto un gran nivel de aceptación y el buen hacer en un trabajo grupal voluntario, desvelando un carácter proactivo por parte del alumno hacia una metodología que ofrece grandes ventajas en su formación

The TGFBR1*6A allele is not associated with susceptibility to colorectal cancer in a Spanish population: a case-control study

Background: TGF-β receptor type I is a mediator of growth inhibitory signals. TGFBR1*6A (rs11466445) is a common polymorphic variant of the TGF-β receptor I gene and has been associated with tumour susceptibility. Nevertheless, the role of this polymorphism as a risk factor for colorectal cancer is controversial. The aim of this study was to assess the association between TGFBR1*6A and colorectal cancer, age, sex, tumour location and tumour stage in a Spanish population. Methods: The case-control study involved 800 Spanish subjects: 400 sporadic colorectal cancer patients and 400 age-, sex-, and ethnic-matched controls. The odds ratio (OR) and 95% confidence interval (95% CI) for the TGFBR1*6A polymorphism were calculated using unconditional logistic regression adjusted for age and sex. Analysis of somatic mutations at the GCG repeat of TGFBR1 exon 1 and germline allele-specific expression were also conducted to obtain further information on the contribution of the TGFBR1*6A allele to CRC susceptibility. Results: There was no statistically significant association between the TGFBR1*6A allele and CRC (p > 0.05). The OR was 1.147 (95% CI: 0.799–1.647) for carriers of the TGFBR1*6A allele and 0.878 (95% CI: 0.306–2.520) for homozygous TGFBR1*6A individuals compared with the reference. The frequency of the polymorphism was not affected by age, sex or tumour stage. The TGFBR1*6A allele was more prevalent among colon tumour patients than among rectal tumour patients. Tumour somatic mutations were found in only two of 69 cases (2.9%). Both cases involved a GCG deletion that changed genotype 9A/9A in normal DNA to genotype 9A/8A. Interestingly, these two tumours were positive for microsatellite instability, suggesting that these mutations originated because of a deficient DNA mismatch repair system. Allele-specific expression of the 9A allele was detected in seven of the 14 heterozygous 9A/6A tumour cases. This could have been caused by linkage disequilibrium of the TGFBR1*6A allele with mutations that cause allele-specific expression, as was recently suggested. Conclusion: Our results suggest that the TGFBR1*6A allele does not confer an increased risk of colorectal cancer in the Spanish population.The research was supported in part by grants from the Generalitat Valenciana in Spain (AP106/06) and the Biomedical Research Foundation of the Hospital of Elche (FIBElx-02/2007). T.M-B was a recipient of a fellowship from the Spanish Society of Medical Oncology

Co‐occurrence of germline pathogenic variants for different hereditary cancer syndromes in patients with Lynch syndrome

Background: Lynch syndrome (LS) is a hereditary condition characterized by a high risk of colorectal cancer, endometrial cancer, and other neoplasia associated with germline alterations in DNA mismatch repair genes. The classical genetic diagnostic strategy for LS consists of the Sanger sequencing of genes associated with the suspected syndrome. Next‐generation sequencing (NGS) enables the simultaneous sequencing of a large number of hereditary cancer genes. Here, we aimed to study whether other germline pathogenic variants of hereditary cancer genes are present in patients with LS. Methods: A cohort of 84 probands with a previous genetic diagnosis of LS by Sanger sequencing was reanalyzed using NGS via a commercial panel of 94 hereditary cancer genes by hybrid capture. The American College of Medical Genetics and Genomics criteria were used to classify the clinical significance of the variants. The findings of NGS were confirmed by Sanger sequencing. When possible, genetic analyses of the new findings in the proband's relatives were also performed by Sanger sequencing. Results: We identified five families (6%), out of 84, with at least two germline pathogenic variants conferring to high or moderate risk in different dominant cancer‐predisposing genes: [MLH1‐BRCA2‐NBN], [MLH1‐BRCA1], [MSH2‐ATM], [MSH6‐NF1], and [MLH1‐FANCA]. Interestingly, only one out of these five families exhibited a clinical phenotype associated with the new pathogenic variants. The family with three pathogenic variants of the [MLH1‐BRCA2‐NBN] genes showed a high aggregation of tumors associated with LS and breast and ovarian cancer syndrome. Conclusions: Our results showed that the co‐occurrence of more than one pathogenic variant in cancer‐predisposing genes was remarkable among cases of LS. In most cases, no clinicial manifestations were associated with the secondary pathogenic variants. Further studies are needed to confirm these findings and elucidate their clinical impact. Reanalysis of LS families should be considered only in families with mixed clinical phenotypes.RFA is a recipient of a Fellowship from the Consellería Educación of the Valencian Community. The studies of ED and ACA are funded by the Acción Juvenil from Consellería Educación of the Valencian Community and the Spanish Ministry of Economy and Competitiveness, respectively. VDO is a recipient of a Fellowship from the Spanish Association Against Cancer (AECC). This work was supported by grants from the Foundation for the Promotion of Health and Biomedical Research in the Valencian Region, FISABIO (Grants: UGP‐14‐192 and UGP‐16‐146), and the Carlos III Health Institute (Grant AES: PI17/01082)

Optimizing polymer lab-on-chip platforms for ultrasonic manipulation: Influence of the substrate

The choice of substrate material in a chip that combines ultrasound with microfluidics for handling biological and synthetic microparticles can have a profound effect on the performance of the device. This is due to the high surface-to-volume ratio that exists within such small structures and acquires particular relevance in polymer-based resonators with 3D standing waves. This paper presents three chips developed to perform particle flow-through separation by ultrasound based on a polymeric SU-8 layer containing channelization over three different substrates: Polymethyl methacrylate (PMMA); Pyrex; and a cracked PMMA composite-like structure. Through direct observations of polystyrene microbeads inside the channel, the three checked chips exhibit their potential as disposable continuous concentration devices with different spatial pressure patterns at frequencies of resonance close to 1 Mhz. Chips with Pyrex and cracked PMMA substrates show restrictions on the number of pressure nodes established in the channel associated with the inhibition of 3D modes in the solid structure. The glass-substrate chip presents some advantages associated with lower energy requirements to collect particles. According to the results, the use of polymer-based chips with rigid substrates can be advantageous for applications that require short treatment times (clinical tests handling human samples) and low-cost fabrication. © 2015 by the authors; licensee MDPI, Basel, Switzerland.The study has been performed in the framework of two Spanish National Research Project BIO2011-30535-C04-01,02,03, “Development of a high throughput for isolation of tumor cells circulating in peripheral blood”.We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).Peer Reviewe

HGUE-C-1 is an atypical and novel colon carcinoma cell line

Background: Colorectal carcinoma is a common cause of cancer. Adjuvant treatments include: 5-fluorouracil administered together with folinic acid, or more recently, oral fluoropyrimidines such as capecitabine, in combination with oxaliplatin or irinotecan. Metastatic colorectal cancer patients can benefit from other additional treatments such as cetuximab or bevacizumab. Methods: Using cell culture techniques, we isolated clonal populations from primary cultures of ascitic effusion derived from a colon cancer patient and after several passages an established cell line, HGUE-C-1, was obtained. Genetic analysis of HGUE-C-1 cells was performed by PCR of selected exons and sequencing. Cell proliferation studies were performed by MTT assays and cell cycle analyses were performed by flow cytometry. Retinoblastoma activity was measured by luciferase assays and proteins levels and activity were analysed by Western blot or immunohistochemistry. Results: We have established a new cell line from ascitic efussion of a colon cancer patient who did not respond to 5-fluorouracil or irinotecan. HGUE-C-1 cells did not show microsatellite instability and did not harbour mutations in KRAS, BRAF, PI3KCA or TP53. However, these cells showed loss of heterozygosity affecting Adenomatous Polyposis Coli and nuclear staining of β-catenin protein. The HGUE-C-1 cell line was sensitive to erlotinib, gefitinib, NVP-BEZ235, rapamycin and trichostatin, among other drugs, but partially resistant to heat shock protein inhibitors and highly resistant to AZD-6244 and oxaliplatin, even though the patient from which this cell line was derived had not been exposed to these drugs. Molecular characterization of this cell line revealed low expression levels and activity of Retinoblastoma protein and elevated basal levels of Erk1/2 activity and p70S6K expression and activity, which may be related to chemoresistance mechanisms. Conclusions: HGUE-C-1 represents a novel and peculiar colon carcinoma model to study chemoresistance to chemotherapeutic agents and to novel anti-neoplasic drugs that interrupt signalling pathways such as the APC/βcatenin, Ras/Raf/Mek/Erk, PI3K/mTOR/p70S6K pathways as well as histone regulation mechanisms.This article has been funded by grants from the Instituto de Salud Carlos III FIS PI080901and FIS PI01202025 to Miguel Saceda

Prevalence of Lynch Syndrome among Patients with Newly Diagnosed Endometrial Cancers

Background: Lynch syndrome (LS) is a hereditary condition that increases the risk for endometrial and other cancers. The identification of endometrial cancer (EC) patients with LS has the potential to influence life-saving interventions. We aimed to study the prevalence of LS among EC patients in our population. Methods: Universal screening for LS was applied for a consecutive series EC. Tumor testing using microsatellite instability (MSI), immunohistochemistry (IHC) for mismatch-repair (MMR) protein expression and MLH1-methylation analysis, when required, was used to select LS-suspicious cases. Sequencing of corresponding MMR genes was performed. Results: One hundred and seventy-three EC (average age, 63 years) were screened. Sixty-one patients (35%) had abnormal IHC or MSI results. After MLH1 methylation analysis, 27 cases were considered suspicious of LS. From these, 22 were contacted and referred for genetic counseling. Nineteen pursued genetic testing and eight were diagnosed of LS. Mutations were more frequent in younger patients (<50 yrs). Three cases had either intact IHC or MSS and reinforce the need of implement the EC screening with both techniques. Conclusion: The prevalence of LS among EC patients was 4.6% (8/173); with a predictive frequency of 6.6% in the Spanish population. Universal screening of EC for LS is recommended.This study was supported by Conselleria Sanidad Comunidad Valenciana, Spain (AP/177/10) (http://www.san.gva.es/); Biomedical Research Foundations of the Alicante University Hospital (PI14/2006 and NI02/2011) (http://www.dep19.san.gva.es/); and the Elche University Hospital, Spain (FIBElx-CO11/03) (http://www.dep20.san.gva.es/)

Genetic and clinical characterization of a novel FH founder mutation in families with hereditary leiomyomatosis and renal cell cancer syndrome

Hereditary leiomyomatosis and renal cell cancer syndrome is a rare autosomal dominant hereditary syndrome. Previously, we published the largest cohort of FH mutation carriers in Spain and observed a highly recurrent missense heterozygous variant, FH(NM_000143.4):c.1118A > G p.(Asn373Ser), in 104 individuals from 31 apparently unrelated families. Here, we aimed to establish its founder effect and characterize the associated clinical phenotype. Results Haplotype analysis confirmed that families shared a common haplotype (32/38 markers) spanning 0.61-0.82 Mb, indicating this recurrent variant was inherited from a founder ancestor. Cutaneous and uterine leiomyomatosis were diagnosed in 64.6% (64/99) and 98% (50/51) of patients, respectively, and renal cell cancer was present in 10.4% (10/96). The pathogenic FH_c.1118A > G variant is a Spanish founder mutation that originated 12-26 generations ago. We estimate that the variant may have appeared between 1370 and 1720. Individuals carrying this founder mutation had similar frequency of renal cell cancer and a higher frequency of renal cysts and leiomyomas than those in other cohorts of this syndrome. Conclusions In the Spanish province of Alicante there is a high prevalence of HLRCC because of the founder mutation FH c.1118A > G; p.(Asn373Ser). The characterization of founder mutations provides accurate and specific information regarding their penetrance and expressivity. In individuals with suspected HLRCC from the province of Alicante, genetic testing by direct analysis of the founder FH c.1118A > G; p.(Asn373Ser) mutation may be a faster and more efficient diagnostic tool compared with complete gene sequencing

The heritability and patterns of DNA methylation in normal human colorectum

DNA methylation (DNAm) has been linked to changes in chromatin structure, gene expression and disease. The DNAm level can be affected by genetic variation; although, how this differs by CpG dinucleotide density and genic location of the DNAm site is not well understood. Moreover, the effect of disease causing variants on the DNAm level in a tissue relevant to disease has yet to be fully elucidated. To this end, we investigated the phenotypic profiles, genetic effects and regional genomic heritability for 196080 DNAm sites in healthy colorectum tissue from 132 unrelated Colombian individuals. DNAm sites in regions of low-CpG density were more variable, on average more methylated and were more likely to be significantly heritable when compared with DNAm sites in regions of high-CpG density. DNAm sites located in intergenic regions had a higher mean DNAm level and were more likely to be heritable when compared with DNAm sites in the transcription start site (TSS) of a gene expressed in colon tissue. Within CpG-dense regions, the propensity of the DNAm level to be heritable was lower in the TSS of genes expressed in colon tissue than in the TSS of genes not expressed in colon tissue. In addition, regional genetic variation was associated with variation in local DNAm level no more frequently for DNAm sites within colorectal cancer risk regions than it was for DNAm sites outside such regions. Overall, DNAm sites located in different genomic contexts exhibited distinguishable profiles and may have a different biological function

Reflexiones sobre el aprendizaje de la histología en biología y en ciencias de la salud

El conocimiento de la estructura, composición y función del organismo es fundamental en la formación de los graduados de Ciencias Biomédicas y de Ciencias de la Salud. Las células y los tejidos son los componentes básicos para comprender el normal funcionamiento del organismo y sus procesos patológicos. Por otra parte, la Histología es una de las disciplinas básicas con mayor cantidad de publicaciones sobre temas relacionados con su docencia. Sin embargo, a pesar de la abrumadora literatura existente, todavía no hay criterios claros sobre la pertinencia de los contenidos, los métodos en el aprendizaje y la evaluación de esta materia. En este trabajo; tras una revisión bibliográfica y a partir de la información aportada por profesores de Histología, se lo analizan las principales cuestiones que plantea el proceso de enseñanza aprendizaje de la Histología en el ámbito de Biomedicina (Biología, Medicina, Veterinaria, Enfermería, Nutrición, Fisioterapia, Biotecnología, etc.), a saber: su pertinencia, los contenidos a impartir, las estrategias y los medios didácticos a emplear y los métodos de evaluación. A partir de todos estos datos se describen los principales puntos fuertes y débiles de la Histología con las correspondientes sugerencias de cambio y adaptación al entorno actual

TGFBR1 Intralocus Epistatic Interaction as a Risk Factor for Colorectal Cancer

In colorectal cancer (CRC), an inherited susceptibility risk affects about 35% of patients, whereas high-penetrance germline mutations account for <6% of cases. A considerable proportion of sporadic tumors could be explained by the coinheritance of multiple low-penetrance variants, some of which are common. We assessed the susceptibility to CRC conferred by genetic variants at the TGFBR1 locus. We analyzed 14 polymorphisms and the allele-specific expression (ASE) of TGFBR1 in 1025 individuals from the Spanish population. A case-control study was undertaken with 504 controls and 521 patients with sporadic CRC. Fourteen polymorphisms located at the TGFBR1 locus were genotyped with the iPLEX Gold (MassARRAY-Sequenom) technology. Descriptive analyses of the polymorphisms and haplotypes and association studies were performed with the SNPator workpackage. No relevant associations were detected between individual polymorphisms or haplotypes and the risk of CRC. The TGFBR1*9A/6A polymorphism was used for the ASE analysis. Heterozygous individuals were analyzed for ASE by fragment analysis using cDNA from normal tissue. The relative level of allelic expression was extrapolated from a standard curve. The cutoff value was calculated with Youden's index. ASE was found in 25.4% of patients and 16.4% of controls. Considering both bimodal and continuous types of distribution, no significant differences between the ASE values of patients and controls were identified. Interestingly, a combined analysis of the polymorphisms and ASE for the association with CRC occurrence revealed that ASE-positive individuals carrying one of the most common haplotypes (H2: 20.7%) showed remarkable susceptibility to CRC (RR: 5.25; 95% CI: 2.547–5.250; p<0.001) with a synergy factor of 3.7. In our study, 54.1% of sporadic CRC cases were attributable to the coinheritance of the H2 haplotype and TGFBR1 ASE. These results support the hypothesis that the allelic architecture of cancer genes, rather than individual polymorphisms, more accurately defines the CRC risk