Magnetic solid-phase extraction based on restricted-access molecularly imprinted polymers for ultrarapid determination of ractopamine residues from food samples by capillary electrophoresis.

An ultrafast, efficient, and eco-friendly method combining magnetic solid phase extraction and capillary electrophoresis with diode array detection have been developed to determine ractopamine residues in food samples. A restricted access material based on magnetic and mesoporous molecularly imprinted polymer has been properly synthesized and characterized, demonstrating excellent selectivity and high adsorbent capacity. Short-end injection capillary electrophoresis method was optimized: 75 mM triethylamine pH 7 as BGE, -20 kV, 50 mbar by hydrodynamic injection during 8 s, and capillary temperature at 25 °C; reaching ultrafast ractopamine analysis (∼0.6 min) with good peak asymmetry, and free from interfering and/or baseline noise. After sample preparation optimization, the conditions were: 1000 µL of sample at pH 6, 20 mg of adsorbent, stirring time of 120 s, 250 µL of ultrapure water as washing solvent, 1000 µL of methanol: acetic acid (7: 3, v/v) as eluent, and the adsorbent can be reused four times. In these conditions, the analytical method showed recoveries around to 100 %, linearity ranged from 9.74 to 974.0 µg kg-1, correlation coefficient (r) ≥ 0,99 in addition to adequate precision, accuracy, and robustness. After proper validation, the method was successfully applied in the analysis ractopamine residues in bovine milk and bovine and porcine muscle.

Diving dinosaurs? Caveats on the use of bone compactness and pFDA for inferring lifestyle.

The lifestyle of spinosaurid dinosaurs has been a topic of lively debate ever since the unveiling of important new skeletal parts for Spinosaurus aegyptiacus in 2014 and 2020. Disparate lifestyles for this taxon have been proposed in the literature; some have argued that it was semiaquatic to varying degrees, hunting fish from the margins of water bodies, or perhaps while wading or swimming on the surface; others suggest that it was a fully aquatic underwater pursuit predator. The various proposals are based on equally disparate lines of evidence. A recent study by Fabbri and coworkers sought to resolve this matter by applying the statistical method of phylogenetic flexible discriminant analysis to femur and rib bone diameters and a bone microanatomy metric called global bone compactness. From their statistical analyses of datasets based on a wide range of extant and extinct taxa, they concluded that two spinosaurid dinosaurs (S. aegyptiacus, Baryonyx walkeri) were fully submerged "subaqueous foragers," whereas a third spinosaurid (Suchomimus tenerensis) remained a terrestrial predator. We performed a thorough reexamination of the datasets, analyses, and methodological assumptions on which those conclusions were based, which reveals substantial problems in each of these areas. In the datasets of exemplar taxa, we found unsupported categorization of taxon lifestyle, inconsistent inclusion and exclusion of taxa, and inappropriate choice of taxa and independent variables. We also explored the effects of uncontrolled sources of variation in estimates of bone compactness that arise from biological factors and measurement error. We found that the ability to draw quantitative conclusions is limited when taxa are represented by single data points with potentially large intrinsic variability. The results of our analysis of the statistical method show that it has low accuracy when applied to these datasets and that the data distributions do not meet fundamental assumptions of the method. These findings not only invalidate the conclusions of the particular analysis of Fabbri et al. but also have important implications for future quantitative uses of bone compactness and discriminant analysis in paleontology.

Estudio observacional de pacientes de una Unidad de Lípidos en tratamiento hipolipemiante para prevención primaria y secundaria: Estudio ULFI / Observational study of patients from a Lipid Unit on lipid-modifying therapy for primary and secondary prevention: ULFI Study

Objetivos: Evaluar la consecución de los objetivos de colesterol unido a lipoproteínas de baja densidad (cLDL) establecidos por las guías europeas de manejo de las dislipemias de 2019 y de prevención cardiovascular de 2021, describir el tratamiento hipolipemiante realizado, analizar el logro de los objetivos según el tratamiento hipolipemiante recibido y estudiar los factores asociados al éxito terapéutico. Diseño: Estudio observacional con 185 pacientes de ambos sexos de 18 años o más en tratamiento hipolipemiante para prevención primaria o secundaria, atendidos en la Unidad de Lípidos. Resultados: El 62,1% de los pacientes presentó un riesgo cardiovascular (RCV) muy alto según la guía de 2019, y el 60,5% según la de 2021. Del total de casos, el 22,7% logró un control adecuado del cLDL según la guía de 2019 y el 20% lo hizo de acuerdo con la de 2021. El 47,6% de los pacientes recibió tratamiento hipolipemiante de muy alta intensidad y el 14,1% lo recibió de extremadamente alta intensidad. El 76% de los sujetos con muy alto RCV en tratamiento hipolipemiante de extremadamente alta intensidad logró los objetivos terapéuticos de ambas guías. En el análisis multivariante, los factores asociados al éxito terapéutico fueron la presencia de enfermedad cardiovascular arteriosclerótica, la intensidad del tratamiento hipolipemiante, la diabetes mellitus y el consumo bajo o moderado de alcohol. Conclusiones: El control de la dislipemia es mejorable. Los tratamientos hipolipemiantes de alta o extremadamente alta intensidad pueden contribuir a optimizar el control de los pacientes con mayor RCV. (AU)

Objectives: To evaluate the achievement of low-density lipoprotein cholesterol (LDLc) goals established by the 2019 European Guidelines for the Management of Dyslipidemias and 2021 Cardiovascular Disease Prevention Guidelines, describe the lipid-lowering treatment received, analyze the achievement of goals according to the lipid-lowering treatment received and study the factors associated with therapeutic success. Design: Observational study that included 185 patients of both sexes aged 18 or over undergoing lipid-lowering treatment for primary or secondary prevention, attended at the Lipid Unit. Results: 62.1% of the patients had a very high cardiovascular risk (CVR) according to the 2019 guidelines, and 60.5% according to the 2021 guidelines. Of the total cases, 22.7% achieved adequate control of LDLc according to the 2019 guidelines and 20% according to the 2021 guidelines. 47.6% of the patients received very high intensity lipid-lowering treatment, and 14.1% received extremely high intensity lipid-lowering treatment. 76% of subjects with very high CVR on extremely high intensity lipid-lowering treatment achieved the therapeutic objectives of both guides. In the multivariate analysis, factors associated with therapeutic success were the presence of arteriosclerotic cardiovascular disease, the intensity of lipid-lowering treatment, diabetes mellitus, and low to moderate alcohol consumption. Conclusions: Dyslipidemia control is improvable. High or extremely high intensity lipid-lowering treatments can contribute to optimizing control of patients with higher CVR. (AU)

Magnetic and mesoporous molecularly imprinted polymer synthesized by rational computation design: Sample preparation and analysis of ractopamine.

Through density functional theory calculations was studied theoretically the formation process of a magnetic and mesoporous molecularly imprinted polymer for ractopamine (RAC), evaluating the molecular electrostatic potential map, functional monomers, functional monomer / template stoichiometry and crosslink agents. The results revealed that the best conditions for the synthesis were established with acrylic acid as functional monomer in a 1: 4 stoichiometry using acetonitrile as the solvent and ethylene glycol dimethacrylate as crosslink agent. It was observed that nine hydrogen bonds established between the RAC and acrylic acid play a key role on the pre-polymerization complex. In addition, three analytical methods using HPLC, UHPLC and CE instruments were optimized for rapid analysis. The adsorbent was experimentally synthesized considering the best conditions found at the molecular level and characterized by FTIR, DRX, TGA, SEM, TEM, surface analysis, and wettability. After that, the synthesized material was used in magnetic solid phase extraction combined with capillary electrophoresis in a preliminary RAC recovery study from milk samples. Finally, greenness metric with a score of 0.55 have been obtained for the sample preparation procedure using the online AGREEprep metric.

Analysis of the Mutational Landscape of Osteosarcomas Identifies Genes Related to Metastasis and Prognosis and Disrupted Biological Pathways of Immune Response and Bone Development.

Osteosarcoma (OS) is the most prevalent type of bone tumor, but slow progress has been achieved in disentangling the full set of genomic events involved in its initiation and progression. We assessed by NGS the mutational spectrum of 28 primary OSs from Brazilian patients, and identified 445 potentially deleterious SNVs/indels and 1176 copy number alterations (CNAs). TP53 was the most recurrently mutated gene, with an overall rate of ~60%, considering SNVs/indels and CNAs. The most frequent CNAs (~60%) were gains at 1q21.2q21.3, 6p21.1, and 8q13.3q24.22, and losses at 10q26 and 13q14.3q21.1. Seven cases presented CNA patterns reminiscent of complex events (chromothripsis and chromoanasynthesis). Putative RB1 and TP53 germline variants were found in five samples associated with metastasis at diagnosis along with complex genomic patterns of CNAs. PTPRQ, KNL1, ZFHX4, and DMD alterations were prevalent in metastatic or deceased patients, being potentially indicative of poor prognosis. TNFRSF11B, involved in skeletal system development and maintenance, emerged as a candidate for osteosarcomagenesis due to its biological function and a high frequency of copy number gains. A protein-protein network enrichment highlighted biological pathways involved in immunity and bone development. Our findings reinforced the high genomic OS instability and heterogeneity, and led to the identification of novel disrupted genes deserving further evaluation as biomarkers due to their association with poor outcomes.

Observational study of patients from a Lipid Unit on lipid-modifying therapy for primary and secondary prevention: ULFI Study. / Estudio observacional de pacientes de una Unidad de Lípidos en tratamiento hipolipemiante para prevención primaria y secundaria: Estudio ULFI.

OBJECTIVES: To evaluate the achievement of low-density lipoprotein cholesterol (LDLc) goals established by the 2019 European Guidelines for the Management of Dyslipidemias and 2021 Cardiovascular Disease Prevention Guidelines, describe the lipid-lowering treatment received, analyze the achievement of goals according to the lipid-lowering treatment received and study the factors associated with therapeutic success. DESIGN: Observational study that included 185 patients of both sexes aged 18 or over undergoing lipid-lowering treatment for primary or secondary prevention, attended at the Lipid Unit. RESULTS: 62.1% of the patients had a very high cardiovascular risk (CVR) according to the 2019 guidelines, and 60.5% according to the 2021 guidelines. Of the total cases, 22.7% achieved adequate control of LDLc according to the 2019 guidelines and 20% according to the 2021 guidelines. 47.6% of the patients received very high intensity lipid-lowering treatment, and 14.1% received extremely high intensity lipid-lowering treatment. 76% of subjects with very high CVR on extremely high intensity lipid-lowering treatment achieved the therapeutic objectives of both guides. In the multivariate analysis, factors associated with therapeutic success were the presence of arteriosclerotic cardiovascular disease, the intensity of lipid-lowering treatment, diabetes mellitus, and low to moderate alcohol consumption. CONCLUSIONS: Dyslipidemia control is improvable. High or extremely high intensity lipid-lowering treatments can contribute to optimizing control of patients with higher CVR.

Fine-tuning of the spin-crossover properties of Fe(III) complexes via ligand design.

Exploring the chemical space of a given ligand aiming to modulate its ligand field strength is a versatile strategy for the fine-tuning of physical properties such as the transition temperature (T1/2) of spin-crossover (SCO) complexes. The computational study presented herein aims at systematically exploring the extent to which the ligand substituent effects can modulate T1/2 in two families of Fe(III) SCO systems with a N4O2 coordination environment and at identifying the best descriptors for fast and accurate prediction of changes in T1/2 upon ligand functionalization. B3LYP* calculations show that the attachment of substituents to ß-ketoiminato fragments (L1) leads to drastic changes in T1/2, while functionalization of phenolato moieties (L2) allows for a finer degree of control over T1/2. Natural Bond Orbital (NBO) charges of the donor atoms, Hammett parameters for both para and meta-functionalization of L2, and Swain-Lupton parameters for L1 and para-functionalization of L2 have been found to be the suitable descriptors for predicting the changes in T1/2. Further analysis of the ligand-field splitting in such systems rationalizes the observed trends and shows that ligand substituents modify both the σ and π bonds between the Fe(III) center and the ligands. Thus, we provide simple yet reliable guidelines for the rational design of new SCO systems with specific values of T1/2 based on their ligand design.

DNA methylation patterns suggest the involvement of DNMT3B and TET1 in osteosarcoma development.

DNA methylation may be involved in the development of osteosarcomas. Osteosarcomas commonly arise during the bone growth and remodeling in puberty, making it plausible to infer the involvement of epigenetic alterations in their development. As a highly studied epigenetic mechanism, we investigated DNA methylation and related genetic variants in 28 primary osteosarcomas aiming to identify deregulated driver alterations. Methylation and genomic data were obtained using the Illumina HM450K beadchips and the TruSight One sequencing panel, respectively. Aberrant DNA methylation was spread throughout the osteosarcomas genomes. We identified 3146 differentially methylated CpGs comparing osteosarcomas and bone tissue samples, with high methylation heterogeneity, global hypomethylation and focal hypermethylation at CpG islands. Differentially methylated regions (DMR) were detected in 585 loci (319 hypomethylated and 266 hypermethylated), mapped to the promoter regions of 350 genes. These DMR genes were enriched for biological processes related to skeletal system morphogenesis, proliferation, inflammatory response, and signal transduction. Both methylation and expression data were validated in independent groups of cases. Six tumor suppressor genes harbored deletions or promoter hypermethylation (DLEC1, GJB2, HIC1, MIR149, PAX6, and WNT5A), and four oncogenes presented gains or hypomethylation (ASPSCR1, NOTCH4, PRDM16, and RUNX3). Our analysis also revealed hypomethylation at 6p22, a region that contains several histone genes. Copy-number changes in DNMT3B (gain) and TET1 (loss), as well as overexpression of DNMT3B in osteosarcomas provide a possible explanation for the observed phenotype of CpG island hypermethylation. While the detected open-sea hypomethylation likely contributes to the well-known osteosarcoma genomic instability, enriched CpG island hypermethylation suggests an underlying mechanism possibly driven by overexpression of DNMT3B likely resulting in silencing of tumor suppressors and DNA repair genes.

High-dose chemotherapy with autologous stem cell transplantation for patients with extracranial germ cell tumors - experience of two Brazilian pediatric centers.

Malignant extracranial germ cell tumors (GCTs) are rare in pediatric patients and are usually extremely sensitive to chemotherapy. Relapsed or refractory tumors, although rare, established the need for second-line therapies, including high-dose chemotherapy with autologous stem cell transplantation (HDCT/ASCT). However, there are few data on its use in children with GCTs. We present a retrospective analysis of all patients diagnosed with extracranial GCTs who received HDCT/ASCT at two Brazilian pediatric cancer centers from May 1999 to December 2019. We identified a total of 34 patients with a median age at diagnosis of 2.8 years (range, 0 to 18.8), who received HDCT/ASCT. Most patients (73%) received carboplatin, etoposide and melphalan (CEM) as a HDCT regimen. Fourteen patients received a second-line conventional dose chemotherapy (CDCT), 14 received a third-line CDCT and five received even a fourth-line CDCT prior to HDCT/ASCT. After a median follow-up of 22.7 months (range, 0.3 to 198.1), 16 patients had died after tumor relapse/progression and 2 patients died from HDCT/ASCT toxicity. We observed a 5-year OS of 47.1% and 5-year EFS of 44.1%. The 5-year OS for patients referred for HDCT/ASCT with progressive disease was 10% compared to 62.5% for those who achieved disease control before HDCT/ASCT (p = 0.001). In our experience, heavily pretreated children and adolescents with extracranial GCTs achieved considerable survival rates with HDCT/ASCT since, at least, partial control of their disease was possible before starting HDCT/ASCT. The role of HDCT/ASCT in pediatric patients with GCTs should be investigated in prospective trials.

Validación de contenido y fiabilidad del instrumento "Conocimiento, actitud y comportamiento sobre hidratación y reposición de líquidos", en deportistas chilenos

Introducción: Existen estrategias direccionadas a mejorar el rendimiento de los deportistas, algunas estableciendo recomendaciones respecto a la hidratación y reposición de líquidos, con el propósito de asegurar un balance hídrico adecuado en todas las etapas de la práctica deportiva. Sin embargo, existe una carencia de instrumentos adecuados y confiables que permitan explorar esta temática. Objetivo: La presente investigación tiene como objetivo validar por contenido y evaluar la fiabilidad del instrumento "Conocimiento, actitud y comportamiento respecto a la hidratación y reposición de líquidos" en deportistas adolescentes y adultos jóvenes en Chile. Métodos: La adaptación cultural se realizó a través del método Delphi con un panel de expertos en el área, para la evaluación de la fiabilidad se aplicó el cuestionario de manera online a una muestra de 384 deportistas de diferentes disciplinas en dos oportunidades para determinar la estabilidad temporal. Resultados: Todos los ítems lograron porcentajes de acuerdo superior al 80% según el estadístico V de Aiken, en relación con la fiabilidad se obtuvo un Alfa de Cronbach de 0,78 lo que evidencia una consistencia interna aceptable, adicionalmente se obtuvo un Coeficiente de Correlación intraclase de 0,64 entre las dos aplicaciones del cuestionario presentando una fiabilidad regular a buena. Conclusión: Este cuestionario puede ser empleado como una herramienta confiable para ser utilizada en futuras investigaciones en el área de la nutrición deportiva, además como un instrumento que aporta al quehacer de distintos profesionales para apoyar a planificar y evaluar estrategias que propendan al mejoramiento del rendimiento deportivo.

Introduction: There are strategies aimed at improving the performance of athletes, some of which establish recommendations regarding hydration and fluid replacement, with the purpose of ensuring an adequate water balance at all stages of sports practice. However, there is a lack of adequate and reliable instruments to explore this issue. Objective: The present research aims to validate the content and evaluate the reliability of the instrument "Knowledge, attitude and behavior regarding hydration and fluid replacement" among adolescent and young adult athletes in Chile. Methods: The cultural adaptation was carried out through the Delphi method with a panel of experts in the area. For the evaluation of reliability, the questionnaire was applied online to a sample of 384 athletes from different disciplines on two occasions to determine temporal stability. Results: All items achieved percentages of agreement higher than 80% according to Aiken's V statistic, in relation to reliability a Cronbach's Alpha of 0.78 was obtained, which shows an acceptable internal consistency, additionally an intraclass Correlation Coefficient of 0.64 was obtained between the two applications of the questionnaire presenting a regular to good reliability. Conclusion: This questionnaire can be used as a reliable tool for future research around sports nutrition, as well as an instrument that contributes to the work of different professionals to support the planning and evaluation of strategies aimed at improving sports performance.

Spinosaurus is not an aquatic dinosaur.

A predominantly fish-eating diet was envisioned for the sail-backed theropod dinosaur Spinosaurus aegyptiacus when its elongate jaws with subconical teeth were unearthed a century ago in Egypt. Recent discovery of the high-spined tail of that skeleton, however, led to a bolder conjecture that S. aegyptiacus was the first fully aquatic dinosaur. The 'aquatic hypothesis' posits that S. aegyptiacus was a slow quadruped on land but a capable pursuit predator in coastal waters, powered by an expanded tail. We test these functional claims with skeletal and flesh models of S. aegyptiacus. We assembled a CT-based skeletal reconstruction based on the fossils, to which we added internal air and muscle to create a posable flesh model. That model shows that on land S. aegyptiacus was bipedal and in deep water was an unstable, slow-surface swimmer (<1 m/s) too buoyant to dive. Living reptiles with similar spine-supported sails over trunk and tail are used for display rather than aquatic propulsion, and nearly all extant secondary swimmers have reduced limbs and fleshy tail flukes. New fossils also show that Spinosaurus ranged far inland. Two stages are clarified in the evolution of Spinosaurus, which is best understood as a semiaquatic bipedal ambush piscivore that frequented the margins of coastal and inland waterways.

Potential New Therapeutic Approaches for Cisplatin-Resistant Testicular Germ Cell Tumors.

BACKGROUND: Testicular germ cell tumors (TGCTs), a group of heterogeneous neoplasms, are the most frequent tumors of teenagers and young men, with the incidence rising worldwide. High cure rates can be achieved through cisplatin (CDDP)-based treatment, but approximately 10% of patients present refractory disease and virtually no treatment alternatives. Here, we explored new strategies to treat CDDP-resistant. METHODS: In vitro TGCT CDDP-resistance model was established and differential mRNA expression profiles were evaluated using NanoString technology. Then, TGCT cell lines were treated with four potential drugs (PCNA-I1, ML323, T2AA, and MG-132) to overcome CDDP-resistance. RESULTS: We found several differentially expressed genes related to DNA repair and cell cycle regulation on CDDP-resistant cell line (NTERA-2R) compared to parental cell line (NTERA-2P), and the proteasome inhibitor MG-132 demonstrated cytotoxic activity in all cell lines evaluated, even at a nanomolar range. MG-132 also enhanced cell lines' sensitivity to CDDP, increasing apoptosis in both NTERA-2P and NTERA-2R. CONCLUSIONS: MG-132 emerges as a potential new drug to treat CDDP-resistant TGCT. Targeted therapy based on molecular mechanism insights may contribute to overcome acquired chemotherapy CDDP-resistance.

Composition of Nonextractable Polyphenols from Sweet Cherry Pomace Determined by DART-Orbitrap-HRMS and Their In Vitro and In Vivo Potential Antioxidant, Antiaging, and Neuroprotective Activities.

Sweet cherry pomace is an important source of phenolic compounds with beneficial health properties. As after the extraction of phenolic compounds, a phenolic fraction called nonextractable polyphenols (NEPs) remains usually retained in the extraction residue, alkaline and acid hydrolyses and enzymatic-assisted extraction (EAE) were carried out in this work to recover NEPs from the residue of conventional extraction from sweet cherry pomace. In vitro and in vivo evaluation of the antioxidant, antihypertensive, antiaging, and neuroprotective capacities employing Caenorhabditis elegans was achieved for the first time. Extractable phenolic compounds and NEPs were separated and identified by families by high-performance thin-layer chromatography (HPTLC) with UV/Vis detection. A total of 39 phenolic compounds were tentatively identified in all extracts by direct analysis in real-time high-resolution mass spectrometry (DART-Orbitrap-HRMS). EAE extracts presented the highest in vitro and in vivo antioxidant capacity as well as the highest in vivo antiaging and neuroprotective capacities. These results showed that NEPs with interesting biological properties are retained in the extraction residue, being usually underestimated and discarded.

Characterization of 3D Printed Yttria-Stabilized Zirconia Parts for Use in Prostheses.

The main aim of the present paper is to study and analyze surface roughness, shrinkage, porosity, and mechanical strength of dense yttria-stabilized zirconia (YSZ) samples obtained by means of the extrusion printing technique. In the experiments, both print speed and layer height were varied, according to a 22 factorial design. Cuboid samples were defined, and three replicates were obtained for each experiment. After sintering, the shrinkage percentage was calculated in width and in height. Areal surface roughness, Sa, was measured on the lateral walls of the cuboids, and total porosity was determined by means of weight measurement. The compressive strength of the samples was determined. The lowest Sa value of 9.4 µm was obtained with low layer height and high print speed. Shrinkage percentage values ranged between 19% and 28%, and porosity values between 12% and 24%, depending on the printing conditions. Lowest porosity values correspond to low layer height and low print speed. The same conditions allow obtaining the highest average compressive strength value of 176 MPa, although high variability was observed. For this reason, further research will be carried out about mechanical strength of ceramic 3D printed samples. The results of this work will help choose appropriate printing conditions extrusion processes for ceramics.

Accurate calculation of spin-state energy gaps in Fe(III) spin-crossover systems using density functional methods.

Fe(III) complexes are receiving ever-increasing attention as spin crossover (SCO) systems because they are usually air stable, as opposed to Fe(II) complexes, which are prone to oxidation. Here, we present the first systematic study exclusively devoted to assess the accuracy of several exchange-correlation functionals when it comes to predicting the energy gap between the high-spin (S = 5/2) and the low-spin (S = 1/2) states of Fe(III) complexes. Using a dataset of 24 different Fe(III) hexacoordinated complexes, it is demonstrated that the B3LYP* functional is an excellent choice not only for predicting spin-state energy gaps for Fe(III) complexes undergoing spin-transitions but also for discriminating Fe(III) complexes that are either low- or high-spin in the whole range of temperatures. Our benchmark study has led to the identification of a very versatile Fe(III) compound whose SCO properties can be engineered upon changing a single axial ligand. Overall, this work demonstrates that B3LYP* is a reliable functional for screening new spin-crossover systems with tailored properties.

ETV4 plays a role on the primary events during the adenoma-adenocarcinoma progression in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancers worldwide; it is the fourth leading cause of death in the world and the third in Brazil. Mutations in the APC, DCC, KRAS and TP53 genes have been associated with the progression of sporadic CRC, occurring at defined pathological stages of the tumor progression and consequently modulating several genes in the corresponding signaling pathways. Therefore, the identification of gene signatures that occur at each stage during the CRC progression is critical and can present an impact on the diagnosis and prognosis of the patient. In this study, our main goal was to determine these signatures, by evaluating the gene expression of paired colorectal adenoma and adenocarcinoma samples to identify novel genetic markers in association to the adenoma-adenocarcinoma stage transition. METHODS: Ten paired adenoma and adenocarcinoma colorectal samples were subjected to microarray gene expression analysis. In addition, mutations in APC, KRAS and TP53 genes were investigated by DNA sequencing in paired samples of adenoma, adenocarcinoma, normal tissue, and peripheral blood from ten patients. RESULTS: Gene expression analysis revealed a signature of 689 differentially expressed genes (DEG) (fold-change> 2, p< 0.05), between the adenoma and adenocarcinoma paired samples analyzed. Gene pathway analysis using the 689 DEG identified important cancer pathways such as remodeling of the extracellular matrix and epithelial-mesenchymal transition. Among these DEG, the ETV4 stood out as one of the most expressed in the adenocarcinoma samples, further confirmed in the adenocarcinoma set of samples from the TCGA database. Subsequent in vitro siRNA assays against ETV4 resulted in the decrease of cell proliferation, colony formation and cell migration in the HT29 and SW480 colorectal cell lines. DNA sequencing analysis revealed KRAS and TP53 gene pathogenic mutations, exclusively in the adenocarcinomas samples. CONCLUSION: Our study identified a set of genes with high potential to be used as biomarkers in CRC, with a special emphasis on the ETV4 gene, which demonstrated involvement in proliferation and migration.

Immunophenotypic characteristics of juvenile myelomonocytic leukaemia and their relation with the molecular subgroups of the disease.

The diagnosis of juvenile myelomonocytic leukaemia (JMML) is based on clinical, laboratory and molecular features but immunophenotyping [multiparametric flow cytometry (MFC)] has not been used routinely. In the present study, we describe the flow cytometric features at diagnosis with special attention to the distribution of monocytic subsets and the relation between MFC and molecular subgroups. MFC was performed with an eight-colour platform based on Euroflow. We studied 33 JMML cases. CD34+ /CD117+ /CD13+ cells >2% was found in 25 cases, and 51·5% presented an aberrant expression of CD7. A decrease of CD34+ /CD19+ /CD10+ cells was seen in eight cases and in four they were absent. The granulocytic population had a decreased side scatter in 29 cases. Bone marrow monocytic precursors were increased in 28 patients, with a decrease in classical monocytes (median 80·7%) and increase in CD16+ (intermediate and non-classical). A more pronounced increase in myeloid CD34+ cells was seen in patients with Neurofibromatosis type 1 (NF1) and tyrosine-protein phosphatase non-receptor type 11 (PTPN11), with aberrant CD7 expression in four of six and 10/12 patients respectively. Thus, JMML shows an immunophenotypic profile similar to myelodysplastic syndromes, and a different monocyte subset distribution when compared with chronic MML. MFC proved to be an important diagnostic tool that can help in differential diagnosis with other clonal diseases with monocytosis.

Insights in Osteosarcoma by Proton Nuclear Magnetic Resonance Serum Metabonomics.

Pediatric osteosarcoma outcomes have improved over the last decades; however, patients who do not achieve a full resection of the tumor, even after aggressive chemotherapy, have the worst prognosis. At a genetic level, osteosarcoma presents many alterations, but there is scarce information on alterations at metabolomic levels. Therefore, an untargeted nuclear magnetic resonance metabonomic approach was used to reveal blood serum alterations, when samples were taken from 21 patients with osteosarcoma aged from 12-20 (18, 86%) to 43 (3, 14%) years before any anticancer therapy were collected. The results showed that metabolites differed greatly between osteosarcoma and healthy control serum samples, especially in lipids, aromatic amino acids (phenylalanine and tyrosine), and histidine concentrations. Besides, most of the loading plots point to protons of the fatty acyls (-CH3 and -CH2-) from very-low- and low-density lipoproteins and cholesterol, as crucial metabolites for discrimination of the patients with osteosarcoma from the healthy samples. The relevance of blood lipids in osteosarcoma was highlighted when analyzed together with the somatic mutations disclosed in tumor samples from the same cohort of patients, where six genes linked to the cholesterol metabolism were found being altered too. The high consistency of the discrimination between osteosarcoma and healthy control blood serum suggests that nuclear magnetic resonance could be successfully applied for osteosarcoma diagnostic and prognostic purposes, which could ameliorate the clinical efficacy of therapy.

The Brazilian TP53 mutation (R337H) and sarcomas.

Sarcomas represent less than 1% of all solid neoplasms in adults and over 20% in children. Their etiology is unclear, but genetic susceptibility plays an important role in this scenario. Sarcoma is central in Li-Fraumeni Syndrome (LFS), a familial predisposition cancer syndrome. In Brazil, the high prevalence of p.Arg337His mutations in the TP53 gene brings about a unique condition: a cluster of LFS. In the present work, we studied 502 sarcoma patients not selected by age or family history in an attempt to assess the impact of the so-called "Brazilian germline TP53 mutation" (p.Arg337His) on this tumor type. We found that 8% of patients are carriers, with leiomyosarcoma being the main histologic type of sarcoma, corresponding to 52.5% of the patients with the mutated TP53 gene. These findings emphasize the importance of genetic counseling and can better guide the management of sarcoma patients.

Ontogenetic similarities between giraffe and sauropod neck osteological mobility.

The functional morphology of sauropod dinosaur long necks has been studied extensively, with virtual approaches yielding results that are difficult to obtain with actual fossils, due to their extreme fragility and size. However, analyses on virtual fossils have been questioned on several of their premises, such as the ability to accurately reconstruct intervertebral tissue with only skeletal data; or whether zygapophyseal overlap can be used to determine the limits of range of motion, since some extreme neck poses in extant giraffes have been claimed not to retain any zygapophyseal overlap. We compared articulation and range of motion in extant giraffes with the exceptionally well-preserved and complete basally branching eusauropod Spinophorosaurus nigerensis from the Middle (?) Jurassic of Niger, under the same virtual paleontology protocols. We examined the articulation and range of motion on grown and young specimens of both Spinophorosaurus and giraffes in order to record any potential changes during ontogeny. Also, the postures of virtual giraffes were compared with previously published data from living animals in the wild. Our analyses show that: (i) articulation of virtual bones in osteologically neutral pose (ONP) does enable accurate prediction of the amount of inter-vertebral space in giraffes and, roughly, in Spinophorosaurus; (ii) even the most extreme neck postures attained by living giraffes in the wild do not require to disarticulate cervical vertebrae; (iii) both living giraffes and Spinophorosaurus have large intervertebral spaces between their cervical centra in early ontogenetical stages, which decrease as ontogeny advances; and (iv) that grown specimens have a greater osteological range of motion in living giraffes and Spinophorosaurus.