Carcinoma de tiroides una verdadera pandemia?

El diagnóstico de cáncer de tiroides se ha incrementado y las posibilidades de detección de una enfermedad subclínica son altas, toda vez que disponemos de herramientas de detección más sensibles y de fácil acceso. Por ende, el clínico requiere conocer la historia natural del nódulo tiroideo y del carcinoma papilar de tiroides de bajo riesgo para brindar a su paciente el mejor tratamiento basado en la evidencia clínica. El objetivo de esta revision es reconocer los elementos clínicos que han condicionado el aumento inusitado de casos de cáncer de tiroides. Conclusión: El sobrediagnóstico del cáncer de tiroides es una realidad, que se posibilita por el uso extendido de biopsia por aspiración con aguja fina ((BACAF)) después de la detección de un nódulo tiroideo, en gran parte de manera incidental, sin acarrear la mayoría de las veces un mejor pronóstico después de su tratamiento.

Improved display of synthetic IgG-binding domains on the baculovirus surface

Improved display of foreign protein moieties in combination with beneficial alteration of the viral surface properties should be of value for targeted and enhanced gene delivery. Here, we describe a vector based on Autographa californica multiple nucleopolyhedrovirus (AcMNPV) displaying synthetic IgG-bincling domains (ZZ) of protein A fused to the transmembrane anchor of vesicular stomatitis virus (VSV) G protein. This display vector was equipped with a GFP/EGFP expression cassette enabling fluorescent detection in both insect and mammalian cells. The virus construct displayed the biologically active fusion protein efficiently and showed increased binding capacity to IgG. As the display is carried out using a membrane anchor of foreign origin, gp64 is left intact for virus entry, which may increase gene expression in the transduced mammalian cells. In addition, the viral vector can be targeted to any desired cell type via binding of ZZ domains when an appropriate IgG antibody is available.

Protein kinase C down-regulation, and not transient activation, correlates with melanocyte growth

The nontumorigenic, immortal line of murine melanocytes, Mel-ab, requires the continual presence of biologically active phorbol esters for growth (R. E. Wilson et al., Cancer Res., 49: 711–716, 1989). Comparable treatments of B16 murine melanoma cells result in partial inhibition of cell proliferation. The role of protein kinase C (PKC) in the modulation of growth of cells from these two melanocytic cell lines has been investigated. Significant levels of PKC were present in quiescent Mel-ab cells as determined by Western blotting, whereas no immunoreactive protein was detected in cell extracts from either proliferating Mel-ab or B16.F1 cells. Phosphorylation of a Mr 80,000 protein, which by one- and two-dimensional gel analysis comigrated with the known Mr 80,000 protein substrate of PKC in fibroblasts, was induced in 12-O-tetradecanoylphorbol-13-acetate-stimulated quiescent Mel-ab cells but not in proliferating Mel-ab cells or B16.F1 melanoma cells. Direct measurement of PKC activity in these cells demonstrated a 10-fold greater level of activity in quiescent Mel-ab cells (262 ± 50 pmol/min/mg SD) compared with growing cells (22.8 ± 11.8 pmol/min/mg SD). An intermediate level of activity was detected in proliferating B16.F1 melanoma cells (148.5 ± 20.4 pmol/min/mg SD). The subcellular distribution of PKC was dependent upon the growth state of the cells such that quiescent Mel-ab cells displayed a higher level of activity in the cytosol, whereas growing Melab cells displayed greater activity in the particulate fraction. Like many other transformed lines, B16.F1 melanoma cells constitutively expressed the majority of enzyme activity in the particulate fraction. Measurement of [3H]phorbol ester binding in intact cells paralleled the PKC activation data such that quiescent Mel-ab cells displayed binding of 1612 ± 147 cpm/106 cells, whereas proliferating Mel-ab and B16.F1 melanoma cells displayed binding of 652 ± 28 and 947 ± 81 cpm/106 cells, respectively. Membrane-permeant diacylglycerol analogues, which activated but did not down-regulate PKC, were devoid of growth-stimulating effects on melanocytes, even in the presence of the specific diacylglycerol kinase inhibitor, R59022. Together, these data show that PKC down-regulation, and not activation, correlates with the growth of melanocytes in culture.

The effect of cruciferous and leguminous sprouts on genotoxicity, in vitro and in vivo

Vegetable consumption is associated with a reduced risk of colorectal cancer, which is the second most common cancer after lung/breast cancer within Europe. Some putative protective phytochemicals are found in higher amounts in young sprouts than in mature plants. The effect of an extract of mixed cruciferous and legume sprouts on DNA damage induced by H(2)O(2) was measured in HT29 cells using single cell microgelelectrophoresis (comet). Significant antigenotoxic effect (P < or = 0.05) was observed when HT29 cells were pre-incubated with the extract (100 and 200 microL/mL) for 24 hours and then challenged with H(2)O(2). A parallel design intervention study was carried out on 10 male and 10 female healthy adult volunteers (mean age = 25.5 years) fed 113 g of cruciferous and legume sprouts daily for 14 days. The effect of the supplementation was measured on a range of parameters, including DNA damage in lymphocytes (comet), the activity of various detoxifying enzymes (glutathione S-transferase, glutathione peroxidase, and superoxide dismutase), antioxidant status using the ferric reducing ability of plasma assay, plasma antioxidants (uric acid, ascorbic acid, and alpha-tocopherol), blood lipids, plasma levels of lutein, and lycopene. A significant antigenotoxic effect against H(2)O(2)-induced DNA damage was shown in peripheral blood lymphocytes of volunteers who consumed the supplemented diet when compared with the control diet (P = 0.04). No significant induction of detoxifying enzymes was observed during the study, neither were plasma antioxidant levels or activity altered. The results support the theory that consumption of cruciferous vegetables is linked to a reduced risk of cancer via decreased damage to DNA.

A central role for heme iron in colon carcinogenesis associated with red meat intake

Epidemiology shows that red and processed meat intake is associated with an increased risk of colorectal cancer. Heme iron, heterocyclic amines and endogenous N-nitroso compounds (NOC) are proposed to explain this effect, but their relative contribution is unknown. Our study aimed at determining, at nutritional doses, which is the main factor involved and proposing a mechanism of cancer promotion by red meat. The relative part of heme iron (1% in diet), heterocyclic amines (PhIP+MeIQx, 50+25 μg/kg in diet) and NOC (induced by NaNO2+NaNO3 0.17+0.23 g/l of drinking water) was determined by a factorial design and preneoplastic endpoints in chemically-induced rats and validated on tumors in Min mice. The molecular mechanisms (genotoxicity, cytotoxicity) were analyzed in vitro in normal and Apc- deficient cell lines and confirmed on colon mucosa. Heme iron increased the number of preneoplastic lesions but dietary heterocyclic amines and NOC had no effect on carcinogenesis in rats. Dietary hemoglobin increased tumor load in Min mice (control diet: 67±39 mm2; 2,5% hemoglobin diet: 114±47 mm2, p=0.004). In vitro, fecal water from rats given hemoglobin was rich in aldehydes and was cytotoxic to normal cells, but not to premalignant cells. The aldehydes 4-hydroxynonenal and 4-hydroxyhexenal were more toxic to normal versus mutated cells and were only genotoxic to normal cells. Genotoxicity was also observed in colon mucosa of mice given hemoglobin. These results highlight the role of heme iron in the promotion of colon cancer by red meat and suggest that heme iron could initiate carcinogenesis through lipid peroxidation.

Neural stem cells adopt tumorigenic properties by constitutively activated NF-kappaB and subsequent VEGF up-regulation

One of the challenges in stem cell research is to avoid transformation during cultivation. We studied high passage subventricular zone derived neural stem cells (NSCs) cultures of adult rats in the absence of growth factors epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). We termed this culture exogenous growth factor independent neural stem cells (GiNSCs). GiNSCs expressed stemness markers, displayed a high constitutive NF-kappaB activity and an increased, aberrant, polyploid DNA content. GiNSCs showed a tumorigenic phenotype and formed colonies in a soft agar assay. Microarray analysis showed the up-regulation of the NF-kappaB target gene vascular endothelial growth factor (VEGF). In contrast, proneuronal genes were down-regulated. Under neuronal differentiation conditions GiNSCs adopted a glioma-like phenotype, with nuclear p53, preserving high amounts of Nestin positive cells and prolonged proliferation. Neutralization of VEGF strongly inhibited proliferation and induced differentiation. In a gain of function approach, the transfection of NSCs with constitutively active upstream kinase IKK-2 led to constitutively activated NF-kappaB, proliferation in absence of growth factors and augmented VEGF secretion. In a rescue experiment a reduction of NF-kappaB activity by overexpression of IkappaB-AA1 was able to shift the morphology toward an elongated cell form, increased cell death, and decreased proliferation. Thus GiNSCs may provide a potent tool in cancer research, as their exogenous cytokine independent proliferation and their constitutively high NF-kappaB expression presumes cancerous properties observed in gliomas. In addition, this study might add a novel mechanism for detecting oncogenic transformation in therapeutic stem cell cultures.

Pooled analysis of recent studies on magnetic fields and childhood leukaemia

BACKGROUND: Previous pooled analyses have reported an association between magnetic fields and childhood leukaemia. We present a pooled analysis based on primary data from studies on residential magnetic fields and childhood leukaemia published after 2000. METHODS: Seven studies with a total of 10 865 cases and 12 853 controls were included. The main analysis focused on 24-h magnetic field measurements or calculated fields in residences. RESULTS: In the combined results, risk increased with increase in exposure, but the estimates were imprecise. The odds ratios for exposure categories of 0.1-0.2 mu T, 0.2-0.3 mu T and >= 0.3 mu T, compared with <0.1 mu T, were 1.07 (95% Cl 0.81-1.41), 1.16 (0.69-1.93) and 1.44 (0.88-2.36), respectively. Without the most influential study from Brazil, the odds ratios increased somewhat. An increasing trend was also suggested by a nonparametric analysis conducted using a generalised additive model. CONCLUSIONS: Our results are in line with previous pooled analyses showing an association between magnetic fields and childhood leukaemia. Overall, the association is weaker in the most recently conducted studies, but these studies are small and lack methodological improvements needed to resolve the apparent association. We conclude that recent studies on magnetic fields and childhood leukaemia do not alter the previous assessment that magnetic fields are possibly carcinogenic. British Journal of Cancer (2010) 103, 1128-1135. doi: 10.1038/sj.bjc.6605838 www.bjcancer.com (c) 2010 Cancer Research UK

Alternative splicing and genetic diversity: silencers are more frequently modified by SNVs associated with alternative exon/intron borders

With the availability of a large amount of genomic data it is expected that the influence of single nucleotide variations (SNVs) in many biological phenomena will be elucidated. Here, we approached the problem of how SNVs affect alternative splicing. First, we observed that SNVs and exonic splicing regulators (ESRs) independently show a biased distribution in alternative exons. More importantly, SNVs map more frequently in ESRs located in alternative exons than in ESRs located in constitutive exons. By looking at SNVs associated with alternative exon/intron borders (by their common presence in the same cDNA molecule), we observed that a specific type of ESR, the exonic splicing silencers (ESSs), are more frequently modified by SNVs. Our results establish a clear association between genetic diversity and alternative splicing involving ESSs.

Evaluation of Quantitative RT-PCR Using Nonamplified and Amplified RNA

Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) is a standard assay in molecular medicine for gene expression analysis. Samples from incisional/needle biopsies, laser-microdissected tumor cells and other biologic sources, normally available in clinical cancer studies, generate very small amounts of RNA that are restrictive for expression analysis. As a consequence, an RNA amplification procedure is required to assess the gene expression levels of such sample types. The reproducibility and accuracy of relative gene expression data produced by sensitive methodology as qRT-PCR when cDNA converted from amplified (A) RNA is used as template has not yet been properly addressed. In this study, to properly evaluate this issue, we performed 1 round of linear RNA amplification in 2 breast cell lines (C5.2 and HB4a) and assessed the relative expression of 34 genes using cDNA converted from both nonamplified (NA) and A RNA. Relative gene expression was obtained from beta actin or glyceraldehyde 3-phosphate dehydrogenase normalized data using different dilutions of cDNA, wherein the variability and fold-change differences in the expression of the 2 methods were compared. Our data showed that 1 round of linear RNA amplification, even with suboptimal-quality RNA, is appropriate to generate reproducible and high-fidelity qRT-PCR relative expression data that have similar confidence levels as those from NA samples. The use of cDNA that is converted from both A and NA RNA in a single qRT-PCR experiment clearly creates bias in relative gene expression data.

Defective transcription/repair factor IN recruitment to specific UV lesions in trichothiodystrophy syndrome

Most trichothiodystrophy (TTD) patients present mutations in the xeroderma pigmentosum D (XPD) gene, coding for a subunit of the transcription/repair factor IIH (TFHH) complex involved in nucleotide excision repair (NER) and transcription. After UV irradiation, most TTD/XPD patients are more severely affected in the NER of cyclobutane pyrimidine dimers (CPD) than of 6-4-photoproducts (6-4PP). The reasons for this differential DNA repair defect are unknown. Here we report the first study of NER in response to CPDs or 6-4PPs separately analyzed in primary fibroblasts. This was done by using heterologous photorepair; recombinant adenovirus vectors carrying photolyases enzymes that repair CPD or 64PP specifically by using the energy of light were introduced in different cell lines. The data presented here reveal that some mutations affect the recruitment of TFHH specifically to CPDs, but not to 6-4PPs. This deficiency is further confirmed by the inability of TTD/XPD cells to recruit, specifically for CPDs, NER factors that arrive in a TFIIH-dependent manner later in the NER pathway. For 6-4PPs, we show that TFHH complexes carrying an NH2-terminal XPD mutated protein are also deficient in recruitment of NER proteins downstream of TFUH. Treatment with the histone deacetylase inhibitor trichostatin A allows the recovery of TFHH recruitment to CPDs in the studied TTD cells and, for COOH-terminal XPD mutations, increases the repair synthesis and survival after UV, suggesting that this defect can be partially related with accessibility of DNA damage in closed chromatin regions.

p53 Mutant Human Glioma Cells Are Sensitive to UV-C-Induced Apoptosis Due to Impaired Cyclobutane Pyrimidine Dimer Removal

The p53 protein is a key regulator of cell responses to DNA damage, and it has been shown that It sensitizes glioma cells to the alkylating agent temozolomide by up-regulating the extrinsic apoptotic pathway, whereas it increases the resistance to chloroethylating agents, such as ACNU and BCNU, probably by enhancing the efficiency of DNA repair. However, because these agents induce a wide variety of distinct DNA lesions, the direct Importance of DNA repair is hard to access. Here, it is shown that the Induction of photoproducts by UV light (UV-C) significantly Induces apoptosis In a p53-mutated glioma background. This Is caused by a reduced level of photoproduct repair, resulting In the persistence of DNA lesions in p53-mutated glioma cells. UV-C-Induced apoptosis in p53 mutant glioma cells Is preceded by strong transcription and replication inhibition due to blockage by unrepaired photolesions. Moreover, the results Indicate that UV-C-induced apoptosis of p53 mutant glioma cells Is executed through the intrinsic apoptotic pathway, with Bcl-2 degradation and sustained Bax and Bak up-regulation. Collectively, the data Indicate that unrepaired DNA lesions Induce apoptosis In p53 mutant gliomas despite the resistance of these gliomas to temozolomide, suggesting that efficiency of treatment of p53 mutant gliomas might be higher with agents that Induce the formation of DNA lesions whose global genomic repair is dependent on p53. (Mol Cancer Res 2009;7(2):237-46)

Objective Detection and Delineation of Oral Neoplasia Using Autofluorescence Imaging

Although the oral cavity is easily accessible to inspection, patients with oral cancer most often present at a late stage, leading to high morbidity and mortality. Autofluorescence imaging has emerged as a promising technology to aid clinicians in screening for oral neoplasia and as an aid to resection, but current approaches rely on subjective interpretation. We present a new method to objectively delineate neoplastic oral mucosa using autofluorescence imaging. Autofluorescence images were obtained from 56 patients with oral lesions and 11 normal volunteers. From these images, 276 measurements from 159 unique regions of interest (ROI) sites corresponding to normal and confirmed neoplastic areas were identified. Data from ROIs in the first 46 subjects were used to develop a simple classification algorithm based on the ratio of red-to-green fluorescence; performance of this algorithm was then validated using data from the ROIs in the last 21 subjects. This algorithm was applied to patient images to create visual disease probability maps across the field of view. Histologic sections of resected tissue were used to validate the disease probability maps. The best discrimination between neoplastic and nonneoplastic areas was obtained at 405 nm excitation; normal tissue could be discriminated from dysplasia and invasive cancer with a 95.9% sensitivity and 96.2% specificity in the training set, and with a 100% sensitivity and 91.4% specificity in the validation set. Disease probability maps qualitatively agreed with both clinical impression and histology. Autofluorescence imaging coupled with objective image analysis provided a sensitive and noninvasive tool for the detection of oral neoplasia.

Analyzing Ru(III)-dmso and Ru(III)-dms motifs in compounds used in the synthesis of the antimetastatic agents

The chemistry of Ru(III) complexes containing dmso as a ligand has become an interesting area in the cancer treatment field. Because of this, structural knowledge and chemistry of the moiety Ru(III)-dmso have become important to cancer research. The crystal structures of the compounds mer-[RuCl(3)(dms)(3)] (1) and mer-[RuCl(3)(dms)(2)(dmso)]:mer-[RuCl(3)(dms)(3)] (2) were determined by X-ray crystallography and a speciation of the presence of intramolecular hydrogen bond in these structures has been studied. Compound (1) crystallizes in the orthorhombic space group, Pna2(1); a = 16.591(8) angstrom, b = 8.724(2) angstrom. c = 10.547(3) angstrom; Z = 12 and (2) crystallizes in the space group, P2(1)/C: a = 11.9930(2) angstrom, b = 7.9390(2) angstrom, c = 15.8700(3) angstrom, beta = 93.266(1)degrees, Z = 2. From the X-ray structures solved in this work, were possible to suggest an interpretation for the broad lines observed in the EPR spectra of the Ru(III) compounds explored here. Also, the exchange interactions detected by EPR spectroscopy in solid state and in solution, confirm the presence of van der Waals interactions such as C-H center dot center dot center dot Cl in the compounds (1), (2) and (3). The use of techniques such as IR, UV-vis, (1)H NMR and EPR Spectroscopy and Cyclic Voltammetry were applied in this work to analyze the behavior of these metallocompounds. (c) 2008 Elsevier B.V. All rights reserved.

Statistical coupling analysis of aspartic proteinases based on crystal structures of the Trichoderma reesei enzyme and its complex with pepstatin A

The crystal structures of an aspartic proteinase from Trichoderma reesei (TrAsP) and of its complex with a competitive inhibitor, pepstatin A, were solved and refined to crystallographic R-factors of 17.9% (R(free)=21.2%) at 1.70 angstrom resolution and 15.81% (R(free) = 19.2%) at 1.85 angstrom resolution, respectively. The three-dimensional structure of TrAsP is similar to structures of other members of the pepsin-like family of aspartic proteinases. Each molecule is folded in a predominantly beta-sheet bilobal structure with the N-terminal and C-terminal domains of about the same size. Structural comparison of the native structure and the TrAsP-pepstatin complex reveals that the enzyme undergoes an induced-fit, rigid-body movement upon inhibitor binding, with the N-terminal and C-terminal lobes tightly enclosing the inhibitor. Upon recognition and binding of pepstatin A, amino acid residues of the enzyme active site form a number of short hydrogen bonds to the inhibitor that may play an important role in the mechanism of catalysis and inhibition. The structures of TrAsP were used as a template for performing statistical coupling analysis of the aspartic protease family. This approach permitted, for the first time, the identification of a network of structurally linked residues putatively mediating conformational changes relevant to the function of this family of enzymes. Statistical coupling analysis reveals coevolved continuous clusters of amino acid residues that extend from the active site into the hydrophobic cores of each of the two domains and include amino acid residues from the flap regions, highlighting the importance of these parts of the protein for its enzymatic activity. (C) 2008 Elsevier Ltd. All rights reserved.