Eva Fernández

Expression of cathepsins B and S in the progression of prostate carcinoma

Cathepsins B and S (CatB, CatS) are lysosomal cysteine proteases which, among other functions, appear to play a role in cancer progression in different tumor models due to their matrix‐degrading properties. To investigate their possible involvement in the development of prostate carcinoma, we immunohistochemically analyzed CatB and CatS in 38 primary human prostatic adenocarcinomas, as well as concomitant high‐grade prostatic intra‐epithelial neoplasia, nodular hyperplasia and normal tissue. CatB expression was observed in 28 (74%) and CatS in 32 (84%) carcinomas, being concomitant in 24 cases (63%). High‐grade intra‐epithelial neoplasia expressed CatB in 20/23 cases (87%), and a similar result was obtained for CatS, with expression of both coinciding in 18 cases (78%). In non‐neoplastic tissue, strong expression of both proteases was observed in macrophages, inflamed glands and transitional metaplasia, whereas atrophic glands and basal cells of normal glands displayed intense CatB positivity. We conclude that CatB and CatS are often expressed together in neoplastic prostatic cells from pre‐invasive to invasive and clinically detectable stages, suggesting a putative role in local invasion, though other functions cannot be ruled out.

Ancient DNA from an Early Neolithic Iberian population supports a pioneer colonization by first farmers

The Neolithic transition has been widely debated particularly regarding the extent to which this revolution implied a demographic expansion from the Near East. We attempted to shed some light on this process in northeastern Iberia by combining ancient DNA (aDNA) data from Early Neolithic settlers and published DNA data from Middle Neolithic and modern samples from the same region. We successfully extracted and amplified mitochondrial DNA from 13 human specimens, found at three archaeological sites dated back to the Cardial culture in the Early Neolithic (Can Sadurní and Chaves) and to the Late Early Neolithic (Sant Pau del Camp). We found that haplogroups with a low frequency in modern populations—N* and X1—are found at higher frequencies in our Early Neolithic population (∼31%). Genetic differentiation between Early and Middle Neolithic populations was significant (FST∼0.13, P < 10−5), suggesting that genetic drift played an important role at this time. To improve our understanding of the Neolithic demographic processes, we used a Bayesian coalescence‐based simulation approach to identify the most likely of three demographic scenarios that might explain the genetic data. The three scenarios were chosen to reflect archaeological knowledge and previous genetic studies using similar inferential approaches. We found that models that ignore population structure, as previously used in aDNA studies, are unlikely to explain the data. Our results are compatible with a pioneer colonization of northeastern Iberia at the Early Neolithic characterized by the arrival of small genetically distinctive groups, showing cultural and genetic connections with the Near East.

Nonhepatosplenic γδ T-cell lymphomas represent a spectrum of aggressive cytotoxic T-cell lymphomas with a mainly extranodal presentation.

&ggr;&dgr; T cells represent a minor T-cell subset that is mainly distributed in mucosal surfaces. Two distinct lymphomas derived from these cells have been recognized: hepatosplenic &ggr;&dgr; T-cell lymphoma (HSTL) and primary cutaneous &ggr;&dgr; T-cell lymphoma (PCGD-TCL). However, whether other anatomic sites may also be involved and whether they represent a spectrum of the same disease are not well studied. The lack of T-cell receptor (TCR)&bgr; expression has been used to infer a &ggr;&dgr; origin when other methods are not available. We studied 35 T-cell tumors suspected to be &ggr;&dgr; TCL using monoclonal antibodies reactive with TCR &dgr; or &ggr; in paraffin sections. We were able to confirm &ggr;&dgr; chain expression in 22 of 35 cases. We identified 8 PCGD-TCLs, 6 HSTLs, and 8 &ggr;&dgr; TCLs without hepatosplenic or cutaneous involvement involving mainly extranodal sites. Two such cases were classified as enteropathy-associated T-cell lymphoma, type II. The other &ggr;&dgr; TCL presented in the intestine, lung, tongue, orbit, and lymph node. In addition, we observed 13 cases with mainly extranodal involvement that lacked any TCR expression (“TCR silent”). In all cases, a natural killer cell origin was excluded. In conclusion, the lack of TCR&bgr; expression does not always predict &ggr;&dgr;-T-cell derivation, as TCR silent cases may be found. The recognition of &ggr;&dgr; TCL presenting in extranodal sites other than skin and liver/spleen expands the clinical spectrum of these tumors. However, non-HSTL &ggr;&dgr; TCL do not seem to represent a single entity. The relationship of these tumors with either HSTL or PCGD-TCL requires further study.

Brief Communication: Ancient Nuclear DNA and Kinship Analysis: The Case of a Medieval Burial in San Esteban Church in Cuellar (Segovia, Central Spain)

The aim of this work was to investigate a very common situation in the archaeological and anthropological context: the study of a burial site containing several individuals, probably related genetically, using ancient DNA techniques. We used available ancient DNA and forensic protocols to obtain reliable results on archaeological material. The results also enabled molecular sex determination to be compared with osteological data. Specifically, a modified ancient DNA extraction method combined with the amplification of nuclear markers with the AmpFlSTR®MiniFiler™ kit(Applied Biosystems) was used. Seven medieval individuals buried in four niches dated in the 15th Century at San Esteban Church in Cuellar (Segovia, Central Spain) were analyzed by the proposed method, and four of seven provided complete autosomal short tandem repeat (STRs) profiles. Kinship analyses comprising paternity and sibship relations were carried out with pedigree-specific software used in forensic casework. A 99.98% paternity probability was established between two individuals, although lower percentages (68%) were obtained in other cases, and some hypothetical kinship relations were excluded. The overall results could eventually provide evidence for reconstructing the historical record.

Tissue macroarrays (mIcrochops) for gene expression analysis

Abstract. We describe a simple system of tissue arraying with multiple tissue fragments obtained with a biopsy punch from selected areas of paraffin blocks. The new blocks thus constructed allow multiple tissue sections in which the uniform shape of the fragments coupled with a geometrical display and a significant amount of tissue per case allows a dependable, cost-effective way to screen tumors or other kinds of tissues with techniques such as immunohistochemistry. This system avoids the disadvantages of previous laborious methods of tissue arraying, such as expensive equipment and scarce tissue sampling, and it can be implemented in any institution with minimal cost and elaboration.

A Paleoneurohistological Study of 3,000-Year-Old Mummified Brain Tissue from the Mediterranean Bronze Age

Objectives: Mummified nervous tissue is very rarely found in ancient remains and usually corresponds to corpses which were frozen or preserved in bogs, conditions which limit tissue autolysis and bacterial degradation. Here, we show the unusual finding of spontaneously mummified brain tissue from several individuals from the little known megalithic talaiotic culture of the island of Minorca, dating approximately 3,000 years before present and corresponding to the late Mediterranean Bronze Age. Methods: These individuals were part of an intact burial site containing 66 subjects. Intracraneal samples were carefully rehydrated with Sandison’s solution. We used classical histochemical as well as 2D and 3D (scanning) electron-microscopic techniques. Results: We provide evidence of the nervous nature of the samples as well as a detailed description of the morphological features of these ancient tissues. The intracranial material consisted of well-preserved eosinophilic reticular tissue and, although mostly absent, some exceptional pigment-containing neurons were identified. Conclusions: We present a detailed morphological analysis which can provide valuable information and guidelines for the interpretation of this scarce type of mummified samples and provide explanations for this surprising preservation.

Analysis of the IGHV region in Burkitt's lymphomas supports a germinal center origin and a role for superantigens in lymphomagenesis

The analysis of immunoglobulin heavy chain variable (IGHV) region may disclose the influence of antigens in Burkitts lymphomas (BL). IGHV sequences from 38 patients and 35 cell lines were analyzed. IGHV3 subset genes were the most used and IGHV4-34 gene was overrepresented. IGHV genes were mutated in 98.6% of the cases, 36% acquired potential glycosylation sites, and in 52% somatic-hypermutation-process was ongoing. Binding motifs for superantigens like Staphylococcal protein A and carbohydrate I/i were preserved in 89% of the cases. IGHV analysis of BL cells supports a germinal center origin and points toward a role for superantigens in lymphomagenesis.

Differential Gene Expression Profile Associated to Apoptosis Induced by Dexamethasone in CLL Cells According to IGHV/ZAP-70 Status

Purpose: Glucocorticoids are part of the therapeutic armamentarium of chronic lymphocytic leukemia (CLL) where it has been suggested that cells with unmutated IGHV genes exhibit higher sensitivity. The mechanisms by which glucorticoids are active in CLL are not well elucidated. We aimed to ascertain the activity of dexamethasone in CLL cells according to prognosis and to identify the molecular mechanisms that are influencing the response to this drug. Experimental Design: Sensitivity to dexamethasone was analyzed ex vivo in 50 CLL and compared according to IGHV mutational status and/or ZAP-70 expression. The response was further compared by gene expression profiling (GEP) of selected cases. Expression of genes of interest was validated by quantitative reverse transcriptase PCR. Results: Response to dexamethasone was higher in cases with unmutated IGHV/high ZAP-70 expression, and the levels of induction of the pro-apoptotic Bim protein correlated with the degree of cell death. GEP analysis showed few genes differentially expressed after dexamethasone treatment between mutated and unmutated cases. However, functional annotation analysis showed that unmutated cases had significant enrichment in terms related to apoptosis. Specific analysis of genes of interest conducted in a large series disclosed that in unmutated IGHV cells, FKBP5 expression was higher at baseline and after dexamethasone exposure and that GILZ was more induced by dexamethasone treatment in these cases. Conclusions: Unmutated IGHV/high ZAP-70 CLL cells exhibit better response to dexamethasone treatment, which is accompanied by a differential expression of genes involved in the glucocorticoid receptor pathway and by an increased induction of genes related to apoptosis. Clin Cancer Res; 18(21); 5924–33. ©2012 AACR.

HACIA EL ORIGEN DE LOS VASCOS SECUENCIAS DE DNA MITOCONDRIAL ANTIGUO DEL PAÍS VASCO

Toward the origin of the basques. Ancient mitochondrial DNA sequences of the Basque country. ABSTRACT.- The task of reconstructing the human past has generated more knowledge of the objects than of their manufacturers. Many of the technological advances, social and structural changes of the pre- and proto- history periods have been explained by demographic expansions and migratory movements. In recent years, Mo- lecular Archaeology has been contributing to the study of Prehistory by adding new information about the biolo- gical characteristics of ancient human populations. The direct genetic study of our ancestors with sophisticated molecular analysis techniques brings us closer to each particular individual. In time, it will be possible to draw up genetic maps of past populations and, thus, contrast the theories of regional and general population period by period. Among the different types of molecular analysis, those of ancient DNA seem to be the most interesting in spite of the technical limitations and the degradation of the DNA, since they deal directly with ancient human remains. This is the first publication of mitochondrial DNA sequences obtained from teeth of the Neolithic epoch in the Cave of Atxuri, and of the medieval period in San Juan de Momoitio, Garai (Biscay), which, together with those of the necropolis of the Alto de la Ermita of Amezaga (Alava) are, so far, the only ones published from the Iberian Peninsula. The results show clear mutational differences with respect to the more common sequence of current Europeans. In none of the 12 samples does the analysis detect haplogroup V, which is much debated in the current scientific literature about the origin of the Basques. The contribution of these sequences is a qualita- tive leap in the study of prehistory. Due to the small number of individuals available to date, general conclusions cannot be drawn.

Calcium, Calmodulin and Phosphoinositides in Leaflet Movements Mediated by Phytochrome

Calcium transduction pathways are involved in plant signaling, including light and hormonal responses (Sanders et al 1999). Calmodulin and phosphoinositide metabolism may also be other components of the transduction pathway (Yang 1996). Cytoplasmic Ca2+ has been related with some phytochrome-mediated responses such as protoplast swelling, germination of fern spores and seed germination (Tretyn et al 1991). Microinjection experiments reported by Neuhaus et al (1993) strongly support this involvement. Thus, a pre-eminent role of Ca2+ in phytochrome transduction pathways has been proposed (Roux 1994). Protein phosphorylation and calcium-binding proteins such as calmodulin, could be downstream in the phytochrome transduction pathway (Schafer et al 1997).