Samba, a Xenopus hnRNP Expressed in Neural and Neural Crest Tissues

RNA binding proteins regulate gene expression at the posttranscriptional level and play important roles in embryonic development. Here, we report the cloning and expression of Samba, a Xenopus hnRNP that is maternally expressed and persists at least until tail bud stages. During gastrula stages, Samba is enriched in the dorsal regions. Subsequently, its expression is elevated only in neural and neural crest tissues. In the latter, Samba expression overlaps with that of Slug in migratory neural crest cells. Thereafter, Samba is maintained in the neural crest derivatives, as well as other neural tissues, including the anterior and posterior neural tube and the eyes. Overexpression of Samba in the animal pole leads to defects in neural crest migration and cranial cartilage development. Thus, Samba encodes a Xenopus hnRNP that is expressed early in neural and neural crest derivatives and may regulate crest cells migratory behavior. Developmental Dynamics 238:204-209, 2009. (C) 2008 Wiley-Liss, Inc.

Insights into the organization of dorsal spinal cord pathways from an evolutionarily conserved raldh2 intronic enhancer

Comparative studies of the tetrapod raldh2 (aldh1a2) gene, which encodes a retinoic acid (RA) synthesis enzyme, have led to the identification of a dorsal spinal cord enhancer. Enhancer activity is directed dorsally to the roof plate and dorsal-most (dl1) interneurons through predicted Tcf- and Cdx-homeodomain binding sites and is repressed ventrally via predicted Tgif homeobox and ventral Lim-homeodomain binding sites. Raldh2 and Math1/Cath1 expression in mouse and chicken highlights a novel, transient, endogenous Raldh2 expression domain in dl1 interneurons, which give rise to ascending circuits and intraspinal commissural interneurons, suggesting roles for RA in the ontogeny of spinocerebellar and intraspinal proprioceptive circuits. Consistent with expression of raldh2 in the dorsal interneurons of tetrapods, we also found that raldh2 is expressed in dorsal interneurons throughout the agnathan spinal cord, suggesting ancestral roles for RA signaling in the ontogenesis of intraspinal proprioception.

Spectrum and Prevalence of FP/TMEM127 Gene Mutations in Pheochromocytomas and Paragangliomas

Context Pheochromocytomas and paragangliomas are genetically heterogeneous neural crest-derived neoplasms. We recently identified germline mutations of the novel transmembrane-encoding gene FP/TMEM127 in familial and sporadic pheochromocytomas consistent with a tumor suppressor effect. Objectives To examine the prevalence and spectrum of FP/TMEM127 mutations in pheochromocytomas and paragangliomas and to test the effect of mutations in vitro. Design, Setting, and Participants We sequenced the FP/TMEM127 gene in 990 individuals with pheochromocytomas and/or paragangliomas, including 898 previously unreported cases without mutations in other susceptibility genes from 8 independent worldwide referral centers between January 2009 and June 2010. A multiplex polymerase chain reaction-based method was developed to screen for large gene deletions in 545 of these samples. Confocal microscopy of 5 transfected mutant proteins was used to determine their subcellular localization. Main Outcome Measures The frequency and type of FP/TMEM127 mutation or deletion was assessed and correlated with clinical variables; the subcellular localization of 5 overexpressed mutants was compared with wild-type FP/TMEM127 protein. Results We identified 19 potentially pathogenic FP/TMEM127 germline mutations in 20 independent families, but no large deletions were detected. All mutation carriers had adrenal tumors, including 7 bilateral (P=2.7 x 10(-4)) and/or with familial disease (5 of 20 samples; P=.005). The median age at disease onset in the FP/TMEM127 mutation group was similar to that of patients without a mutation (41.5 vs 45 years, respectively; P=.54). The most common presentation was that of a single benign adrenal tumor in patients older than 40 years. Malignancy was seen in 1 mutation carrier (5%). Expression of 5 novel FP/TMEM127 mutations in cell lines revealed diffuse localization of the mutant proteins in contrast with the discrete multiorganelle distribution of wild-type TMEM127. Conclusions Germline mutations of FP/TMEM127 were associated with pheochromocytoma but not paraganglioma and occured in an age group frequently excluded from genetic screening algorithms. Disease-associated mutations disrupt intracellular distribution of the FP/TMEM127 protein. JAMA. 2010;304(23):2611-2619 www.jama.com

Germline mutations in TMEM127 confer susceptibility to pheochromocytoma

Pheochromocytomas, which are catecholamine-secreting tumors of neural crest origin, are frequently hereditary(1). However, the molecular basis of the majority of these tumors is unknown(2). We identified the transmembrane-encoding gene TMEM127 on chromosome 2q11 as a new pheochromocytoma susceptibility gene. In a cohort of 103 samples, we detected truncating germline TMEM127 mutations in approximately 30% of familial tumors and about 3% of sporadic-appearing pheochromocytomas without a known genetic cause. The wild-type allele was consistently deleted in tumor DNA, suggesting a classic mechanism of tumor suppressor gene inactivation. Pheochromocytomas with mutations in TMEM127 are transcriptionally related to tumors bearing NF1 mutations and, similarly, show hyperphosphorylation of mammalian target of rapamycin (mTOR) effector proteins. Accordingly, in vitro gain-of-function and loss-of-function analyses indicate that TMEM127 is a negative regulator of mTOR. TMEM127 dynamically associates with the endomembrane system and colocalizes with perinuclear (activated) mTOR, suggesting a subcompartmental-specific effect. Our studies identify TMEM127 as a tumor suppressor gene and validate the power of hereditary tumors to elucidate cancer pathogenesis.

Differential Antitumor Effects of IgG and IgM Monoclonal Antibodies and Their Synthetic Complementarity-Determining Regions Directed to New Targets of B16F10-Nex2 Melanoma Cells

Malignant melanoma has increased incidence worldwide and causes most skin cancer-related deaths. A few cell surface antigens that can be targets of antitumor immunotherapy have been characterized in melanoma. This is an expanding field because of the ineffectiveness of conventional cancer therapy for the metastatic form of melanoma. In the present work, antimelanoma monoclonal antibodies (mAbs) were raised against B16F10 cells (subclone Nex4, grown in murine serum), with novel specificities and antitumor effects in vitro and in vivo. MAb A4 (IgG2ak) recognizes a surface antigen on B16F10-Nex2 cells identified as protocadherin beta(13). It is cytotoxic in vitro and in vivo to B16F10-Nex2 cells as well as in vitro to human melanoma cell lines. MAb A4M (IgM) strongly reacted with nuclei of permeabilized murine tumor cells, recognizing histone 1. Although it is not cytotoxic in vitro, similarly with mAb A4, mAb A4M significantly reduced the number of lung nodules in mice challenged intravenously with B16F10-Nex2 cells. The V(H) CDR3 peptide from mAb A4 and V(L) CDR1 and CDR2 from mAb A4M showed significant cytotoxic activities in vitro, leading tumor cells to apoptosis. A cyclic peptide representing A4 CDR H3 competed with mAb A4 for binding to melanoma cells. MAb A4M CDRs L1 and L2 in addition to the antitumor effect also inhibited angiogenesis of human umbilical vein endothelial cells in vitro. As shown in the present work, mAbs A4 and A4M and selected CDR peptides are strong candidates to be developed as drugs for antitumor therapy for invasive melanoma.

Vertical alveolar crest bone maintenance around implants in two-stage surgery: an in situ study in dogs

The aim of this study was to evaluate in situ changes in the alveolar crest bone height around immediate implant-supported crowns in comparison to tooth-supported crowns (control) with the cervical margins located at the bone crest level, without occlusal load. In Group I, after extraction of 12 mandibular premolars from 4 adult dogs, implants from Branemark System (MK III TiU RP 4.0 x 11.5 mm) were placed to retain complete acrylic crowns. In Group II, premolars were prepared to receive complete metal crowns. Sixteen weeks after placement of the crowns (38 weeks after tooth extraction), the height of the alveolar bone crest was measured with a digital caliper. Data were analyzed statistically by the Mann-Whitney test at 5% significance level. The in situ analysis showed no statistically significant difference (p=0.880) between the implant-supported and the tooth-supported groups (1.528 + 0.459 mm and 1.570 + 0.263 mm, respectively). Based on the findings of the present study, it may be concluded that initial peri-implant bone loss may result from the remodeling process necessary to establish the biological space, similar to which occurs with tooth-supported crowns.

Central chemoreceptors and neural mechanisms of cardiorespiratory control

The arterial partial pressure (P CO2) of carbon dioxide is virtually constant because of the close match between the metabolic production of this gas and its excretion via breathing. Blood gas homeostasis does not rely solely on changes in lung ventilation, but also to a considerable extent on circulatory adjustments that regulate the transport of CO2 from its sites of production to the lungs. The neural mechanisms that coordinate circulatory and ventilatory changes to achieve blood gas homeostasis are the subject of this review. Emphasis will be placed on the control of sympathetic outflow by central chemoreceptors. High levels of CO2 exert an excitatory effect on sympathetic outflow that is mediated by specialized chemoreceptors such as the neurons located in the retrotrapezoid region. In addition, high CO2 causes an aversive awareness in conscious animals, activating wake-promoting pathways such as the noradrenergic neurons. These neuronal groups, which may also be directly activated by brain acidification, have projections that contribute to the CO2-induced rise in breathing and sympathetic outflow. However, since the level of activity of the retrotrapezoid nucleus is regulated by converging inputs from wake-promoting systems, behavior-specific inputs from higher centers and by chemical drive, the main focus of the present manuscript is to review the contribution of central chemoreceptors to the control of autonomic and respiratory mechanisms.

Engagement of multifocal neural circuits during recall of autobiographical happy events

Happy emotional states have not been extensively explored in functional magnetic resonance imaging studies using autobiographic recall paradigms. We investigated the brain circuitry engaged during induction of happiness by standardized script-driven autobiographical recall in 11 healthy subjects (6 males), aged 32.4 ± 7.2 years, without physical or psychiatric disorders, selected according to their ability to vividly recall personal experiences. Blood oxygen level-dependent (BOLD) changes were recorded during auditory presentation of personal scripts of happiness, neutral content and negative emotional content (irritability). The same uniform structure was used for the cueing narratives of both emotionally salient and neutral conditions, in order to decrease the variability of findings. In the happiness relative to the neutral condition, there was an increased BOLD signal in the left dorsal prefrontal cortex and anterior insula, thalamus bilaterally, left hypothalamus, left anterior cingulate gyrus, and midportions of the left middle temporal gyrus (P < 0.05, corrected for multiple comparisons). Relative to the irritability condition, the happiness condition showed increased activity in the left insula, thalamus and hypothalamus, and in anterior and midportions of the inferior and middle temporal gyri bilaterally (P < 0.05, corrected), varying in size between 13 and 64 voxels. Findings of happiness-related increased activity in prefrontal and subcortical regions extend the results of previous functional imaging studies of autobiographical recall. The BOLD signal changes identified reflect general aspects of emotional processing, emotional control, and the processing of sensory and bodily signals associated with internally generated feelings of happiness. These results reinforce the notion that happiness induction engages a wide network of brain regions.

Neural networks and statistical analysis for classification of corneal videokeratography maps based on Zernike coefficients: a quantitative comparison

PURPOSE: The main goal of this study was to develop and compare two different techniques for classification of specific types of corneal shapes when Zernike coefficients are used as inputs. A feed-forward artificial Neural Network (NN) and discriminant analysis (DA) techniques were used. METHODS: The inputs both for the NN and DA were the first 15 standard Zernike coefficients for 80 previously classified corneal elevation data files from an Eyesys System 2000 Videokeratograph (VK), installed at the Departamento de Oftalmologia of the Escola Paulista de Medicina, São Paulo. The NN had 5 output neurons which were associated with 5 typical corneal shapes: keratoconus, with-the-rule astigmatism, against-the-rule astigmatism, "regular" or "normal" shape and post-PRK. RESULTS: The NN and DA responses were statistically analyzed in terms of precision ([true positive+true negative]/total number of cases). Mean overall results for all cases for the NN and DA techniques were, respectively, 94% and 84.8%. CONCLUSION: Although we used a relatively small database, results obtained in the present study indicate that Zernike polynomials as descriptors of corneal shape may be a reliable parameter as input data for diagnostic automation of VK maps, using either NN or DA.

Learning processes and the neural analysis of conditioning

Classical and operant conditioning principles, such as the behavioral discrepancy-derived assumption that reinforcement always selects antecedent stimulus and response relations, have been studied at the neural level, mainly by observing the strengthening of neuronal responses or synaptic connections. A review of the literature on the neural basis of behavior provided extensive scientific data that indicate a synthesis between the two conditioning processes based mainly on stimulus control in learning tasks. The resulting analysis revealed the following aspects. Dopamine acts as a behavioral discrepancy signal in the midbrain pathway of positive reinforcement, leading toward the nucleus accumbens. Dopamine modulates both types of conditioning in the Aplysia mollusk and in mammals. In vivo and in vitro mollusk preparations show convergence of both types of conditioning in the same motor neuron. Frontal cortical neurons are involved in behavioral discrimination in reversal and extinction procedures, and these neurons preferentially deliver glutamate through conditioned stimulus or discriminative stimulus pathways. Discriminative neural responses can reliably precede operant movements and can also be common to stimuli that share complex symbolic relations. The present article discusses convergent and divergent points between conditioning paradigms at the neural level of analysis to advance our knowledge on reinforcement.

A new species of Baurusuchus (Crocodyliformes, Mesoeucrocodylia) from the Upper Cretaceous of Brazil, with the first complete postcranial skeleton described for the family Baurusuchidae

The present work describes a new species of Baurusuchidae from Upper Cretaceous sediments of the Bauru Basin, and provides the first complete postcranial description for the family. Many postcranial features observed in the new species are also present in other notosuchian taxa, and are thus considered plesiomorphic for the genus. These are: long cervical neural spines; robust deltopectoral crest of the humerus; large proximal portion in the radiale that contacts the ulna; ulnare anterior distal projection; supra-acetabular crest well developed laterally; post-acetabular process posterodorsally deflected; presence of an anteromedial crest in the femur; fourth trocanter of femur posteriorly positioned; tibia with a laterally curved shaft; calcaneum tuber posteroventrally oriented; osteoderms ornamented with grooves and imbricated in the tail. On the other hand, we found the following sacral and carpal features to be unique among all mesoeucrocodylians analyzed: transverse processes of sacral vertebrae dorsolaterally deflected; presence of a longitudinal crest in the lateral surface of sacral vertebrae; pisiform carpal with a condyle-like surface. The majority of these cited features corroborates a cursorial locomotion for the new species described in the present study, suggesting that members of the family Baurusuchidae were also cursorial species.

Os histricomorfos sul-americanos: uma análise comparativa do desenvolvimento embriológico

O objetivo do estudo foi realizar uma análise embriológica dos roedores histricomorfos (paca, cutia, preá e capivara), a fim de comparar com a de outros roedores e com a morfogênese humana um padrão embriológico. Utilizaram-se 8 espécimes de roedores sendo, 2 embriões para cada espécie coletada, ambas em inicio de gestação. Estes foram retirados dos úteros gestantes através de ovariosalpingohisterectomia parcial, seguido de fixação em solução de paraformaldeído 4%. Para as mensurações de Crow-Rump, adotou-se como referência a crista nucal numa extremidade e da última vértebra sacral na extremidade oposta. De forma geral, os embriões analisados mostraram as seguintes características morfológicas: divisão dos arcos branquiais, o não fechamento do neuróporo cranial em alguns embriões estudados, a curvatura cranial acentuada e os somitos delimitados e individualizados. O broto dos membros apresentava-se em desenvolvimento em formato de remo, além da impressão cardíaca e fígado. Na região caudal, visualizou-se a curvatura crânio-caudal, a vesícula óptica com e sem pigmentação da retina, a abertura do tubo neural na região do quarto ventrículo encefálico, a fosseta nasal e a formação das vesículas encefálicas. Concluímos que desenvolvimento embriológico dos roedores histricomorfos pode ser comparado à morfogênese de ratos, cobaios, coelhos e humanos nos diferentes estágios de desenvolvimento, tomando apenas o cuidado com as particularidades de cada espécie, além da implementação de tecnologias reprodutivas, especialmente a de embriões, a qual requer o conhecimento do desenvolvimento pré-implantação referente às fases de desenvolvimento.

Evaluation of follicular lymphoid depletion in the Bursa of Fabricius: an alternative methodology using digital image analysis and artificial neural networks

Fifty Bursa of Fabricius (BF) were examined by conventional optical microscopy and digital images were acquired and processed using Matlab® 6.5 software. The Artificial Neuronal Network (ANN) was generated using Neuroshell® Classifier software and the optical and digital data were compared. The ANN was able to make a comparable classification of digital and optical scores. The use of ANN was able to classify correctly the majority of the follicles, reaching sensibility and specificity of 89% and 96%, respectively. When the follicles were scored and grouped in a binary fashion the sensibility increased to 90% and obtained the maximum value for the specificity of 92%. These results demonstrate that the use of digital image analysis and ANN is a useful tool for the pathological classification of the BF lymphoid depletion. In addition it provides objective results that allow measuring the dimension of the error in the diagnosis and classification therefore making comparison between databases feasible.

Classification of breast masses using a committee machine of artificial neural networks

This work proposes a new approach using a committee machine of artificial neural networks to classify masses found in mammograms as benign or malignant. Three shape factors, three edge-sharpness measures, and 14 texture measures are used for the classification of 20 regions of interest (ROIs) related to malignant tumors and 37 ROIs related to benign masses. A group of multilayer perceptrons (MLPs) is employed as a committee machine of neural network classifiers. The classification results are reached by combining the responses of the individual classifiers. Experiments involving changes in the learning algorithm of the committee machine are conducted. The classification accuracy is evaluated using the area A. under the receiver operating characteristics (ROC) curve. The A, result for the committee machine is compared with the A, results obtained using MLPs and single-layer perceptrons (SLPs), as well as a linear discriminant analysis (LDA) classifier Tests are carried out using the student's t-distribution. The committee machine classifier outperforms the MLP SLP, and LDA classifiers in the following cases: with the shape measure of spiculation index, the A, values of the four methods are, in order 0.93, 0.84, 0.75, and 0.76; and with the edge-sharpness measure of acutance, the values are 0.79, 0.70, 0.69, and 0.74. Although the features with which improvement is obtained with the committee machines are not the same as those that provided the maximal value of A(z) (A(z) = 0.99 with some shape features, with or without the committee machine), they correspond to features that are not critically dependent on the accuracy of the boundaries of the masses, which is an important result. (c) 2008 SPIE and IS&T.

Activation of Neural and Pluripotent Stem Cell Signatures Correlates with Increased Malignancy in Human Glioma

The presence of stem cell characteristics in glioma cells raises the possibility that mechanisms promoting the maintenance and self-renewal of tissue specific stem cells have a similar function in tumor cells. Here we characterized human gliomas of various malignancy grades for the expression of stem cell regulatory proteins. We show that cells in high grade glioma co-express an array of markers defining neural stem cells (NSCs) and that these proteins can fulfill similar functions in tumor cells as in NSCs. However, in contrast to NSCs glioma cells co-express neural proteins together with pluripotent stem cell markers, including the transcription factors Oct4, Sox2, Nanog and Klf4. In line with this finding, in high grade gliomas mesodermal-and endodermal-specific transcription factors were detected together with neural proteins, a combination of lineage markers not normally present in the central nervous system. Persistent presence of pluripotent stem cell traits could only be detected in solid tumors, and observations based on in vitro studies and xenograft transplantations in mice imply that this presence is dependent on the combined activity of intrinsic and extrinsic regulatory cues. Together these results demonstrate a general deregulated expression of neural and pluripotent stem cell traits in malignant human gliomas, and indicate that stem cell regulatory factors may provide significant targets for therapeutic strategies.