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PubMed1991

Maturation of the corneal endothelial tight junction.

Apical tight junctional formation of the rabbit corneal endothelium was examined by freeze-fracture analysis and measurement of paracellular permeability to 5(6)-carboxyfluorescein. Freeze-fracture analysis indicated that apical tight junction formation of the rabbit corneal endothelium is a dynamic process. At birth, there are few tight junctional strands present and a minimal barrier for paracellular diffusion. As the rabbit matures, a more complex network of anastomosing tight junctional strands begins to encircle the cell perimeter under the apical folds. However, even in the mature animal (3 months), there are discontinuities and free ends in the network, thus suggesting that the barrier is not complete even at this stage. Paracellular permeability measurements using 5(6)-carboxyfluorescein as a tracer corroborate these anatomic findings. Endothelial paracellular flux measurements steadily decrease as the rabbit matures from birth to young adult. This indicates that the tight junctional network is increasing in complexity and progressively limiting the flow of substances through the intercellular space.

Animals↗
PubMed1995

A complex network regulates expression of eps and other virulence genes of Pseudomonas solanacearum.

We have discovered an unusual and complex regulatory network used by the phytopathogen Pseudomonas solanacearum to control transcription of eps, which encodes for production of its primary virulence factor, the exopolysaccharide EPS I. The major modules of this network were shown to be three separate signal transduction systems: PhcA, a LysR-type transcriptional regulator, an dual two-component regulatory systems, VsrA/VsrD and VsrB/VsrC. Using lacZ fusions and RNA analysis, we found that both PhcA and VsrA/VsrD control transcription of another network component, xpsR, which in turn acts in conjunction with vsrB/vsrC to increase transcription of the eps promoter by > 25-fold. Moreover, gel shift DNA binding assays showed that PhcA specifically binds to the xpsR promoter region. Thus, the unique XpsR protein interconnects the three signal transduction systems, forming a network for convergent control of EPS I in simultaneous response to multiple environmental inputs. In addition, we demonstrate that each individual signaling system of the network also acts independently to divergently regulate other unique sets of virulence factors. The purpose of this complex network may be to allow this phytopathogen to both coordinately or independently regulate diverse virulence factors in order to cope with the dynamic situations and conditions encountered during interactions with plants.

Amino Acid Sequence↗
PubMed1999

Is lower income associated with greater biopsychosocial morbidity? Implications for physicians working with underserved patients.

BACKGROUND: Previous studies have established a powerful relationship between socioeconomic position and health. However, there has been little attention given to the association between income, biopsychosocial morbidity, and decline in health over time among primary care patients. METHODS: Data were collected using a survey mailed to patients receiving care at a family medicine center and through a follow-up survey mailed 2 years later. The independent association between various biopsychosocial measures and family income was assessed through stepwise linear regression. After controlling for baseline health status, the effect of family income on health status at follow-up was assessed. RESULTS: Data were available from 922 active family medicine patients who responded to the initial survey and from 655 who responded to the follow-up survey. In bivariate analyses, lower family income was significantly associated with poorer health status, greater psychological distress, more family dysfunction, less social support, more behavioral risk factors, higher rates of obesity and uncontrolled blood pressure, poorer physical and mental health status, and more medical diagnoses. In a multivariate analysis, age, sex, marital status, race, social network, family criticism, smoking, fat consumption, and health status were independently associated with family income. After controlling for covariates, including baseline health status, family income was a significant predictor of health status at follow-up. CONCLUSIONS: Family income is associated with biopsychosocial morbidity and health decline. Physicians who care for poorer patients will likely be confronted by challenging and complex biopsychosocial problems.

Adult↗
PubMed1989

Involvement of microtubules and microfilaments in the control of the nuclear movement during maturation of mouse oocyte.

We confirm that the centrifugal migration of the chromosomes in maturing mouse oocytes depends on a microfilament-mediated process. We investigated the role of the cytoskeleton in the germinal vesicle (GV) behavior of oocytes prevented from resuming meiosis by either activators of protein kinase A or activators of protein kinase C. A time-lapse microcinematography study demonstrates that GV immobilization by isobutylmethylxanthine (IBMX) is overcome by colcemid (COL), nocodazole (NOC), and taxol and that cytochalasin D (CCD) reversibly immobilizes the GV of oocytes treated with either IBMX + COL (or NOC) or 12-O-tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C, known to allow a programmed GV cortical translocation. An immunofluorescence analysis shows that the disorganization of a perinuclear microtubule network is the very first cytological clue of maturation. IBMX promotes its persistence while NOC, COL, and taxol induce its immediate disappearance. It is concluded that elements of the cytoplasmic microtubular complex (CMTC) are passively involved in the control of the setting up of a "centrifugal displacement property" (CDP) by counteracting a motive force provided by the microfilament cytoskeleton. Finally, TPA induces a clearcut reorganization instead of a total disorganization of the CMTC. This reorganization is, however, sufficient to allow the microfilaments to drive the GV displacement.

Actin Cytoskeleton↗
PubMed1976

Kindling in the frog: development of spontaneous epileptiform activity.

The present report is the first demonstration of the kindling phenomenon in a non-mammalian, poikilothermic species. Brief (2 sec) localized electrical stimulation was delivered to the hippocampal cortex of one hemisphere of the partially paralyzed bullfrog, Rana catesbeiana, at current levels just sufficient to elicit brief after-discharge (AD) on first application. Succeeding stimulations, once per hour, gave rise to more and more prolonged AD involving first the side stimulated (1 degree focus) and secondly the opposite hemisphere (2 degree focus). In addition to these stimulus-dependent alterations, after several hours spontaneous, paroxysmal discharges occurred in interstimulus intervals. They appeared first in the stimulated hemisphere, then as evoked spikes in the secondary hemisphere, and finally independently on the two sides. The waveshape of the spontaneous epileptiform potentials exhibited an extraordinary constancy from complex to complex which was specific to each animal. Such wave-shape coherence implies a constancy to both the anatomical substrate and the sequency of synaptic engagement each time spontaneous paroxysmal activity invades the network. The model is especially suitable for analysis of temperature-dependent biochemical events responsible for this functional transformation.

Animals↗
PubMed1999

Roles of ephrinB ligands and EphB receptors in cardiovascular development: demarcation of arterial/venous domains, vascular morphogenesis, and sprouting angiogenesis.

Eph receptor tyrosine kinases and their cell-surface-bound ligands, the ephrins, regulate axon guidance and bundling in the developing brain, control cell migration and adhesion, and help patterning the embryo. Here we report that two ephrinB ligands and three EphB receptors are expressed in and regulate the formation of the vascular network. Mice lacking ephrinB2 and a proportion of double mutants deficient in EphB2 and EphB3 receptor signaling die in utero before embryonic day 11.5 (E11.5) because of defects in the remodeling of the embryonic vascular system. Our phenotypic analysis suggests complex interactions and multiple functions of Eph receptors and ephrins in the embryonic vasculature. Interaction between ephrinB2 on arteries and its EphB receptors on veins suggests a role in defining boundaries between arterial and venous domains. Expression of ephrinB1 by arterial and venous endothelial cells and EphB3 by veins and some arteries indicates that endothelial cell-to-cell interactions between ephrins and Eph receptors are not restricted to the border between arteries and veins. Furthermore, expression of ephrinB2 and EphB2 in mesenchyme adjacent to vessels and vascular defects in ephB2/ephB3 double mutants indicate a requirement for ephrin-Eph signaling between endothelial cells and surrounding mesenchymal cells. Finally, ephrinB ligands induce capillary sprouting in vitro with a similar efficiency as angiopoietin-1 (Ang1) and vascular endothelial growth factor (VEGF), demonstrating a stimulatory role of ephrins in the remodeling of the developing vascular system.

Animals↗
PubMed1999

PrM- and cell-binding domains of the dengue virus E protein.

The E-prM proteins of flaviviruses are unusual complexes which play important roles in virus assembly and fusion modulation and in potential immunity-inducing vaccines. Despite their importance, little is known about the biogenesis and structural organization of E-prM complexes. Pulse-chase radiolabeling of dengue virus-infected Vero cells demonstrated a rapid interassociation of E and prM proteins, and sucrose gradient sedimentation analysis suggested that E-prM complexes progressed from simple heteromers to more densely sedimenting structures indicating increased multimerization. E-prM heteromers of even higher complexity were observed in virus particles, suggesting an intracellular assembly process which results in the networking of E-prM subunits into a lattice-like structure found in virus particles. Trypsin cleavage of E-prM-containing virus particles resulted in the release of a soluble 45-kDa fragment of the E protein which retained cell-binding activity. The results suggest that E-prM interactions in dengue virus particles are largely mediated by domains in the carboxy-terminal anchoring domain of E, while cell-binding activity is retained in a trypsin-releasable ectodomain of the E protein.

Animals↗
PubMedDate not supplied

Accurate spike synchronization in cortex.

In view of the enormous capacity and complexity of mammalian brains it is evident that a detailed account of their anatomy and physiology alone cannot lead to a complete understanding of their function. Computer simulation and mathematical analysis of abstract, yet biologically realistic models for neurons and networks yield additional and useful information about the interplay of the underlying anatomical structures, the physiological processes operating on the neuronal substrate, and the resulting brain functions during the performance of behavioral tasks. We discuss some contributions of such experiment-guided theory to the issue of accurate spike synchronization.

Animals↗
PubMed1998

Cytokines and cell adhesion molecules in cerebral ischemia: experimental bases and therapeutic perspectives.

The possibility of reopening an occluded cerebral artery by means of thrombolysis has renewed interest in a number of the several mechanisms that are active during acute cerebral ischemia. Over recent years, it has become apparent that leukocytes play a central role not only during the healing stage of brain infarction but also during the early phases of cerebral ischemia, when it is postulated that these cells produce harmful effects, particularly in the presence of reperfusion. This review is based on the critical analysis of more than 150 publications dealing with the role of leukocytes and some inflammatory mediators (cytokines, chemokines, and adhesion molecules) in cerebral ischemia. Animal studies indicate that leukocyte involvement is promoted by a variety of inflammatory molecules produced immediately after the onset of cerebral ischemia. Considerable experimental evidence suggests that these mediators play a key role in the progression from ischemia to irreversible injury (ie, cellular death and necrosis). However, the precise role of each molecule alone remains to be further elucidated as well as in relation to the complex network existing among different mediators. Progress in our understanding of the inflammatory mechanisms operating in cerebral ischemia has enabled the testing of new compounds with promising results in animals; in contrast, one recent controlled trial of an anti-leukocyte molecule in acute stroke patients showed negative results. This discrepancy may derive in part from our incomplete understanding of the complexity of the inflammatory mechanisms involved in cerebral ischemia. Our analysis suggests that until sufficient knowledge of the underlying disease mechanisms is acquired, more care should be taken when testing new and potentially efficacious drugs in stroke patients.

Animals↗
PubMed1999

Analysis of site-specific phosphorylation of the retinoblastoma protein during cell cycle progression.

Differential phosphorylation of the retinoblastoma protein plays a pivotal role in cell cycle regulation. The retinoblastoma protein is specifically phosphorylated during the cell cycle by cyclin-dependent kinase complexes which intersect with many cellular signaling networks. Since the loss of the retinoblastoma signaling pathways occurs in a wide variety of human tumors, understanding the significance of site-specific phosphorylation can clarify the role of selected cyclin-dependent kinase complexes during cell cycle progression. Here we describe the phosphospecificity and cellular characterization of a panel of polyclonal antibodies that recognize unique phosphorylation sites within the retinoblastoma protein. These reagents were used to validate authentic cellular retinoblastoma phosphorylation sites at amino acids 780, 795, and 807/811 correlating with the G1-S transition.

Animals↗
PubMed1993

Localization of ribosomal DNA in Rhagoletis pomonella (Diptera: Tephritidae) by in situ hybridization.

The ribosomal DNA (rDNA) of Rhagoletis pomonella was localized within both salivary gland polytene chromosomes and somatic cell mitotic chromosomes by in situ hybridization using the heterospecific Drosophila melanogaster rDNA clone, Dm238. In situ hybridization analysis of polytene nuclei showed that R. pomonella rDNA is located in the nucleolus and adjacent granular network of chromosome 1. The site of origin of rDNA is within this isomorphic granular network. The preservation of nucleolar ultrastructure in some polytene chromosome preparations allowed light microscope localization of R. pomonella rDNA to the apparent periphery of fibrillar centres within fibrillar complexes. In somatic cell nuclei, DM238 hybridized to the nucleolus organizing region (NOR) located on chromosome 1 at the site of the secondary constriction. The frequency distribution of heteromorphisms for rDNA content, differential appearance of secondary constrictions, non-pairing of the NOR and differences in homologue lengths suggests that the structural differentiation of this region in chromosome 1 is sex linked. This the first published description of the salivary gland polytene chromosomes from R. pomonella, and we include a tentative karyotype description, polytene chromosome maps and comments on their suitability for banding and molecular analysis.

Animals↗
PubMed1998

Analysis and simulation of complex interactions during dynamic microfiltration of Escherichia coli suspensions.

Microfiltration is an important unit operation in downstream processing. However, due to the influence of membrane fouling, prediction of the filtration performance for biological suspensions is difficult. This paper describes a modeling approach that allows a comprehensive description of filtration performance. On the basis of experimental data and linguistic information, a specific artificial neural network was developed that predicts the process behavior within a certain range of parameters. This approach allows us to analyze influences of fermentation on filtration. By using extensive simulations, the interactions of 17 parameters were examined and the fouling causes determined. The model was developed for cell harvesting of Escherichia coli through a shear-enhanced module. The method can be applied to any cross-flow filtration process.

Bacteriological Techniques↗
PubMed1997

Probabilistic independence networks for hidden Markov probability models.

Graphical techniques for modeling the dependencies of random variables have been explored in a variety of different areas, including statistics, statistical physics, artificial intelligence, speech recognition, image processing, and genetics. Formalisms for manipulating these models have been developed relatively independently in these research communities. In this paper we explore hidden Markov models (HMMs) and related structures within the general framework of probabilistic independence networks (PINs). The paper presents a self-contained review of the basic principles of PINs. It is shown that the well-known forward-backward (F-B) and Viterbi algorithms for HMMs are special cases of more general inference algorithms for arbitrary PINs. Furthermore, the existence of inference and estimation algorithms for more general graphical models provides a set of analysis tools for HMM practitioners who wish to explore a richer class of HMM structures. Examples of relatively complex models to handle sensor fusion and coarticulation in speech recognition are introduced and treated within the graphical model framework to illustrate the advantages of the general approach.

Algorithms↗
PubMed1995

Artificial neural networks for decision support in clinical medicine.

Connectionist models such as neural networks are alternatives to linear, parametric statistical methods. Neural networks are computer-based pattern recognition methods with loose similarities with the nervous system. Individual variables of the network, usually called 'neurones', can receive inhibitory and excitatory inputs from other neurones. The networks can define relationships among input data that are not apparent when using other approaches, and they can use these relationships to improve accuracy. Thus, neural nets have substantial power to recognize patterns even in complex datasets. Neural network methodology has outperformed classical statistical methods in cases where the input variables are interrelated. Because clinical measurements usually derive from multiple interrelated systems it is evident that neural networks might be more accurate than classical methods in multivariate analysis of clinical data. This paper reviews the use of neural networks in medical decision support. A short introduction to the basics of neural networks is given, and some practical issues in applying the networks are highlighted. The current use of neural networks in image analysis, signal processing and laboratory medicine is reviewed. It is concluded that neural networks have an important role in image analysis and in signal processing. However, further studies are needed to determine the value of neural networks in the analysis of laboratory data.

Clinical Medicine↗
PubMed1998

Feed forward neural networks for the analysis of censored survival data: a partial logistic regression approach.

Flexible modelling in survival analysis can be useful both for exploratory and predictive purposes. Feed forward neural networks were recently considered for flexible non-linear modelling of censored survival data through the generalization of both discrete and continuous time models. We show that by treating the time interval as an input variable in a standard feed forward network with logistic activation and entropy error function, it is possible to estimate smoothed discrete hazards as conditional probabilities of failure. We considered an easily implementable approach with a fast selection criteria of the best configurations. Examples on data sets from two clinical trials are provided. The proposed artificial neural network (ANN) approach can be applied for the estimation of the functional relationships between covariates and time in survival data to improve model predictivity in the presence of complex prognostic relationships.

Clinical Trials as Topic↗
PubMedDate not supplied

The connectional organization of the cortico-thalamic system of the cat.

Data on connections between the areas of the cerebral cortex and nuclei of the thalamus are too complicated to analyse with naked intuition. Indeed, the complexity of connection data is one of the major challenges facing neuroanatomy. Recently, systematic methods have been developed and applied to the analysis of the connectivity in the cerebral cortex. These approaches have shed light on the gross organization of the cortical network, have made it possible to test systematically theories of cortical organization, and have guided new electrophysiological studies. This paper extends the approach to investigate the organization of the entire cortico-thalamic network. An extensive collation of connection tracing studies revealed approximately 1500 extrinsic connections between the cortical areas and thalamic nuclei of the cat cerebral hemisphere. Around 850 connections linked 53 cortical areas with each other, and around 650 connections linked the cortical areas with 42 thalamic nuclei. Non-metric multidimensional scaling, optimal set analysis and non-parametric cluster analysis were used to study global connectivity and the 'place' of individual structures within the overall scheme. Thalamic nuclei and cortical areas were in intimate connectional association. Connectivity defined four major thalamo-cortical systems. These included three broadly hierarchical sensory or sensory/motor systems (visual and auditory systems and a single system containing both somatosensory and motor structures). The highest stations of these sensory/motor systems were associated with a fourth processing system composed of prefrontal, cingulate, insular and parahippocampal cortex and associated thalamic nuclei (the 'fronto-limbic system'). The association between fronto-limbic and somato-motor systems was particularly close.

Animals↗
PubMed1978

The Slovak-American "swaddling ethos": homeostat for family dynamics and cultural continuity.

This paper explores the systemic relationships among cultural ethos, family dynamics, personality configuration, and child-rearing patterns among multigeneration Slovak-Americans. The "swaddling ethos" is posited to serve as a homeostat whose regulatory function can be discerned through the analysis of family structure and process, in particular through the explication of values, affective patterns, roles, boundaries, and structural units within the family. The core of the ethos is a dependency-security complex that attaches the individual to an extended family network of obligation, indebtedness, and reciprocity (ethnic, Slovak); induces rebellion against such attachment (mainstream American); and undermines efforts toward separation-individuation, resulting in the perpetuation of an ethnic tradition.

Attitude↗
PubMed1993

Application of artificial neural networks for reduction of false-positive detections in digital chest radiographs.

A methodology based on the fuzzy set theory and the convolution neural network (CNN) architecture is proposed to tackle the problem of reducing false-positive rate in automatic lung nodule detection. The CNN which simulates human visual mechanism was trained by a supervised back-propagation algorithm based on fuzzy membership functions. The training and testing database consists of image blocks (each 32 x 32 pixels) of suspected lung nodule areas (nodule candidates) which were generated from our pre-scanning program [1]. A linguistic label was assigned to each nodule candidate of the training set, then the label was converted to a membership value through a pre-defined membership function and used as teaching signal (desired outputs) during the network learning. Before the nodule candidate was fed to the network input, it was pre-processed to reduce the complex background noise and the contrast discrepancy resulted from film development. During the network testing phase, a defuzzification process was applied to decipher the trained network's output triggered by the nodule candidate in the testing set. Finally, a Receiver Operating Characteristic (ROC) analysis was used to evaluate the CNN's performance based on the defuzzified output of the testing database. Preliminary results showed an average Az (the performance index) of 0.84 which is equivalent to 0.80 true-positive detection (sensitivity) with an average 2-3 false-positive detections per chest image.

Computer Simulation↗