INSECT SCIENCE, Issue 2 2003
LIN Xin-da
Abstract Wg/Wnt signaling is a key signaling pathway in Drosophila. Many genes involved in Wingless(wg) signal transduction pathway downstream of Wg, or it s vertebrate Wg homologue Wnt, have been identified. Transduction of the Wg signal downstream of Wg is mediated by nuclear TCF/LEF-1, through association with Armadillo (Arm),-catenin. Pygopus (pygo) is a new identified component in this pathway. Cellular localization experiment showed that pygo was expressed specifically in the nucleus. The expression profile of pygo in embryos was examined using in situ hybridization. Although pygo expressed ubiquitously in the embryos, it expressed at relatively high level in pre-blastoderm embryos which indicate a high degree of maternally provided message, followed by a low level of ubiquitous zygotic expression. This continues into larval tissues (including wing disc, eye disc and leg disc), where pygo appears to be expressed at low level. Comparison of pygo expression levels, in the wing disc, eye disc and leg disc, showed pygo expression level in the wing disc pouch and leg disc were relative higher. [source]

EVIDENCE FOR A SPECIALIZED LOCALIZATION OF THE CHLOROPLAST ATP-SYNTHASE SUBUNITS ,, ,, AND , IN THE EYESPOT APPARATUS OF CHLAMYDOMONAS REINHARDTII (CHLOROPHYCEAE),

JOURNAL OF PHYCOLOGY, Issue 2 2007
Melanie Schmidt
The eyespot apparatus (EA) of Chlamydomonas reinhardtii P. A. Dang. consists of two layers of carotenoid-rich lipid globules subtended by thylakoids. The outermost globule layer is additionally associated with the chloroplast envelope membranes and the plasma membrane. In a recent proteomic approach, we identified 202 proteins from isolated EAs of C. reinhardtii via at least two peptides, including, for example, structural components, signalling-related proteins, and photosynthetic-related membrane proteins. Here, we have analyzed the proteins of the EA with regard to their topological distribution using thermolysin to find out whether the arrangement of globules and membranes provides protection mechanisms for some of them. From about 230 protein spots separated on two-dimensional gels, the majority were degraded by thermolysin. Five major protein spots were protected against the action of this protease. These proteins and some that were degradable were identified by mass spectrometry. Surprisingly, the thermolysin-resistant proteins represented the , and , subunits of the soluble CF1 complex of the chloroplast ATP synthase. Degradable proteins included typical membrane proteins like LHCs, demonstrating that thermolysin is not in general sterically prevented by the EA structure from reaching membrane-associated proteins. A control experiment showed that the CF1 complex of thylakoids is efficiently degraded by thermolysin. Blue native PAGE of thermolysin-treated EAs followed by SDS-PAGE revealed that the , and , subunits are present in conjunction with the , subunit in a thermolysin-resistant complex. These results provide strong evidence that a significant proportion of these ATP-synthase subunits have a specialized localization and function within the EA of C. reinhardtii. [source]

Perinatal development of the rat kidney: Apoptosis and epidermal growth factor

CONGENITAL ANOMALIES, Issue 3 2003
Toshiya Okada
ABSTRACT, Localization of apoptotic cells in the kidney of perinatal rats was examined by the terminal deoxynucleotidyl transferase,mediated d,UTP,biotin nick end labeling (TUNEL) method and electron microscopy. Perinatal changes in the percentage of kidney cells with DNA fragmentation were determined by flow cytometric analysis. Through observation of two successive sections, the relationship between the localization of the epidermal growth factor receptor (EGFR) positive cells and TUNEL positive cells in the kidney was determined. From fetal day 18 to neonatal day 5, TUNEL positive cells were noted in immature glomeruli, collecting ducts and interstitium. Electron microscopically, chromatin condensed nuclei and apoptotic bodies were seen in the same tissue component as the TUNEL positive cells. The percentage of DNA fragmented cells significantly increased from fetal days 18 to 20 and significantly decreased from fetal days 20 to 22, while they still remained low in the neonatal period. The TUNEL positive cells in immature glomeruli and collecting ducts were not reactive to the EGFR antibody. The TUNEL positive cells were not observed in the proximal tubular cells, which were positive to EGFR antibody. These results indicate that apoptotic cells are present in the kidney throughout the perinatal period in the rat and that EGF plays an important role in perinatal development of the rat kidney. [source]

Localization and functional characterization of the human NKCC2 isoforms

ACTA PHYSIOLOGICA, Issue 3 2010
I. Carota
Abstract Aim:, Salt reabsorption across the apical membrane of cells in the thick ascending limb (TAL) of Henle is primarily mediated by the bumetanide-sensitive Na+/K+/2Cl, cotransporter NKCC2. Three full-length splice variants of NKCC2 (NKCC2B, NKCC2A and NKCC2F) have been described. The NKCC2 isoforms have specific localizations and transport characteristics, as assessed for rabbit, rat and mouse. In the present study, we aimed to address the localization and transport characteristics of the human NKCC2 isoforms. Methods:, RT-PCR, in situ hybridization and uptake studies in Xenopus oocytes were performed to characterize human NKCC2 isoforms. Results:, All three classical NKCC2 isoforms were detected in the human kidney; in addition, we found splice variants with tandem duplicates of the variable exon 4. Contrary to rodents, in which NKCC2F is the most abundant NKCC2 isoform, NKCC2A was the dominant isoform in humans; similarly, isoform-specific in situ hybridization showed high expression levels of human NKCC2A along the TAL. Compared to NKCC2B and NKCC2F, human NKCC2A had the lowest Cl, affinity as determined by 86Rb+ uptake studies in oocytes. All NKCC2 isoforms were more efficiently inhibited by bumetanide than by furosemide. A sequence analysis of the amino acids encoded by exon 4 variants revealed high similarities between human and rodent NKCC2 isoforms, suggesting that differences in ion transport characteristics between species may be related to sequence variations outside the highly conserved sequence encoded by exon 4. Conclusion:, The human NKCC2 is an example of how differential splicing forms the basis for a diversification of transporter protein function. [source]

An Investigation of Localization as an Element of Cognitive Fit in Accounting Model Representations

DECISION SCIENCES, Issue 1 2001
Cheryl Dunn
Abstract Cognitive fit, a correspondence between task and data representation format, has been demonstrated to lead to superior task performance by individual users and has been posited as an explanation for performance differences among users of various problem representations such as tables, graphs, maps, and schematic faces. The current study extends cognitive fit to accounting models and integrates cognitive fit theory with the concept of localization to provide additional evidence for how cognitive fit works. Two accounting model representations are compared in this study, the traditional DCA (Debit-Credit-Account) accounting model and the REA (Resources-Events-Agents) accounting model. Results indicate that the localization of relevant objects or linkages is important in establishing cognitive fit. [source]

Localization of Indian hedgehog and PTH/PTHrP receptor expression in relation to chondrocyte proliferation during mouse bone development

DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 2 2005
Helen E. MacLean
We have developed a useful approach to examine the pattern of gene expression in comparison to cell proliferation, using double in situ hybridization and immunofluorescence. Using this system, we examined the expression of Indian hedgehog (Ihh) and PTH/PTHrP receptor (PPR) mRNA in relation to chondrocyte proliferation during embryonic mouse bone development. Both genes are expressed strongly in prehypertrophic and early hypertrophic chondrocytes, and there is a strong correlation between upregulation of both Ihh and PPR expression and chondrocyte cell cycle arrest. At embryonic day (E14.5), PPR mRNA upregulation begins in the columnar chondrocytes just prior to cell cycle exit, but at later time points expression is only observed in the postproliferative region. In contrast, Ihh mRNA expression overlaps slightly with the region of columnar proliferating chondrocytes at all stages. This study provides further evidence that in the developing growth plate, cell cycle exit and upregulation of Ihh and PPR mRNA expression are coupled. [source]

Localization of Lutheran, a novel laminin receptor, in normal, knockout, and transgenic mice suggests an interaction with laminin ,5 in vivo

DEVELOPMENTAL DYNAMICS, Issue 1 2001
Casey L. Moulson
Abstract Laminins are major components of all basement membranes. One laminin that has garnered particular interest, due to its widespread expression pattern and importance during development, is the laminin ,5 chain. In vitro studies have suggested that the Lutheran blood group glycoprotein/basal cell adhesion molecule (Lu), an Ig superfamily transmembrane protein, is a receptor for laminins containing the ,5 chain. However, there are no in vivo studies showing that these proteins are capable of interacting in tissues. We have isolated the mouse ortholog of Lu and characterized its expression and localization in mouse tissues. Lu was primarily found on the basal surface of epithelial cells and on muscle cells adjacent to basement membranes containing laminin ,5. In addition, there was both a dramatic reduction in the basal concentration of Lu in mice lacking laminin ,5, and a significant increase in Lu protein in transgenic mice overexpressing laminin ,5. Together, these data provide the first in vivo evidence for an interaction between Lu and laminin ,5 and support the hypothesis that Lu is a laminin ,5 receptor. We propose that laminin ,5 is involved in concentrating Lu on the basal surface of epithelial cells. This may be one mechanism by which basement membrane signals are transmitted to the cell. © 2001 Wiley-Liss, Inc. [source]

Localization of estrogen receptor-, and -,mRNA in brain areas controlling sexual behavior in Japanese quail

DEVELOPMENTAL NEUROBIOLOGY, Issue 2 2006
Krister Halldin
Abstract Two estrogen receptors (ERs), denoted ER, and ER,, have been identified in humans and various animal species, including the Japanese quail. Estrogens play a key role in sexual differentiation and in activation of sexual behavior in Japanese quail. The distribution of ER, in the brain of male and female adult quail has previously been studied using immunohistochemistry, whereas in situ hybridization has been employed to study the distribution of ER, mRNA in males only. In this article, we used in situ hybridization to study the distribution of mRNAs for both ER, and ER, in brain areas controlling sexual behavior of Japanese quail. Our results show that both ER, mRNA and ER, mRNA are localized in areas important for sexual behavior, such as the preoptic area and associated limbic areas, in both males and females. Moreover, we found differences in distribution of mRNA for the two receptors in these areas. The results of this article support previously reported data and provide novel data on localization of ER mRNAs in adult quail brain of both sexes. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2005 [source]

Localization of KCNC1 (Kv3.1) potassium channel subunits in the avian auditory nucleus magnocellularis and nucleus laminaris during development

DEVELOPMENTAL NEUROBIOLOGY, Issue 2 2003
Suchitra Parameshwaran-Iyer
Abstract The KCNC1 (previously Kv3.1) potassium channel, a delayed rectifier with a high threshold of activation, is highly expressed in the time coding nuclei of the adult chicken and barn owl auditory brainstem. The proposed role of KCNC1 currents in auditory neurons is to reduce the width of the action potential and enable neurons to transmit high frequency temporal information with little jitter. Because developmental changes in potassium currents are critical for the maturation of the shape of the action potential, we used immunohistochemical methods to examine the developmental expression of KCNC1 subunits in the avian auditory brainstem. The KCNC1 gene gives rise to two splice variants, a longer KCNC1b and a shorter KCNC1a that differ at the carboxy termini. Two antibodies were used: an antibody to the N-terminus that does not distinguish between KCNC1a and b isoforms, denoted as panKCNC1, and another antibody that specifically recognizes the C terminus of KCNC1b. A comparison of the staining patterns observed with the panKCNC1 and the KCNC1b specific antibodies suggests that KCNC1a and KCNC1b splice variants are differentially regulated during development. Although panKCNC1 immunoreactivity is observed from the earliest time examined in the chicken (E10), a subcellular redistribution of the immunoproduct was apparent over the course of development. KCNC1b specific staining has a late onset with immunostaining first appearing in the regions that map high frequencies in nucleus magnocellularis (NM) and nucleus laminaris (NL). The expression of KCNC1b protein begins around E14 in the chicken and after E21 in the barn owl, relatively late during ontogeny and at the time that synaptic connections mature morphologically and functionally. © 2003 Wiley Periodicals, Inc. J Neurobiol 55: 165,178, 2003 [source]

Localization analysis in softening RC frame structures

EARTHQUAKE ENGINEERING AND STRUCTURAL DYNAMICS, Issue 2 2003
Ali R. Khaloo
Abstract This paper discusses the sensitivity of softening reinforced concrete frame structures to the changes in input ground motion and investigates the possibility of localizations for this type of structure in static and dynamic analysis. A finite element model is used in which the sections resisting force are calculated using a proposed differential hysteretic model. This model is especially developed for modelling softening behaviour under cyclic loading. To obtain parameters of the differential model the moment,curvature of each section is evaluated using a microplane constitutive law for concrete and bi-linear elasto-plastic law for reinforcements. The capability of the procedure is verified by comparing results with available experimental data at element level, which shows good accuracy of the procedure. The effect of possible changes in ground motion is assessed using a non-stationary Kanai,Tajimi process. This process is used to generate ground motions with approximately the same amplitude and frequency content evolution as those of base ground motion. The possibility of localization in static and dynamic loading is investigated using two structures. A measure for the possibility of localization in code-designed structures is obtained. This study indicates that localization may occur in ordinary moment-resisting structures located in high seismic zones. Localization may result in substantial drift in global response and instability due to P,, effect. Also, it is shown that the structure becomes very sensitive to the input ground motion. It is concluded that allowance by some design codes of the use of ordinary moment-resisting frames in regions with high seismicity should be revised or improvements should be made in the detailing requirements at critical sections of these structures. Copyright © 2002 John Wiley & Sons, Ltd. [source]

Positional Localization: Three-Dimensional Transthoracic Echocardiography Techniques for the Measurement of Cardiac Mass, Volume, and Function

ECHOCARDIOGRAPHY, Issue 8 2000
Andrew M. Keller M.D.
An accurate and reproducible determination of cardiac volume and mass is important for the selection and timing of therapeutic interventions. Quantitative three-dimensional echocardiography has evolved to provide these measurements with the use of a noninvasive, readily available, and inexpensive technique. We introduce and review the principle of positional localization as well as the clinical application of this technique for the measurement of cardiac volume and mass. [source]

Localization of processes involved in methanogenic degradation of rice straw in anoxic paddy soil

ENVIRONMENTAL MICROBIOLOGY, Issue 8 2001
Kristin Glissmann
In anoxic paddy soil, rice straw is decomposed to CH4 and CO2 by a complex microbial community consisting of hydrolytic, fermenting, syntrophic and methanogenic microorganisms. Here, we investigated which of these microbial groups colonized the rice straw and which were localized in the soil. After incubation of rice straw in anoxic soil slurries for different periods, the straw pieces were removed from the soil, and both slurry and straw were studied separately. Although the potential activities of polysaccharolytic enzymes were higher in the soil slurry than in the straw incubations, the actual release of reducing sugars was higher in the straw incubations. The concentrations of fermentation products, mainly acetate and propionate, increased steadily in the straw incubations, whereas only a little CH4 was formed. In the soil slurries, on the other hand, fermentation products were low, whereas CH4 production was more pronounced. The production of CH4 or of fermentation products in the separated straw and soil incubations accounted in sum for 54,82% of the CH4 formed when straw was not removed from the soil. Syntrophic propionate degradation to acetate, CO2 and H2 was thermodynamically more favourable in the soil than in the straw fraction. These results show that hydrolysis and primary fermentation reactions were mainly localized on the straw pieces, whereas the syntrophic and methanogenic reactions were mainly localized in the soil. The percentage of bacterial relative to total microbial 16S rRNA content was higher on the straw than in the soil, whereas it was the opposite for the archaeal 16S rRNA content. It appears that rice straw is mainly colonized by hydrolytic and fermenting bacteria that release their fermentation products into the soil pore water where they are further degraded to CH4. Hence, complete methanogenic degradation of straw in rice soil seems to involve compartmentalization. [source]

Vapor phase transport of unexploded ordnance compounds through soils

ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 10 2002
Raghunathan Ravikrishna
Abstract Unexploded ordnance(UXO) is a source of concern at several U.S. Department of Defense(DOD) sites. Localization of munitions and fate and transport of the explosive compounds from these munitions are a major issue of concern. A set of laboratory experiments were conducted in specially designed flux chambers to measure the evaporative flux of three explosive compounds (2,4-dinitrotoluene, 2,6-dinitrotoluene, and 1,3-dinitrobenzene) from three different soils. The effect of different soil moisture contents, the relative humidity of air contacting the soil surface, and soil temperature on the chemical fluxes were evaluated. A diffusion model was used to describe the chemical transport mechanism in the soil pore air. The soil-air partition constant was treated as a fit parameter in the model because of the uncertainty in the a priori estimation. The model predicts the qualitative trends of the experimental fluxes satisfactorily. Under extremely dry conditions, the flux decreased more rapidly than that predicted by the model. The fluxes from soils at 24°C were higher than those at 14°C, indicating a larger volatilization driving force at the higher temperature. [source]

Localization of deposited polycyclic aromatic hydrocarbons in leaves of Plantago

ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 5 2001
Martine I. Bakker
Abstract After deposition to foliage, polycyclic aromatic hydrocarbons (PAHs) may remain on the leaf surface, accumulate in the cuticular wax, or diffuse into the remaining interior of the plant. In a field study, the location of deposited PAHs in the leaves of two Plantago species was determined. To this aim, leaves of Plantago major and Plantago media were divided into three fractions. First, the leaves were washed (wash-off fraction), then cuticular wax was extracted (wax fraction). Finally, the remaining leaf material was extracted (interior fraction). The presence of PAHs could be demonstrated in all three fractions. For both plants, the distribution of PAHs over the three fractions changed with molecular weight (mol wt) of the PAHs. The wash-off fraction increased with increasing molecular weight, likely because high molecular-weight PAHs occur predominantly bound to particles, which can be readily washed off from the leaves. In contrast, the amount of PAHs detected in the interior of the leaves decreased with increasing molecular weight. This can be explained by a slow desorption of the PAHs from the particles and a low diffusion rate of the larger molecules. This study shows that washing reduces the amount of high molecular-weight PAHs on plant surfaces. Therefore, washing of leafy vegetables is important to minimize human dietary intake of PAHs. [source]

Neuromagnetic Source Localization of Epileptiform Activity in Patients with Graphogenic Epilepsy

EPILEPSIA, Issue 11 2006
Naoaki Tanaka
Summary:,Purpose: To clarify the source localization of epileptiform activity by using magnetoencephalography (MEG) in patients with graphogenic epilepsy. Methods: MEG and simultaneous EEG were recorded with a 204-channel whole-head MEG system in two patients with graphogenic epilepsy. During the MEG recordings, the patients performed a set of tasks comprising mental arithmetic calculation, speaking, moving the right arm in a manner resembling writing, writing, and thinking of writing. Equivalent current dipoles (ECD) were calculated for epileptiform discharges on MEG by using a single-dipole model. The ECD were superimposed on the magnetic resonance images of the patients. Results: The task of writing provoked seizures, in which both patients jerked the right arms. Thinking of writing also induced these seizures. In both patients, EEG associated with the seizures showed bursts of spike-and-slow-wave complexes predominantly in the centroparietal region. MEG also showed epileptiform discharges corresponding to the EEG bursts. ECDs obtained from the discharges were clustered in the left centroparietal area. Conclusions: Thinking of writing was a trigger for the seizures, as well as the task of writing. The source of the epileptiform discharge associated with the seizures was localized in the unilateral centroparietal area. The findings suggest that the centroparietal region plays an important role in the pathophysiology underlying these two graphogenic epilepsy cases. [source]

The Ictal Bradycardia Syndrome: Localization and Lateralization

EPILEPSIA, Issue 4 2006
Jeffrey W. Britton
Summary:,Purpose: Previous studies have established the importance of the insular cortex and temporal lobe in cardiovascular autonomic modulation. Some investigators, based on the results of cortical stimulation response, functional imaging, EEG recordings of seizures, and lesional studies, have suggested that cardiac sympathetic and parasympathetic function may be lateralized, with sympathetic representation lateralized to the right insula, and parasympathetic, to the left. These studies have suggested that ictal bradycardia is most commonly a manifestation of activation of the left temporal and insular cortex. However, the evidence for this is inconsistent. We sought to assess critically the predictable value of ictal bradycardia for seizure localization and lateralization. Methods: In this study, we reviewed the localization of seizure activity in 13 consecutive patients with ictal bradycardia diagnosed during prolonged video-EEG monitoring at Mayo Clinic Rochester. The localization of electrographic seizure activity at seizure onset and bradycardia onset was identified in all patients. In addition, we performed a comprehensive review of the ictal bradycardia literature focusing on localization of seizure activity in ictal bradycardia cases. Results: All occurrences of ictal bradycardia in the 13 identified patients were associated with temporal lobe,onset seizures. However, no consistent lateralization of seizure activity was found at onset of seizure activity or at onset of bradycardia in this population. Seizure activity was bilateral at bradycardia onset in nine of 13 patients. The results from the literature review also showed that a predominance of patients had bilateral activity at bradycardia onset; however, more of the ictal bradycardia cases from the literature had left hemispheric localization of seizure onset. Conclusions: Ictal bradycardia most often occurs in association with bilateral hemispheric seizure activity and is not a consistent lateralizing sign in localizing seizure onset. Our data do not support the existence of a strictly unilateral parasympathetic cardiomotor representation in the left hemisphere, as has been suggested. [source]

Consistent Localization of Interictal Epileptiform Activity on EEGs of Patients with Tuberous Sclerosis Complex

EPILEPSIA, Issue 3 2005
Floor E. Jansen
Summary:,Purpose: We addressed consistent localization of focal interictal epileptiform activity on EEGs of patients with tuberous sclerosis complex (TSC) and epilepsy. Methods: Twenty-one patients with TSC with a 10-year history of epilepsy and interictal epileptiform activity in three or more EEG recordings were included. None of the patients had undergone epilepsy surgery. Local maxima of interictal epileptiform activity were measured from 76 EEG traces and 33 EEG reports. Information about the patients' clinical course was extracted from their medical records. Magnetic resonance imaging (MRI) and neuropsychological examinations were performed. Statistical analysis was performed with the Mann,Whitney U test. Results: In eight patients, interictal epileptiform activity was consistently detected in one or two regions (group 1), and in 13 patients, epileptiform activity was detected in three or more regions (group 2). The number of foci increased throughout the disease course in both groups. Age at seizure onset and IQ were significantly higher in group 1. Complex partial seizures occurred more often in the patients of group 1. In 19 of the 21 patients, the most consistent epileptiform activity was localized in the frontotemporal region. Conclusions: Ninety percent of patients with TSC showed at least one region of consistent interictal epileptiform activity. Patients with one or two regions of epileptiform activity were older at seizure onset, often experienced complex partial seizures, and had mild or no mental deficits. These patients may be candidates for epilepsy surgery. [source]

Subtraction SPECT Coregistered to MRI in Focal Malformations of Cortical Development: Localization of the Epileptogenic Zone in Epilepsy Surgery Candidates

EPILEPSIA, Issue 4 2004
Terence J. O'Brien
Summary: Purpose: To determine the extent to which periictal subtraction single-photon emission computed tomography (SPECT) may improve detection and definition of the epileptogenic zone in patients with focal malformations of cortical development (MCDs). Methods: Subtraction SPECT coregistered to magnetic resonance (MR) images (SISCOM) were constructed for 22 consecutive patients with focal MCDs who underwent periictal SPECT injection (18 ictal and four postictal). In the 17 patients who had epilepsy surgery, concordance between the site of SISCOM localization and site of surgical resection was determined by coregistration of SISCOM images with postoperative MRIs. Results: SISCOM images were localizing in 19 (86%) patients, including eight of the 10 with nonlocalizing MRI. Concordance of SISCOM localization was 91% with MRI localization, 93% with scalp ictal EEG localization, and 100% with intracranial EEG localization. Eight patients whose SISCOM localization was concordant with the surgical resection site had lower postoperative seizure frequency scores (SFSs; p = 0.04) and greater postoperative improvement in SFSs (p = 0.05) than the nine patients whose SISCOM was either nonconcordant or nonlocalizing. On multiple regression analysis, a model combining SISCOM concordance with surgical resection site and extent of MRI lesion resection was predictive of postoperative SFS (R2= 0.47; p = 0.03). Conclusions: Periictal subtraction SPECT using the SISCOM technique provides useful information for seizure localization in patients with focal MCDs, even when MRI is nonlocalizing. [source]

Localization of nAChR subunit mRNAs in the brain of Macaca mulatta

EUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2000
Zhi-Yan Han
Abstract We present here a systematic mapping of nAChR subunit mRNAs in Macaca mulatta brain. A fragment, from the transmembrane segments MIII to MIV of Macaca neuronal nAChR subunits was cloned, and shown to exhibit high identity (around 95%) to the corresponding human subunits. Then, specific oligodeoxynucleotides were synthesized for in situ hybridization experiments. Both ,4 and ,2 mRNA signals were widely distributed in the brain, being stronger in the thalamus and in the dopaminergic cells of the mesencephalon. Most brain nuclei displayed both ,4 and ,2 signals with the exception of some basal ganglia regions and the reticular thalamic nucleus which were devoid of ,4 signal. ,6 and ,3 mRNA signals were selectively concentrated in the substantia nigra and the medial habenula. The strongest signals for ,3 or ,4 mRNAs were found in the epithalamus (medial habenula and pineal gland), whereas there were no specific ,3 or ,4 signals in mesencephalic dopaminergic nuclei. ,5 and ,7 mRNA signals were found in several brain areas, including cerebral cortex, thalamus and substantia nigra, although at a lower level than ,4 and ,2. The distribution of ,3, ,4, ,5, ,6, ,7, ,2, ,3 and ,4 subunit mRNAs in the monkey is substantially similar to that observed in rodent brain. Surprisingly, ,2 mRNA signal was largely distributed in the Macaca brain, at levels comparable with those of ,4 and ,2. This observation represents the main difference between rodent and Macaca subunit mRNA distribution and suggests that, besides ,4,2*, ,2,2* nAChRs constitute a main nAChR isoform in primate brain. [source]

Localization of the A kinase anchoring protein AKAP79 in the human hippocampus

EUROPEAN JOURNAL OF NEUROSCIENCE, Issue 4 2000
Attila Sík
Abstract The phosphorylation state of the proteins, regulated by phosphatases and kinases, plays an important role in signal transduction and long-term changes in neuronal excitability. In neurons, cAMP-dependent protein kinase (PKA), protein kinase C (PKC) and calcineurin (CN) are attached to a scaffold protein, A kinase anchoring protein (AKAP), thought to anchor these three enzymes to specific sites of action. However, the localization of AKAP, and the predicted sites of linked phosphatase and kinase activities, are still unknown at the fine structural level. In the present study, we investigated the distribution of AKAP79 in the hippocampus from postmortem human brains and lobectomy samples from patients with intractable epilepsy, using preembedding immunoperoxidase and immunogold histochemical methods. AKAP79 was found in the CA1, presubicular and subicular regions, mostly in pyramidal cell dendrites, whereas pyramidal cells in the CA3, CA2 regions and dentate granule cells were negative both in postmortem and in surgical samples. In some epileptic cases, the dentate molecular layer and hilar interneurons also became immunoreactive. At the subcellular level, AKAP79 immunoreactivity was present in postsynaptic profiles near, but not attached to, the postsynaptic density of asymmetrical (presumed excitatory) synapses. We conclude that the spatial selectivity for the action of certain kinases and phosphatases regulating various ligand- and voltage-gated channels may be ensured by the selective presence of their anchoring protein, AKAP79, at the majority of glutamatergic synapses in the CA1, but not in the CA2/CA3 regions, suggesting profound differences in signal transduction and long-term synaptic plasticity between these regions of the human hippocampus. [source]

Structural Evolution and Copper-Ion Release Behavior of Cu-pHEMA Hybrids Synthesized In Situ,

ADVANCED ENGINEERING MATERIALS, Issue 11 2009
Yen-Yu Liu
Abstract A novel Cu-pHEMA hybrid was successfully prepared by in situ photopolymerization of 2-hydroxyethyl methacrylate (HEMA) monomer in the presence of Cu(II) copper ions, following an in situ chemical reduction. Experimental observations indicate that intermolecular interactions such as the coupling force and hydrogen bonding between the Cu and the hydroxyl groups further stabilize the hybrid structure to a considerable extent. Localization of the metallic copper particles within the pHEMA network structure as a result of those intermolecular interactions gives rise to the formation of discretely distributed nanocrystallites with particle sizes ranging from 5 to 25,nm in diameter. A crystallographic change of the Cu nanophase from an amorphous-like to a crystalline structure is observed as the H2O:HEMA molar ratio increases, upon synthesis, accompanied with an increase in the particle size. A relatively slow and sustained release of the Cu (in the form of cupric ions) from the hybrids was measured for a time period of about 10 days, which also illustrates a Cu(II)-induced proliferation of the endothelial cells over a relatively small range of release rate of the Cu from the hybrids. Such a new type of Cu-loaded hybrid hydrogel is expected to be compatible and may be considered as a candidate biomaterial for biomedical/therapeutic uses. [source]

Localization of RNAs in oocytes of Eleutherodactylus coqui, a direct developing frog, differs from Xenopus laevis

EVOLUTION AND DEVELOPMENT, Issue 6 2003
Yvonne M. Beckham
SummaryEleutherodactylus coqui develops directly on land to a frog. The large 3.5-mm oocyte of E. coqui has enough yolk to allow development without a feeding tadpole. In the smaller Xenopus laevis oocyte, 1.3 mm in diameter, mRNAs involved in germ layer formation, such as VegT and Vg1, are localized to the vegetal cortex of the oocyte. We hypothesized that an animal shift has occurred in the localization of the E. coqui Orthologs of VegT and Vg1 due to the large egg size. Through a combination of degenerate reverse transcriptase polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE), we cloned 1634 bp of EcVegT and 1377 bp of EcVg1. Northern blot analysis shows that the lengths of these transcripts are 2.5 kb and 1.3 kb, respectively. This result suggests that we have obtained the complete Vg1 transcript, although this transcript has an extremely short 3, untranslated region compared with X. laevis, 256 bp and 1268 bp, respectively. Zygotic expression of EcVegT closely resembles that of VegT, supporting their orthology. Radioactive RT-PCR and in situ hybridization demonstrated the presence of EcVegT and EcVg1 predominantly near the animal pole of the oocyte. RT-PCR showed that the animal blastomeres, formed from the first horizontal cleavage, inherit half of the EcVegT and EcVg1 transcripts, although they contain only about 1% of the embryo volume. Our results indicate major differences between the molecular organization of the eggs of X. laevis and E. coqui. [source]

Biomimetic Composites: Protein Localization in Silica Nanospheres Derived via Biomimetic Mineralization (Adv. Funct.

ADVANCED FUNCTIONAL MATERIALS, Issue 18 2010
Mater.
Abstract Lysozyme-templated precipitation of silica synthesized by sol-gel chemistry produces a composite material with antimicrobial properties. This study investigates the structural properties of the composite material that allow for retention of the antimicrobial activity of lysozyme. Scanning (SEM) and transmission (TEM) electron microscopy reveal that the composite has a hierarchical structure composed of quasi-spherical structures (,450 nm diameter), which are in turn composed of closely packed spherical structures of ,8,10 nm in diameter. Using small-angle neutron scattering (SANS) with contrast variation, the scattering signatures of the lysozyme and silica within the composite were separated. It was determined that the lysozyme molecules are spatially correlated in the material and form clusters with colloidal silica particles. The size of the clusters determined by SANS agrees well with the structural architecture observed by TEM. BET analysis revealed that the surface area of the composite is relatively low (4.73 m2/g). However, after removal of the protein by heating to 200 °C, the surface area is increased by ,20%. In addition to demonstrating a well organized sol-gel synthesis which generates a functional material with antimicrobial applications, the analysis and modeling approaches described herein can be used for characterizing a wide range of mesoporous and ultrastructural materials. [source]

Protein Localization in Silica Nanospheres Derived via Biomimetic Mineralization

ADVANCED FUNCTIONAL MATERIALS, Issue 18 2010
Mateus B. Cardoso
Abstract Lysozyme-templated precipitation of silica synthesized by sol-gel chemistry produces a composite material with antimicrobial properties. This study investigates the structural properties of the composite material that allow for retention of the antimicrobial activity of lysozyme. Scanning (SEM) and transmission (TEM) electron microscopy reveal that the composite has a hierarchical structure composed of quasi-spherical structures (,450 nm diameter), which are in turn composed of closely packed spherical structures of ,8,10 nm in diameter. Using small-angle neutron scattering (SANS) with contrast variation, the scattering signatures of the lysozyme and silica within the composite were separated. It was determined that the lysozyme molecules are spatially correlated in the material and form clusters with colloidal silica particles. The size of the clusters determined by SANS agrees well with the structural architecture observed by TEM. BET analysis revealed that the surface area of the composite is relatively low (4.73 m2/g). However, after removal of the protein by heating to 200 °C, the surface area is increased by ,20%. In addition to demonstrating a well organized sol-gel synthesis which generates a functional material with antimicrobial applications, the analysis and modeling approaches described herein can be used for characterizing a wide range of mesoporous and ultrastructural materials. [source]

Localization of the mosaic transmembrane serine protease corin to heart myocytes

FEBS JOURNAL, Issue 23 2000
John D. Hooper
Corin cDNA encodes an unusual mosaic type II transmembrane serine protease, which possesses, in addition to a trypsin-like serine protease domain, two frizzled domains, eight low-density lipoprotein (LDL) receptor domains, a scavenger receptor domain, as well as an intracellular cytoplasmic domain. In in vitro experiments, recombinant human corin has recently been shown to activate pro-atrial natriuretic peptide (ANP), a cardiac hormone essential for the regulation of blood pressure. Here we report the first characterization of corin protein expression in heart tissue. We generated antibodies to two different peptides derived from unique regions of the corin polypeptide, which detected immunoreactive corin protein of approximately 125,135 kDa in lysates from human heart tissues. Immunostaining of sections of human heart showed corin expression was specifically localized to the cross striations of cardiac myocytes, with a pattern of expression consistent with an integral membrane localization. Corin was not detected in sections of skeletal or smooth muscle. Corin has been suggested to be a candidate gene for the rare congenital heart disease, total anomalous pulmonary venous return (TAPVR) as the corin gene colocalizes to the TAPVR locus on human chromosome 4. However examination of corin protein expression in TAPVR heart tissue did not show evidence of abnormal corin expression. The demonstrated corin protein expression by heart myocytes supports its proposed role as the pro-ANP convertase, and thus a potentially critical mediator of major cardiovascular diseases including hypertension and congestive heart failure. [source]

Localization of gene products using a chromosomally tagged GFP-fusion library in the fission yeast Schizosaccharomyces pombe

GENES TO CELLS, Issue 2 2009
Aki Hayashi
We constructed a library of chromosomally-tagged green fluorescent protein (GFP) fusions in the fission yeast Schizosaccharomyces pombe. This library contains 1058 strains. In each strain, the coding sequence of GFP is integrated at the 3,-end of a particular chromosomal ORF such that the full-length GFP fusion construct is expressed under the control of the original promoter. Integration of the GFP coding sequence at the authentic chromosomal location of each gene was confirmed by PCR. Microscopic screening of these strains detected sufficient levels of GFP signal in 710 strains and allowed assignment of these GFP-fusion gene products with their intracellular localization: 374 proteins were localized in the nucleus, 65 proteins in the nucleolus, 34 proteins at the nuclear periphery, 27 proteins at the plasma membrane and cytoplasmic membranous structures, 24 proteins at the spindle pole body and microtubules, 92 proteins at cytoplasmic structures, and 94 proteins were uniformly distributed throughout the cytoplasm. [source]

Localization of a putative low-penetrance ependymoma susceptibility locus to 22q11 using a chromosome 22 tiling-path genomic microarray

GENES, CHROMOSOMES AND CANCER, Issue 4 2005
Anneke C. J. Ammerlaan
Ependymomas frequently display allelic loss of chromosome 22 in the absence of mutations in the known tumor-suppressor genes on chromosome 22, suggesting the role of an alternative predisposing gene or genes from this chromosome. In an effort to localize these genes, 37 ependymomas derived from 33 patients were analyzed for the presence of copy number changes by use of a high-resolution chromosome 22 genomic microarray. Eighteen ependymomas (49%) displayed an array-CGH profile consistent with monosomy of chromosome 22. However, in 10 of these tumors, the fluorescence ratios for 22q clones scored as deleted were different from those at the single gene copy level. This suggests either analysis of mixed populations of tumor and normal stromal cells or analysis of mixed tumor cell populations with different genetic profiles. Four ependymomas derived from two patients showed overlapping interstitial deletions of 2.2 Mb and ,510 kb. Further analyses revealed that these deletions were present in the constitutional DNA of these two patients as well as in some of their unaffected relatives. Detailed microsatellite analysis of these families refined the commonly deleted segment to a region of 320 kb between markers RH13801 and D22S419. Our results provide additional evidence for the involvement of genes on chromosome 22 in the development of ependymoma and suggest the presence of a low-penetrance ependymoma susceptibility locus at 22q11. © 2005 Wiley-Liss, Inc. [source]

Pseudomigraine With Lymphocytic Pleocytosis: A Calcium Channelopathy?

HEADACHE, Issue 8 2003
Clinical Description of 10 Cases, Genetic Analysis of the Familial Hemiplegic Migraine Gene CACNA1A
Objective.,To report the clinical findings of 10 patients diagnosed with pseudomigraine with lymphocytic pleocytosis and the results of mutational analysis of the CACNA1A gene in 8 of these patients. Background.,Pseudomigraine with lymphocytic pleocytosis, also referred to as headache with neurologic deficits and cerebrospinal fluid lymphocytosis (HaNDL), is characterized by episodic transient neurologic dysfunction associated with moderate to severe headache and cerebrospinal fluid lymphocytic pleocytosis. Episodes are recurrent and the condition is self-limiting. The etiology of this sporadic condition remains unknown, but the episodic nature and its ability to be triggered by angiography is somewhat reminiscent of the phenotypic features of familial hemiplegic migraine, a condition caused by mutations in the CACNA1A gene. Design/Methods.,Utilizing retrospective chart review, we describe the clinical features of pseudomigraine with lymphocytic pleocytosis in 10 patients. Whole blood was taken from 8 patients (2 were lost to follow-up) and used for DNA testing. The CACNA1A gene was screened for mutations using heteroduplex analysis and direct DNA sequencing. Results.,Clinical features of pseudomigraine with lymphocytic pleocytosis included transient episodes of weakness, sensory and visual symptoms, aphasia, and confusion lasting minutes up to 4 hours. Sensory symptoms, typically affecting the face and arm, were the most common presentation. Localization of symptoms did not conform to vascular territories. Headache was typically throbbing and most often bilateral. Genetic analysis did not identify any mutations in the CACNA1A gene. Conclusions.,Similarities between familial hemiplegic migraine and pseudomigraine with lymphocytic pleocytosis include recurrent headache with reversible neurologic deficit, cerebrospinal fluid lymphocytic pleocytosis, and triggers such as angiography. Even so, heteroduplex analysis and DNA sequencing failed to identify any sporadic mutations or shared polymorphisms in the exons or the intron/exon boundaries of the CACNA1A gene. These results do not support a role of the CACNA1A gene in the etiology of pseudomigraine with lymphocytic pleocytosis. [source]

Localization of hepatitis C virus in gastrointestinal mucosa: A possible reservoir for relapse

HEPATOLOGY, Issue 3 2003
Lucia Miglioresi M.D.
No abstract is available for this article. [source]

Enhanced expression of B7-1, B7-2, and intercellular adhesion molecule 1 in sinusoidal endothelial cells by warm ischemia/reperfusion injury in rat liver

HEPATOLOGY, Issue 4 2001
Naosuke Kojima
To elucidate a role of costimulatory molecule and cell adhesion molecule in hepatic ischemia/reperfusion injury, we examined an alteration in B7-1 (CD80), B7-2 (CD86), and intercellular adhesion molecule 1 (ICAM-1; CD54) expression in the rat liver after warm ischemia/reperfusion injury. To induce hepatic warm ischemia in a rat model, both portal vein and hepatic artery entering the left-lateral and median lobes were occluded by clamping for 30 minutes or 60 minutes, and then reperfused for 24 hours. B7-1, B7-2, and ICAM-1 expressions in the liver were analyzed by immunofluorescence staining and real-time reverse transcription polymerase chain reaction (RT-PCR). Although B7-1 and B7-2 expressions were at very low levels in the liver tissues from normal or sham-operated control rats, both B7-1 and B7-2 expressions were enhanced at protein and messenger RNA (mRNA) levels in the affected, left lobes after warm ischemia/reperfusion. ICAM-1 protein and mRNA were constitutively expressed in the liver of normal and sham-operated control rats, and further up-regulated after warm ischemia/reperfusion. Localization of increased B7-1, B7-2, and ICAM-1 proteins, as well as von Willebrand factor as a marker protein for endothelial cells, was confined by immunofluorescence staining to sinusoidal endothelial cells in hepatic lobules. Data from quantitative real-time RT-PCR analysis revealed that B7-1 and B7-2 mRNA levels were elevated in hepatic lobes after warm ischemia/reperfusion (5.13- and 52.9-fold increase, respectively), whereas ICAM-1 mRNA expression was rather constitutive but further enhanced by warm ischemia/reperfusion (4.24-fold increase). These results suggest that hepatic sinusoidal endothelial cells play a pivotal role as antigen-presenting cells by expressing B7-1 and B7-2 in warm hepatic ischemia/reperfusion injury, and that B7-1 and/or B7-2 might be the primary target to prevent early rejection and inflammatory reactions after hepatic ischemia/reperfusion injury associated with liver transplantation. [source]