CHEMISTRY & BIODIVERSITY, Issue 6 2007

Abstract Zervamicin IIB (ZER) is a 16-mer peptaibol that produces voltage-dependent conductances in artificial membranes, a property considered responsible for its antimicrobial activity to mainly Gram -positive microorganisms. In addition, ZER appears to inhibit the locomotor activity of the mouse (see elsewhere in this Issue), probably by affecting the brain. To examine whether the electrophysiological properties of the neuronal cells of the central neural system might be possibly influenced by the pore forming ZER, the present study was undertaken as a first attempt to unravel the molecular mechanism of this biological activity. To this end, membrane permeabilization of the neuron-like rat pheochromocytoma cell (PC12) by the channel-forming ZER was studied with the whole-cell patch-clamp technique, and compared with the permeabilizations of the well-known voltage-gated peptaibol alamethicin F50/5 (ALA) and the cation channel-forming peptide-antibiotic gramicidin D (GRAM). While 1,,M GRAM addition to PC12 cells kept at a membrane potential Vm=0,mV causes an undelayed gradual increase of a leak conductance with a negative reversal potential of ca. ,24,mV, ZER and ALA are ineffective at that concentration and potential. However, if ZER and ALA are added in 5,10,,M concentrations while Vm is kept at ,60,mV, they cause a sudden and strong permeabilization of the PC12 cell membrane after a delay of 1,2,min, usually leading to disintegrating morphology changes of the patched cell but not of the surrounding cells of the culture at that time scale. The zero reversal potential of the established conductance is consistent with the known aselectivity of the channels formed. This sudden permeabilization does not occur within 10,20,min at Vm=0,mV, in accordance with the known voltage dependency of ZER and ALA channel formation in artificial lipid membranes. The permeabilizing action of these peptaibols on the culture as a whole is further supported by K+ -release measurements from a PC12 suspension with a K+ -selective electrode. Further analysis suggested that the permeabilizing action is associated with extra- or intracellular calcium effects, because barium inhibited the permeabilizing effects of ZER and ALA. We conclude, for the membrane of the mammalian neuron-like PC12 cell, that the permeabilizing effects of the peptides ZER and ALA are different from those of GRAM, consistent with earlier studies of these peptides in other (artificial) membrane systems. They are increased by cis -positive membrane potentials in the physiological range and may include calcium entry into the PC12 cell. [source]

Cover Picture: Biomineralized Polysaccharide Capsules for Encapsulation, Organization, and Delivery of Human Cell Types and Growth Factors (Adv. Funct.

ADVANCED FUNCTIONAL MATERIALS, Issue 6 2005
Mater.
Abstract The cover shows biomineralized polysaccharide capsules with specifiable make-up, which can provide microenvironments for stabilization, growth, and differentiation of human cell types, as reported by Oreffo and co-workers on p.,917. The capsules are amenable to complexation with a range of bioactive molecules and cells, offering tremendous potential as multifunctional scaffolds and delivery vehicles in tissue regeneration of hard and soft tissues. The construction of biomimetic microenvironments with specific chemical and physical cues for the organization and modulation of a variety of cell populations is of key importance in tissue engineering. We show that a range of human cell types, including promyoblasts, chondrocytes, adipocytes, adenovirally transduced osteoprogenitors, immunoselected mesenchymal stem cells, and the osteogenic factor, rhBMP-2 (BMP: bone morphogenic protein), can be successfully encapsulated within mineralized polysaccharide capsules without loss of function in vivo. By controlling the extent of mineralization within the alginate/chitosan shell membrane, degradation of the shell wall and release of cells or rhBMP-2 into the surrounding medium can be regulated. In addition, we describe for the first time the ability to generate bead-in-bead capsules consisting of spatially separated cell populations and temporally separated biomolecule release, entrapped within alginate/chitosan shells of variable thickness, mineralization, and stability. Such materials offer significant potential as multifunctional scaffolds and delivery vehicles in tissue regeneration of hard and soft tissues. [source]

Biomineralized Polysaccharide Capsules for Encapsulation, Organization, and Delivery of Human Cell Types and Growth Factors,

ADVANCED FUNCTIONAL MATERIALS, Issue 6 2005
W. Green
Abstract The construction of biomimetic microenvironments with specific chemical and physical cues for the organization and modulation of a variety of cell populations is of key importance in tissue engineering. We show that a range of human cell types, including promyoblasts, chondrocytes, adipocytes, adenovirally transduced osteoprogenitors, immunoselected mesenchymal stem cells, and the osteogenic factor, rhBMP-2 (BMP: bone morphogenic protein), can be successfully encapsulated within mineralized polysaccharide capsules without loss of function in vivo. By controlling the extent of mineralization within the alginate/chitosan shell membrane, degradation of the shell wall and release of cells or rhBMP-2 into the surrounding medium can be regulated. In addition, we describe for the first time the ability to generate bead-in-bead capsules consisting of spatially separated cell populations and temporally separated biomolecule release, entrapped within alginate/chitosan shells of variable thickness, mineralization, and stability. Such materials offer significant potential as multifunctional scaffolds and delivery vehicles in tissue regeneration of hard and soft tissues. [source]

Cell Types Obtained from the Epidural Space of Patients with Low Back Pain/Radiculopathy

PAIN PRACTICE, Issue 3 2009
James E. Heavner PhD
Abstract Background: We investigated if correlations exist between medical history, tissue abnormalities, and cell types retrieved from the epidural space of patients with chronic low back pain (LBP) and chronic radicular pain (RP). Methods: Approval was obtained from the Institutional Review Board for the Protection of Human Subjects to study 191 patients undergoing epiduroscopy. Visual inspection was performed and abnormal areas were identified. A specimen obtained from the area using a cytology brush was processed by the Thin Prep technique. Patients were divided into four groups based on the presence or absence and intensity of LBP and RP. The gender and age of the patients were recorded, as was any history of prior back surgery. Areas of tissue abnormalities were rated according to changes in vascularity and amount of fat, fibrosis, and inflammation. Stenosis was assessed from magnetic resonance imaging or computerized tomography scan images. Cytologic assessments included notations of the presence or absence of erythrocytes, leukocytes, cell groups, lipocytes, spindled cells, and large round cells. Results: There was a significant difference in the number of patients from whom big round cells were obtained who had a high degree of LBP compared with the number of patients who had a high degree of both LBP and RP. Conclusions: The findings provide a foundation for future studies of cells obtained from similar patients with the goal of furthering the understanding of the pathogenesis of LBP/RP. [source]

Defining Cytochemical Markers for Different Cell Types in the Equine Retina

ANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 6 2006
C. A. Deeg
Summary The major cell types in the mammalian retina are photoreceptors, amacrine, horizontal, bipolar, ganglion and Mueller glial cells. Most of the specific cell types are conserved, but cytochemical markers vary between species. The aim of our study was to characterize cytochemically distinctive markers for different cell types in the equine retina. We were able to define specific markers for equine Mueller glial cells and photoreceptor cells. Furthermore, we describe markers for large ganglion cells, horizontal and amacrine cells and a subpopulation of bipolar cells. Additionally, discrimination between the inner plexiform layer and nerve fibre layer can be achieved by expression of syntaxin and neurofilament 200 respectively. [source]

Cell type,specific expression of adenomatous polyposis coli in lung development, injury, and repair

DEVELOPMENTAL DYNAMICS, Issue 8 2010
Aimin Li
Abstract Adenomatous polyposis coli (Apc) is critical for Wnt signaling and cell migration. The current study examined Apc expression during lung development, injury, and repair. Apc was first detectable in smooth muscle layers in early lung morphogenesis, and was highly expressed in ciliated and neuroendocrine cells in the advanced stages. No Apc immunoreactivity was detected in Clara or basal cells, which function as stem/progenitor cell in adult lung. In ciliated cells, Apc is associated mainly with apical cytoplasmic domain. In response to naphthalene-induced injury, Apcpositive cells underwent squamous metaplasia, accompanied by changes in Apc subcellular distribution. In conclusion, both spatial and temporal expression of Apc is dynamically regulated during lung development and injury repair. Differential expression of Apc in progenitor vs. nonprogenitor cells suggests a functional role in cell-type specification. Subcellular localization changes of Apc in response to naphthalene injury suggest a role in cell shape and cell migration. Developmental Dynamics 239:2288,2297, 2010. © 2010 Wiley-Liss, Inc. [source]

Cell type- and region-specific expression of protein kinase C-substrate mRNAs in the cerebellum of the macaque monkey

THE JOURNAL OF COMPARATIVE NEUROLOGY, Issue 2 2003
Noriyuki Higo
Abstract We performed nonradioactive in situ hybridization histochemistry in the monkey cerebellum to investigate the localization of protein kinase C-substrate (growth-associated protein-43 [GAP-43], myristoylated alanine-rich C-kinase substrate [MARCKS], and neurogranin) mRNAs. Hybridization signals for GAP-43 mRNA were observed in the molecular and granule cell layers of both infant and adult cerebellar cortices. Signals for MARCKS mRNA were observed in the molecular, Purkinje cell, and granule cell layers of both infant and adult cortices. Moreover, both GAP-43 and MARCKS mRNAs were expressed in the external granule cell layer of the infant cortex. In the adult cerebellar vermis, signals for both GAP-43 and MARCKS mRNAs were more intense in lobules I, IX, and X than in the remaining lobules. In the adult hemisphere, both mRNAs were more intense in the flocculus and the dorsal paraflocculus than in other lobules. Such lobule-specific expressions were not prominent in the infant cerebellar cortex. Signals for neurogranin, a postsynaptic substrate for protein kinase C, were weak or not detectable in any regions of either the infant or adult cerebellar cortex. The prominent signals for MARCKS mRNA were observed in the deep cerebellar nuclei, but signals for both GAP-43 and neurogranin mRNAs were weak or not detectable. The prominent signals for both GAP-43 and MARCKS mRNAs were observed in the inferior olive, but signals for neurogranin were weak or not detectable. The cell type- and region-specific expression of GAP-43 and MARCKS mRNAs in the cerebellum may be related to functional specialization regarding plasticity in each type of cell and each region of the cerebellum. J. Comp. Neurol. 467:135,149, 2003. © 2003 Wiley-Liss, Inc. [source]

Sodium valproate inhibits glucose transport and exacerbates Glut1-deficiency in vitro

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2005
Hei Yi Wong
Abstract Anticonvulsant sodium valproate interferes with brain glucose metabolism. The mechanism underlying such metabolic disturbance is unclear. We tested the hypothesis that sodium valproate interferes with cellular glucose transport with a focus on Glut1 since glucose transport across the blood-brain barrier relies on this transporter. Cell types enriched with Glut1 expression including human erythrocytes, human skin fibroblasts, and rat astrocytes were used to study the effects of sodium valproate on glucose transport. Sodium valproate significantly inhibited Glut1 activity in normal and Glut1-deficient erythrocytes by 20%,30%, causing a corresponding reduction of Vmax of glucose transport. Similarly, in primary astrocytes as well as in normal and Glut1-deficient fibroblasts, sodium valproate inhibited glucose transport by 20%,40% (P,<,0.05), accompanied by an up to 60% downregulation of GLUT1 mRNA expression (P,<,0.05). In conclusion, sodium valproate inhibits glucose transport and exacerbates Glut1 deficiency in vitro. Our findings imply the importance of prudent use of sodium valproate for patients with compromised Glut1 function. J. Cell. Biochem. © 2005 Wiley-Liss, Inc. [source]

Cellular oxygen sensing, signalling and how to survive translational arrest in hypoxia

ACTA PHYSIOLOGICA, Issue 2 2009
M. Fähling
Abstract Hypoxia is a consequence of inadequate oxygen availability. At the cellular level, lowered oxygen concentration activates signal cascades including numerous receptors, ion channels, second messengers, as well as several protein kinases and phosphatases. This, in turn, activates trans -factors like transcription factors, RNA-binding proteins and miRNAs, mediating an alteration in gene expression control. Each cell type has its unique constellation of oxygen sensors, couplers and effectors that determine the activation and predominance of several independent hypoxia-sensitive pathways. Hence, altered gene expression patterns in hypoxia result from a complex regulatory network with multiple divergences and convergences. Although hundreds of genes are activated by transcriptional control in hypoxia, metabolic rate depression, as a consequence of reduced ATP level, causes inhibition of mRNA translation. In a multi-phase response to hypoxia, global protein synthesis is suppressed, mainly by phosphorylation of eIF2-alpha by PERK and inhibition of mTOR, causing suppression of 5,-cap-dependent mRNA translation. Growing evidence suggests that mRNAs undergo sorting at stress granules, which determines the fate of mRNA as to whether being translated, stored, or degraded. Data indicate that translation is suppressed only at ,free' polysomes, but is active at subsets of membrane-bound ribosomes. The recruitment of specific mRNAs into subcellular compartments seems to be crucial for local mRNA translation in prolonged hypoxia. Furthermore, ribosomes themselves may play a significant role in targeting mRNAs for translation. This review summarizes the multiple facets of the cellular adaptation to hypoxia observed in mammals. [source]

Activation of PLA2 isoforms by cell swelling and ischaemia/hypoxia

ACTA PHYSIOLOGICA, Issue 1-2 2006
I. H. Lambert
Abstract Phospholipase A2 (PLA2) activity is increased in mammalian cells in response to numerous stimuli such as osmotic challenge, oxidative stress and exposure to allergens. The increased PLA2 activity is seen as an increased release of free, polyunsaturated fatty acids, e.g. arachidonic acid and membrane-bound lysophospholipids. Even though arachidonic acid acts as a second messenger in its own most mammalian cells seem to rely on oxidation of the fatty acid into highly potent second messengers via, e.g. cytochrome P450, the cyclo-oxygenase, or the lipoxygenase systems for downstream signalling. Here, we review data that illustrates that stress-induced PLA2 activity involves various PLA2 subtypes and that the PLA2 in question is determined by the cell type and the physiological stress condition. [source]

Cardiac hypertrophy and failure: lessons learned from genetically engineered mice

ACTA PHYSIOLOGICA, Issue 1 2001
Y. Takeishi
Congestive heart failure is a major and growing public health problem. Because of improved survival of myocardial infarction patients produced by thrombolytic therapy or per-cutaneous revascularization it represents the only form of cardiovascular disease with significantly increased incidence and prevalence. Clinicians view this clinical syndrome as the final common pathway of diverse pathologies such as myocardial infarction and haemodynamic overload. Insights into mechanisms for heart failure historically derived from physiological and biochemical studies which identified compensatory adaptations for the haemodynamic burden associated with the pathological condition including utilization of the Frank Starling mechanism, augmentation of muscle mass, and neurohormonal activation to increase contractility. Therapy has largely been phenomenological and designed to prevent or limit the deleterious effects of these compensatory processes. More recently insights from molecular and cell biology have contributed to a more mechanistic understanding of potential causes of cardiac hypertrophy and failure. Many different analytical approaches have been employed for this purpose. These include the use of conventional animal models which permit serial observation of the onset and progression of heart failure and a sequential analysis of underlying biochemical and molecular events. Neonatal murine cardiomyocytes have been a powerful tool to examine in vitro subcellular mechanisms devoid of the confounding functional effects of multicellular preparations and heterogeneity of cell type. Finally, significant progress has been made by utilizing tissue from human cardiomyopathic hearts explanted at the time of orthotopic transplantation. Each of these methods has significant advantages and disadvantages. Arguably the greatest advance in our understanding of cardiac hypertrophy and failure over the past decade has been the exploitation of genetically engineered mice as biological reagents to study in vivo the effects of alterations in the murine genome. The power of this approach, in principle, derives from the ability to precisely overexpress or ablate a gene of interest and examine the phenotypic consequences in a cardiac specific post-natal manner. In contrast to conventional animal models of human disease which employ some form of environmental stress, genetic engineering involves a signal known molecular perturbation which produces the phenotype. [source]

Cell distribution of stress fibres in response to the geometry of the adhesive environment

CYTOSKELETON, Issue 6 2006
Manuel Théry
Abstract Cells display a large variety of shapes when plated in classical culture conditions despite their belonging to a common cell type. These shapes are transitory, since cells permanently disassemble and reassemble their cytoskeleton while moving. Adhesive micropatterns are commonly used to confine cell shape within a given geometry. In addition the micropattern can be designed so as to impose cells to spread upon adhesive and nonadhesive areas. Modulation of the pattern geometry allows the analysis of the mechanisms governing the determination of cell shape in response to external adhesive conditions. In this study, we show that the acquisition of cell shape follows two stages where initially the cell forms contact with the micropattern. Here, the most distal contacts made by the cell with the micropattern define the apices of the cell shape. Then secondly, the cell borders that link two apices move so as to minimise the distance between the two apices. In these cell borders, the absence of an underlying adhesive substrate is overcome by stress fibres forming between the apices, which in turn are marked by an accumulation of focal adhesions. By inhibiting myosin function, cell borders on nonadhesive zones become more concave, suggesting that the stress fibres work against the membrane tension in the cell border. Moreover, this suggested that traction forces are unevenly distributed in stationary, nonmigrating, cells. By comparing the stress fibres in cells with one, two, or three nonadherent cell borders it was reasoned that stress fibre strength is inversely proportional to number. We conclude that cells of a given area can generate the same total sum of tractional forces but that these tractional forces are differently spaced depending on the spatial distribution of its adherence contacts. Cell Motil. Cytoskeleton 2006. © 2006 Wiley-Liss, Inc. [source]

Anterior,posterior patterning of neural differentiated embryonic stem cells by canonical Wnts, Fgfs, Bmp4 and their respective antagonists

DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 8 2009
Marijke Hendrickx
Embryonic stem (ES) cells are pluripotent and can differentiate into every cell type of the body. Next to their potential in regenerative medicine, they are excellent tools to study embryonic development. In this work the processes of neural induction and neural patterning along the antero-posterior (A/P) body axis are studied and evidence suggests a two step mechanism for these events. First, neural induction occurs by default in the primitive ectoderm, forming anterior neural tissue and thereafter, a series of factors can posteriorize this anterior neurectoderm. In a gain-of-function/loss-of-function approach using mouse ES cells, we show that Fgf2 has the strongest caudalizing potential of all Fgfs tested. Furthermore, Bmp4 and Wnt3a, but not Wnt1, can caudalize the neurectodermal cells. The effect of the antagonists of these factors was also examined and though Dkk1 and Noggin clearly have an effect that opposes that of Wnt3a and Bmp4 respectively, they fail to anteriorize the neurectoderm. The patterning effect of SU5402, an Fgf receptor inhibitor, was rather limited. These data confirm that in the mouse, two steps are involved in neural patterning and we show that while Fgf4, Fgf8 and Wnt1 have no strong patterning effect, Fgf2, Wnt3a and Bmp4 are strong posteriorizing factors. [source]

Functional retinoid receptors in budding ascidians

DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2000
Mika Kamimura
A homolog of retinoid X receptors (RXR), named PmRXR, was cloned from the budding ascidian, Polyandrocarpa misakiensis. Gel-shift assays revealed that PmRXR and a previously identified P. misakiensis retinoic acid receptor (PmRAR) formed a complex to bind vertebrate-type retinoic acid response element (RARE). Transfection assays were carried out using a reporter gene containing a RARE upstream of lacZ. Two chimeric effector genes were constructed by placing PmRXR and PmRAR cDNA fragments (containing the DNA-binding, ligand-binding and ligand-dependent transactivation domains) downstream of the human RXR, and RAR, cDNA (covering the N-terminal coding region), respectively. Each chimeric cDNA was ligated to a notochord-specific enhancer. In case the embryos were transfected with all three transgenes and treated with retinoic acid (RA), the reporter gene was activated in the notochord cells. The result suggests that the PmRXR/PmRAR complex functions as an RA-dependent transcriptional activator. The PmRXR mRNA was detected in a mesenchymal cell type, called glomerulocyte, in the developing Polyandrocarpa bud. As this cell type has been shown to express PmRAR mRNA, it seems possible that the PmRXR/PmRAR complex mediates RA signaling in this cell type to induce the expression of genes involved in the morphogenesis of the developing bud. [source]

Developmental expression of Smoc1 and Smoc2 suggests potential roles in fetal gonad and reproductive tract differentiation

DEVELOPMENTAL DYNAMICS, Issue 11 2009
Dorothy E. Pazin
Abstract SMOC1 and SMOC2 are matricellular proteins thought to influence growth factor signaling, migration, proliferation, and angiogenesis. We examined the expression and regulation of Smoc1 and Smoc2 in fetal gonad/mesonephros complexes to discover possible roles for these genes in gonad and mesonephros development. Smoc1 was upregulated at ,E10.75 in a center-to-poles wave in pre-Sertoli and pre-granulosa cells and its expression was greatly reduced in Wt1, Sf1, and Fog2 mutants. After E13.5, Smoc1 was downregulated in an anterior-to-posterior wave in granulosa cells but persisted in Sertoli cells, suggesting a sexually dimorphic requirement in supporting cell lineage differentiation. Smoc2 was expressed in Leydig cells, mesonephroi, and Wnt4 mutant ovaries, but not wildtype ovaries. Using organ culture, we determined that Smoc2 expression was dependent on Hedgehog signaling in testes, mesonephroi, and kidneys. Overall, these results demonstrate that SMOC1 and SMOC2 may mediate intercellular signaling and cell type,specific differentiation during gonad and reproductive tract development. Developmental Dynamics 238:2877,2890, 2009. © 2009 Wiley-Liss, Inc. [source]

Diverse expression patterns of LIM-homeodomain transcription factors (LIM-HDs) in mammalian inner ear development

DEVELOPMENTAL DYNAMICS, Issue 11 2008
Mingqian Huang
Abstract LIM-homeodomain transcription factors (LIM-HDs) are essential in tissue patterning and differentiation. But their expression patterns in the inner ear are largely unknown. Here we report on a study of twelve LIM-HDs, by their tempo-spatial patterns that imply distinct yet overlapping roles, in the developing mouse inner ear. Expression of Lmx1a and Isl1 begins in the otocyst stage, with Lmx1a exclusively in the non-sensory and Isl1 in the prosensory epithelia. The second wave of expression at E12.5 includes Lhx3, 5, 9, Isl2, and Lmx1b in the differentiating sensory epithelia with cellular specificities. With the exception of Lmx1a and Lhx3, all LIM-HDs are expressed in ganglion neurons. Expression of multiple LIM-HDs within a cell type suggests their redundant function. Developmental Dynamics 237:3305,3312, 2008. © 2008 Wiley-Liss, Inc. [source]

ghrelin is a novel target of Pax4 in endocrine progenitors of the pancreas and duodenum

DEVELOPMENTAL DYNAMICS, Issue 1 2008
Qian Wang
Abstract Pax4 -deficient mice have a severe gastrointestinal endocrine deficiency: they lack most pancreatic cells that produce insulin or somatostatin and various duodenal endocrine cell types. Remarkably, Pax4 -deficient mice also have an overabundance of ghrelin-expressing cells in the pancreas and duodenum. Detailed analysis of the Pax4 nullizygous pancreas determined that the mutant islets are largely composed of a distinctive endocrine cell type that expresses ghrelin, glucagon, islet amyloid polypeptide (IAPP), and low levels of Pdx1. Lineage-tracing analysis revealed that most of these unique endocrine cells directly arose from Pax4 -deficient progenitors. Previous in vitro work reported that Pax4 is a transcriptional repressor of islet amyloid polypeptide (IAPP) and glucagon. In this study, we expanded those results by showing that Pax4 is also a repressor of gherlin. Together, our data further support the notion that Pax4 activity is necessary to establish appropriate patterns of gene expression in endocrine progenitors of the digestive tract. Developmental Dynamics 237:51,61, 2008. © 2007 Wiley-Liss, Inc. [source]

Transdifferentiation in developmental biology, disease, and in therapy

DEVELOPMENTAL DYNAMICS, Issue 12 2007
Shifaan Thowfeequ
Abstract Transdifferentiation (or metaplasia) refers to the conversion of one cell type to another. Because transdifferentiation normally occurs between cells that arise from the same region of the embryo, understanding the molecular and cellular events in cell type transformations may help to explain the mechanisms underlying normal development. Here we review examples of transdifferentiation in nature focusing on the possible role of cell type switching in metamorphosis and regeneration. We also examine transdifferentiation in mammals in relation to disease and the use of transdifferentiated cells in cellular therapy. Developmental Dynamics 236:3208,3217, 2007. © 2007 Wiley-Liss, Inc. [source]

Gonadotropin-releasing hormone-1 (GnRH-1) is involved in tooth maturation and biomineralization,

DEVELOPMENTAL DYNAMICS, Issue 11 2007
Jean Tiong
Abstract Gonadotropin releasing-hormone-1 (GnRH-1) is expressed in mouse incisors during development. In this report, we identify (1) cell type(s) that express GnRH-1 throughout tooth development, (2) the GnRH-1 receptor, and (3) the role of GnRH-1/GnRH-1 receptor signaling in tooth maturation. Results show that GnRH-1-positive cells in dental epithelium differentiate and populate multiple tooth structures including ameloblast and papillary layers that are involved in enamel formation and mineralization. The GnRH-1 receptor was present, and in vitro a GnRH-1 antagonist attenuated incisor GnRH-1 cell expression. In vivo, in mice lacking GnRH-1 (,/,), the incisors were discolored, longer, and more curved compared to wildtype. Elemental analysis of calcium, phosphorus, and iron revealed changes in ,/, incisors consistent with GnRH-1 affecting movement of minerals into the dental matrix. In sum, in tooth development a signal transduction pathway exists for GnRH-1 via the GnRH-1 receptor and disruption of such signaling affects incisor growth and biomineralization. Developmental Dynamics 236:2980,2992, 2007. Published 2007 Wiley-Liss, Inc. [source]

NG2 proteoglycan is expressed exclusively by mural cells during vascular morphogenesis

DEVELOPMENTAL DYNAMICS, Issue 2 2001
Ugur Ozerdem
Abstract Immunofluorescence mapping demonstrates that the NG2 proteoglycan is invariably expressed by the mural cell component of mouse neovascular structures. This pattern is independent of the developmental mechanism responsible for formation of the vasculature (vasculogenesis or angiogenesis). Thus, NG2 is expressed in the embryonic heart by cardiomyocytes, in developing macrovasculature by smooth muscle cells, and in nascent microvessels by vascular pericytes. Due to the scarcity of proven markers for developing pericytes, NG2 is especially useful for identification of this cell type. The utility of NG2 as a pericyte marker is illustrated by two observations. First, pericytes are associated with endothelial tubes at an early point in microvessel development. This early interaction between pericytes and endothelial cells has important implications for the role of pericytes in the development and stabilization of microvascular tubes. Second, the pericyte to endothelial cell ratio in developing capillaries varies from tissue to tissue. Because the extent of pericyte investment is likely to affect the physical properties of the vessel in question, it is important to understand the mechanisms that control this process. Additional insight into these and other aspects of vascular morphogenesis should be possible through use of NG2 as a mural cell marker. © 2001 Wiley-Liss, Inc. [source]

MafA transcription factor identifies the early ret-expressing sensory neurons

DEVELOPMENTAL NEUROBIOLOGY, Issue 7 2010
Laure Lecoin
Abstract Dorsal root ganglia proceed from the coalescence of cell bodies of sensory neurons, which have migrated dorsoventrally from the delaminating neural crest. They are composed of different neuronal subtypes with specific sensory functions, including nociception, thermal sensation, proprioception, and mechanosensation. In contrast to proprioceptors and thermonociceptors, little is known about the molecular mechanisms governing the early commitment and later differentiation into mechanosensitive neurons. This is mainly due to the absence of specific molecular markers for this particular cell type. Using knockout mice, we identified the bZIP transcription factor MafA as the first specific marker of a subpopulation of "early c-ret" positive neurons characterized by medium-to-large diameters. This marker will allow further functional characterization of these neurons. © 2010 Wiley Periodicals, Inc. Develop Neurobiol 70:485,497, 2010 [source]

Liposome-mediated transfection of mature taste cells

DEVELOPMENTAL NEUROBIOLOGY, Issue 1 2005
Ana Marie Landin
Abstract The introduction and expression of exogenous DNA in neurons is valuable for analyzing a range of cellular and molecular processes in the periphery, e.g., the roles of transduction-related proteins, the impact of growth factors on development and differentiation, and the function of promoters specific to cell type. However, sensory receptor cells, particularly chemosensory cells, have been difficult to transfect. We have successfully introduced plasmids expressing green and Discosoma Red fluorescent proteins (GFP and DsRed) into rat taste buds in primary culture. Transfection efficiency increased when delaminated taste epithelium was redigested with fresh protease, suggesting that a protective barrier of extracellular matrix surrounding taste cells may normally be present. Because taste buds are heterogeneous aggregates of cells, we used ,-gustducin, neuronal cell adhesion molecule (NCAM), and neuronal ubiquitin carboxyl terminal hydrolase (PGP9.5), markers for defined subsets of mature taste cells, to demonstrate that liposome-mediated transfection targets multiple taste cell types. After testing eight commercially available lipids, we identified one, Transfast, that is most effective on taste cells. We also demonstrate the effectiveness of two common "promiscuous" promoters and one promoter that taste cells use endogenously. These studies should permit ex vivo strategies for studying development and cellular function in taste cells. © 2005 Wiley Periodicals, Inc. J. Neurobiol, 2005 [source]

Medullary thyroid carcinoma presenting as rectangular cell type on fine-needle aspiration

DIAGNOSTIC CYTOPATHOLOGY, Issue 3 2009
Andrew M. Schreiner M.D.
Abstract Medullary thyroid carcinoma typically presents as dyscohesive plasmacytoid, spindled, or polygonal cells on fine-needle aspiration smears. We recently encountered a case of sporadic medullary thyroid carcinoma that presented as a hypercellular aspirate composed of cohesive aggregates of rectangle-shaped cells. The case was mistakenly reported as a hypercellular follicular neoplasm on cytology. Subsequent thyroidectomy revealed medullary carcinoma. We draw attention to this distinctive rectangular cell type as an additional morphology for medullary thyroid carcinoma. Diagn. Cytopathol. 2009. © 2009 Wiley-Liss, Inc. [source]

Cytomorphology of anaplastic giant cell type of medullary thyroid carcinoma,A diagnostic dilemma in an elderly female: A case report

DIAGNOSTIC CYTOPATHOLOGY, Issue 2 2008
Bharat Rekhi M.D., D.N.B, M.I.A.C
No abstract is available for this article. [source]

Detection of a subset of CD30+ anaplastic large cell lymphoma by interphase fluorescence in situ hybridization

DIAGNOSTIC CYTOPATHOLOGY, Issue 2 2003
Hyung Ju C. Shin M.D.
Abstract T/null-cell anaplastic large cell lymphoma (ALCL) is a morphologically and clinically heterogeneous group of non-Hodgkin's lymphoma; to date several morphologic variants have been described on histologic specimens. However, the cytologic features of these variants in the fine-needle aspiration (FNA) specimens have not been well evaluated. The t(2;5)(p23;q35) has been identified in a subset of T/null-ALCL and is known to be associated with a favorable prognosis. We reviewed the cytomorphologic characteristics in 24 FNA specimens of ALCL. In all cases, the diagnosis was confirmed on histologic specimens, and immunohistochemical studies for anaplastic lymphoma kinase (ALK) protein expression were performed on the aspirates. The presence of ALK breakpoints were evaluated in nine cases, using a DNA break-apart probe on chromosome 2 covering the ALK gene by fluorescence in situ hybridization (FISH) techniques. Two hundred cells per case were examined. The results were expressed as the percentage of cells containing more than two signals of chromosome 2 to the total number of cells counted. FNA sites included lymph nodes (20), lung (2), breast (1), and soft tissue (1). The median age of the patients was 56 yr (range, 17,75 yr). Twenty cases had systemic involvement; in four cases, skin was the primary site with secondary involvement of the lymph nodes. All cases were CD30+ by immunohistochemistry; 20 were of T-cell phenotype and 4 were null cell type. The cytologic evaluation revealed typical anaplastic morphology (common type) with many "hallmark cells" in 16 (67%) cases. Other morphologic variants identified were small cell pattern in five cases, monomorphic pattern in two cases, and lymphohistiocytic pattern in one case. FISH studies showed that six (66.7%) of nine cases had at least two signals of chromosome 2, consistent with ALK breakpoints. With careful cytomorphologic evaluation in conjunction with appropriate immunohistochemical studies, a diagnosis of ALCL can be confidently made in the FNA specimens in the cellular aspirates and its morphologic variants also can be recognized. Furthermore, the FNA specimen is suitable in detecting ALK breakpoints by FISH study, permitting rapid identification of a subset of patients with ALCL, who may have a favorable prognosis. Using a commercially available probe, detection of ALK breakpoints in the FNA specimens is simple and can be a useful diagnostic adjunct in cases where distinction from other lymphomas or lymphoid lesions is morphologically difficult. Diagn. Cytopathol. 2003;29:61,66. © 2003 Wiley-Liss, Inc. [source]

Diagnosis of melanoma aspirates on ThinPrep®: The University of Michigan experience

DIAGNOSTIC CYTOPATHOLOGY, Issue 5 2002
Güliz Akdas Barkan M.D.
Abstract The purpose of this study was to compare the cytologic features of melanoma fine-needle aspirates (FNAs) prepared by ThinPrep® (TP) with those in conventional smears (CS) and to identify any diagnostic pitfalls. Fifty-one aspirates diagnosed as melanoma were obtained, 36 of which were prepared by both TP and CS. The preparations were evaluated for cellularity, cell aggregates, cellular appearance, melanin pigment, cytoplasmic, and nuclear features. Categorical data were analyzed by the chi-square test and continuous data by the Wilcoxin-signed rank test. Correlation was determined by Spearman's test for bivariate correlations (rho). Good correlation between the two methods was identified for the following features: cellularity, cell type, bi/multinucleated cells, cytoplasmic features, NC ratio, and presence of macronucleoli. TP exhibits coarser chromatin compared to CS (P = 0.005). Six of 36 CS contained large cellular groups; none of the TP contained them (P = 0.018). Twenty-five of 36 CS contained intranuclear inclusions as opposed to 12/36 TP (P < 0.001). The number of inclusions was significantly reduced on TP. The amount of intracellular melanin was the same with both techniques. Background melanin was markedly reduced on TP except when either trapped by fibrin or attached to cellular clusters (P = 0.006). Background blood was also markedly reduced on TP (P < 0.005). In summary, the cytological features of TP and CS for FNA evaluation of melanoma correlate well; however, one needs to be aware of the cytologic alterations introduced by TP. TP is a sufficient preparation method in the diagnosis of melanoma FNA aspirates when performed by clinicians. It is also a useful adjunct in bloody or low-cellular aspirates, where it tends to reduce the background blood and concentrate the cells. Diagn. Cytopathol. 2002;26:334,339. © 2002 Wiley-Liss, Inc. [source]

Evidence for a widespread involvement of NO in control of photogenesis in bioluminescent fish

ACTA ZOOLOGICA, Issue 4 2010
Jenny Krönström
Abstract Krönström, J. and Mallefet, J. 2009. Evidence for a widespread involvement of NO in control of photogenesis in bioluminescent fish. ,Acta Zoologica (Stockholm) 91: 474,483. The presence of nitric oxide synthase (NOS) and nerve fibres in the photophores of seven bioluminescent fish species (Hygophum benoiti, Myctophum punctatum, Electrona risso, Cyclothone braueri, Vinciguerria attenuata, Maurolicus muelleri and Porichthys notatus) with endogenous photocytes, were investigated. Antibodies directed against neuronal and inducible NOS (n and iNOS respectively) and NADPH-diaphorase activity were used to reveal the locations of NOS, while antibodies directed against acetylated tubulin were used to visualize nerve fibres. The nNOS antibody labelled structures in all investigated photophores except in the organs from P. notatus. The photocytes of P. notatus showed NADPH-diaphorase activity. In the myctophid species, NOS-like immunoreactivity was found in small intracellular structures of the photocytes and in nerve fibres reaching the photocytes. nNOS-positive fibres were also found among lens/filter cells in V. attenuata, and in M. muelleri the cytoplasm of lens/filter cells contained NOS-like material. In C. braueri, a cell type located at a collecting chamber for luminous products in the photophore contained NOS-like material. All photophores received an innervation reaching the photocytes, as well as other components including lens/filter areas. The results of this study comply with an involvement of nitric oxide in the control of bioluminescence in several fish species. [source]

The eye of the freshwater prosobranch gastropod Viviparus viviparus: ultrastructure, electrophysiology and behaviour

ACTA ZOOLOGICA, Issue 1 2006
Valery V. Zhukov
Abstract We used light and electron microscopy to study the retinal organization of the eye of Viviparus viviparus. Electroretinogram (ERG) recordings were used to investigate the electrophysiological responsiveness to flashes of light of varying intensity and colour, behavioural observations were made of phototactic reactions, and optical measurements and calculations related to the path of light rays in the eye were made. The retina contains principally two types of cells: first, photoreceptor cells with both microvilli and cilia, and second, cells, often strongly pigmented, that are supportive in nature. The ERGs obtained were essentially similar in form, amplitude and duration to those known from other gastropods that have exclusively rhabdomeric photoreceptors. Spectral sensitivity curves closely fitted the absorption spectrum of a rhodopsin-like pigment. The spectral sensitivity peak was at 475 nm. Measurements of the refractive indices of the lens gave values of 1.55 for the outer layer and 1.57 for the lens core. None of the snails tested exhibited a ,defensive reflex' and although no preference between light and dark regions was expressed, we nevertheless argue that, on the basis of optical measurements and calculations, the eye of V. viviparus is well-adapted for seeing under water. Our main conclusion is that in the eye of V. viviparus with its ,mixed photoreceptor' cell type, there is an equal probability for microvilli and cilia to function as principal photoreceptive elements. [source]

Locomotory and feeding effectors of the tornaria larva of Balanoglossus biminiensis

ACTA ZOOLOGICA, Issue 2 2001
T. C. Lacalli
Abstract Lacalli, T. C. and Gilmour, T. H. J. 2001. Locomotory and feeding effectors of the tornaria larva of Balanoglossus biminiensis. ,Acta Zoologica (Stockholm) 82: 117,126 The tornaria ciliary bands and oesophagus were examined ultrastructurally to identify the neural components that control larval behaviour. The circumoral ciliary band is known to be innervated in part by fibres from the apical plate and adoral nerve centres. Within the band itself, however, the only neurones we could find were multipolar cells, an unusual cell type with apical processes that traverse the surface of the band. Similar cells occur in the circumoral bands of echinoderm larvae. The tornaria telotroch has a much larger nerve, but no neurones were found either in the band or nearby, so the source of the fibres in the telotroch nerve remains unknown. In addition to having different innervation, the two bands also respond differently to cholinergic agonists, which elicit telotroch arrests but have no visible effect on the circumoral band. The oesophagus has a well-developed musculature and an extensive nerve plexus. During feeding, the oesophagus repeatedly contracts, forcing excess water out along two lateral channels prior to swallowing. These channels are also sites of gill slit formation, so there is evidently a continuity between the water bypass mechanism of the larva and that of the postmetamorphic juvenile. [source]

Nuclear proteome analysis of undifferentiated mouse embryonic stem and germ cells

ELECTROPHORESIS, Issue 11 2008
Nicolas Buhr
Abstract Embryonic stem cells (ESCs) and embryonic germ cells (EGCs) provide exciting models for understanding the underlying mechanisms that make a cell pluripotent. Indeed, such understanding would enable dedifferentiation and reprogrammation of any cell type from a patient needing a cell therapy treatment. Proteome analysis has emerged as an important technology for deciphering these biological processes and thereby ESC and EGC proteomes are increasingly studied. Nevertheless, their nuclear proteomes have only been poorly investigated up to now. In order to investigate signaling pathways potentially involved in pluripotency, proteomic analyses have been performed on mouse ESC and EGC nuclear proteins. Nuclei from ESCs and EGCs at undifferentiated stage were purified by subcellular fractionation. After 2-D separation, a subtractive strategy (subtracting culture environment contaminating spots) was applied and a comparison of ESC, (8.5 day post coïtum (dpc))-EGC and (11.5,dpc)-EGC specific nuclear proteomes was performed. A total of 33 ESC, 53 (8.5,dpc)-EGC, and 36 (11.5,dpc)-EGC spots were identified by MALDI-TOF-MS and/or nano-LC-MS/MS. This approach led to the identification of two isoforms (with and without N -terminal acetylation) of a known pluripotency marker, namely developmental pluripotency associated 5 (DPPA5), which has never been identified before in 2-D gel-MS studies of ESCs and EGCs. Furthermore, we demonstrated the efficiency of our subtracting strategy, in association with a nuclear subfractionation by the identification of a new protein (protein arginine N -methyltransferase 7; PRMT7) behaving as proteins involved in pluripotency. [source]