Mechanisms of splicing-dependent trans-synaptic adhesion by PTPδ-IL1RAPL1/IL-1RAcP for synaptic differentiation

NASA Astrophysics Data System (ADS)

Yamagata, Atsushi; Yoshida, Tomoyuki; Sato, Yusuke; Goto-Ito, Sakurako; Uemura, Takeshi; Maeda, Asami; Shiroshima, Tomoko; Iwasawa-Okamoto, Shiho; Mori, Hisashi; Mishina, Masayoshi; Fukai, Shuya

2015-04-01

Synapse formation is triggered through trans-synaptic interaction between pairs of pre- and postsynaptic adhesion molecules, the specificity of which depends on splice inserts known as `splice-insert signaling codes'. Receptor protein tyrosine phosphatase δ (PTPδ) can bidirectionally induce pre- and postsynaptic differentiation of neurons by trans-synaptically binding to interleukin-1 receptor accessory protein (IL-1RAcP) and IL-1RAcP-like-1 (IL1RAPL1) in a splicing-dependent manner. Here, we report crystal structures of PTPδ in complex with IL1RAPL1 and IL-1RAcP. The first immunoglobulin-like (Ig) domain of IL1RAPL1 directly recognizes the first splice insert, which is critical for binding to IL1RAPL1. The second splice insert functions as an adjustable linker that positions the Ig2 and Ig3 domains of PTPδ for simultaneously interacting with the Ig1 domain of IL1RAPL1 or IL-1RAcP. We further identified the IL1RAPL1-specific interaction, which appears coupled to the first-splice-insert-mediated interaction. Our results thus reveal the decoding mechanism of splice-insert signaling codes for synaptic differentiation induced by trans-synaptic adhesion between PTPδ and IL1RAPL1/IL-1RAcP.

Reduced inhibitory gate in the barrel cortex of Neuroligin3R451C knock‐in mice, an animal model of autism spectrum disorders

PubMed Central

Cellot, Giada; Cherubini, Enrico

2014-01-01

Abstract Neuroligins are postsynaptic adhesion molecules that interacting with presynaptic neurexins ensure the cross‐talk between pre‐ and postsynaptic specializations. Rare mutations in neurexin–neuroligin genes have been linked to autism spectrum disorders (ASDs). One of these, the R451C mutation of the gene encoding for Neuroligin3 (Nlgn3), has been found in patients with familial forms of ASDs. Animals carrying this mutation (NL3R451C knock‐in mice) exhibit impaired social behaviors, reminiscent of those observed in ASD patients, associated with major alterations in both GABAergic and glutamatergic transmission, which vary among different brain regions and at different developmental stages. Here, pair recordings from parvalbumin‐ (PV) expressing basket cells and spiny neurons were used to study GABAergic synaptic signaling in layer IV barrel cortex of NL3R451C mutant mice. We found that the R451C mutation severely affects the probability of GABA release from PV‐expressing basket cells, responsible for controlling via thalamo‐cortical inputs the feed‐forward inhibition. No changes in excitatory inputs to parvalbumin‐positive basket cells or spiny neurons were detected. These data clearly show that primary targets of the NL3 mutation are PV‐expressing basket cells, independently of the brain region where they are localized. Changes in the inhibitory gate of layer IV somatosensory cortex may alter sensory processing in ASD patients leading to misleading sensory representations with difficulties to combine pieces of information into a unified perceptual whole. PMID:25347860

Cationic influences upon synaptic transmission at the hair cell-afferent fiber synapse of the frog

NASA Technical Reports Server (NTRS)

Cochran, S. L.

1995-01-01

The concentrations of inorganic cations (K+, Na+, and Ca2+) bathing the isolated frog labyrinth were varied in order to assess their role in influencing and mediating synaptic transmission at the hair cell-afferent fiber synapse. Experiments employed intracellular recordings of synaptic activity from VIIIth nerve afferents. Recordings were digitized continuously at 50 kHz, and excitatory postsynaptic potentials were detected and parameters quantified by computer algorithms. Particular attention was focused on cationic effects upon excitatory postsynaptic potential frequency of occurrence and excitatory postsynaptic potential amplitude, in order to discriminate between pre- and postsynaptic actions. Because the small size of afferents preclude long term stable recordings, alterations in cationic concentrations were applied transiently and their peak effects on synaptic activity were assessed. Increases in extracellular K+ concentration of a few millimolar produced a large increase in the frequency of occurrence of excitatory postsynaptic potentials with little change in amplitude, indicating that release of transmitter from the hair cell is tightly coupled to its membrane potential. Increasing extracellular Na+ concentration resulted in an increase in excitatory postsynaptic potential amplitude with no significant change in excitatory postsynaptic potential frequency of occurrence, suggesting that the transmitter-gated subsynaptic channel conducts Na+ ions. Decreases in extracellular Ca2+ concentration had little effect upon excitatory postsynaptic potential frequency, but increased excitatory postsynaptic potential frequency and amplitude. These findings suggest that at higher concentrations Ca2+ act presynaptically to prevent transmitter release and postsynaptically to prevent Na+ influx during the generation of the excitatory postsynaptic potential. The influences of these ions on synaptic activity at this synapse are remarkably similar to those reported at the vertebrate neuromuscular junction. The major differences between these two synapses are the neurotransmitters and the higher resting release rate and higher sensitivity of release to increased K+ concentrations of the hair cells over that of motor nerve terminals. These differences reflect the functional roles of the two synapses: the motor nerve terminal response in an all-or-nothing signal consequent from action potential invasion, while the hair cell releases transmitter in a graded fashion, proportionate to the extent of stereocilial deflection. Despite these differences between the two junctions, the similar actions of these elemental cations upon synaptic function at each implies that these ions may participate similarly in the operations of other synapses, independent of the neurotransmitter type.(ABSTRACT TRUNCATED AT 400 WORDS).

Adducin at the Neuromuscular Junction in Amyotrophic Lateral Sclerosis: Hanging on for Dear Life

PubMed Central

Krieger, Charles; Wang, Simon Ji Hau; Yoo, Soo Hyun; Harden, Nicholas

2016-01-01

The neurological dysfunction in amyotrophic lateral sclerosis (ALS)/motor neurone disease (MND) is associated with defective nerve-muscle contacts early in the disease suggesting that perturbations of cell adhesion molecules (CAMs) linking the pre- and post-synaptic components of the neuromuscular junction (NMJ) are involved. To search for candidate proteins implicated in this degenerative process, researchers have studied the Drosophila larval NMJ and find that the cytoskeleton-associated protein, adducin, is ideally placed to regulate synaptic contacts. By controlling the levels of synaptic proteins, adducin can de-stabilize synaptic contacts. Interestingly, elevated levels of phosphorylated adducin have been reported in ALS patients and in a mouse model of the disease. Adducin is regulated by phosphorylation through protein kinase C (PKC), some isoforms of which exhibit Ca2+-dependence, raising the possibility that changes in intracellular Ca2+ might alter PKC activation and secondarily influence adducin phosphorylation. Furthermore, adducin has interactions with the alpha subunit of the Na+/K+-ATPase. Thus, the phosphorylation of adducin may secondarily influence synaptic stability at the NMJ and so influence pre- and post-synaptic integrity at the NMJ in ALS. PMID:26858605

NGL-2 Deletion Leads to Autistic-like Behaviors Responsive to NMDAR Modulation.

PubMed

Um, Seung Min; Ha, Seungmin; Lee, Hyejin; Kim, Jihye; Kim, Kyungdeok; Shin, Wangyong; Cho, Yi Sul; Roh, Junyeop Daniel; Kang, Jaeseung; Yoo, Taesun; Noh, Young Woo; Choi, Yeonsoo; Bae, Yong Chul; Kim, Eunjoon

2018-06-26

Netrin-G ligand 2 (NGL-2)/LRRC4, implicated in autism spectrum disorders and schizophrenia, is a leucine-rich repeat-containing postsynaptic adhesion molecule that interacts intracellularly with the excitatory postsynaptic scaffolding protein PSD-95 and trans-synaptically with the presynaptic adhesion molecule netrin-G2. Functionally, NGL-2 regulates excitatory synapse development and synaptic transmission. However, whether it regulates synaptic plasticity and disease-related specific behaviors is not known. Here, we report that mice lacking NGL-2 (Lrrc4 -/- mice) show suppressed N-Methyl-D-aspartate receptor (NMDAR)-dependent synaptic plasticity in the hippocampus. NGL-2 associates with NMDARs through both PSD-95-dependent and -independent mechanisms. Moreover, Lrrc4 -/- mice display mild social interaction deficits and repetitive behaviors that are rapidly improved by pharmacological NMDAR activation. These results suggest that NGL-2 promotes synaptic stabilization of NMDARs, regulates NMDAR-dependent synaptic plasticity, and prevents autistic-like behaviors from developing in mice, supporting the hypothesis that NMDAR dysfunction contributes to autism spectrum disorders. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Functional dependence of neuroligin on a new non-PDZ intracellular domain

PubMed Central

Shipman, Seth L; Schnell, Eric; Hirai, Takaaki; Chen, Bo-Shiun; Roche, Katherine W; Nicoll, Roger A

2011-01-01

Neuroligins, a family of postsynaptic adhesion molecules, are important in synaptogenesis through a well-characterized trans-synaptic interaction with neurexin. In addition, neuroligins are thought to drive postsynaptic assembly through binding of their intracellular domain to PSD-95. However, there is little direct evidence to support the functional necessity of the neuroligin intracellular domain in postsynaptic development. We found that presence of endogenous neuroligin obscured the study of exogenous mutated neuroligin. We therefore used chained microRNAs in rat organotypic hippocampal slices to generate a reduced background of endogenous neuroligin. On this reduced background, we found that neuroligin function was critically dependent on the cytoplasmic tail. However, this function required neither the PDZ ligand nor any other previously described cytoplasmic binding domain, but rather required a previously unknown conserved region. Mutation of a single critical residue in this region inhibited neuroligin-mediated excitatory synaptic potentiation. Finally, we found a functional distinction between neuroligins 1 and 3. PMID:21532576

Emerging Synaptic Molecules as Candidates in the Etiology of Neurological Disorders

PubMed Central

Torres, Viviana I.; Vallejo, Daniela

2017-01-01

Synapses are complex structures that allow communication between neurons in the central nervous system. Studies conducted in vertebrate and invertebrate models have contributed to the knowledge of the function of synaptic proteins. The functional synapse requires numerous protein complexes with specialized functions that are regulated in space and time to allow synaptic plasticity. However, their interplay during neuronal development, learning, and memory is poorly understood. Accumulating evidence links synapse proteins to neurodevelopmental, neuropsychiatric, and neurodegenerative diseases. In this review, we describe the way in which several proteins that participate in cell adhesion, scaffolding, exocytosis, and neurotransmitter reception from presynaptic and postsynaptic compartments, mainly from excitatory synapses, have been associated with several synaptopathies, and we relate their functions to the disease phenotype. PMID:28331639

Evidence that protons act as neurotransmitters at vestibular hair cell-calyx afferent synapses.

PubMed

Highstein, Stephen M; Holstein, Gay R; Mann, Mary Anne; Rabbitt, Richard D

2014-04-08

Present data support the conclusion that protons serve as an important neurotransmitter to convey excitatory stimuli from inner ear type I vestibular hair cells to postsynaptic calyx nerve terminals. Time-resolved pH imaging revealed stimulus-evoked extrusion of protons from hair cells and a subsequent buildup of [H(+)] within the confined chalice-shaped synaptic cleft (ΔpH ∼ -0.2). Whole-cell voltage-clamp recordings revealed a concomitant nonquantal excitatory postsynaptic current in the calyx terminal that was causally modulated by cleft acidification. The time course of [H(+)] buildup limits the speed of this intercellular signaling mechanism, but for tonic signals such as gravity, protonergic transmission offers a significant metabolic advantage over quantal excitatory postsynaptic currents--an advantage that may have driven the proliferation of postsynaptic calyx terminals in the inner ear vestibular organs of contemporary amniotes.

Increased Excitatory Synaptic Transmission of Dentate Granule Neurons in Mice Lacking PSD-95-Interacting Adhesion Molecule Neph2/Kirrel3 during the Early Postnatal Period.

PubMed

Roh, Junyeop D; Choi, Su-Yeon; Cho, Yi Sul; Choi, Tae-Yong; Park, Jong-Sil; Cutforth, Tyler; Chung, Woosuk; Park, Hanwool; Lee, Dongsoo; Kim, Myeong-Heui; Lee, Yeunkum; Mo, Seojung; Rhee, Jeong-Seop; Kim, Hyun; Ko, Jaewon; Choi, Se-Young; Bae, Yong Chul; Shen, Kang; Kim, Eunjoon; Han, Kihoon

2017-01-01

Copy number variants and point mutations of NEPH2 (also called KIRREL3 ) gene encoding an immunoglobulin (Ig) superfamily adhesion molecule have been linked to autism spectrum disorders, intellectual disability and neurocognitive delay associated with Jacobsen syndrome, but the physiological roles of Neph2 in the mammalian brain remain largely unknown. Neph2 is highly expressed in the dentate granule (DG) neurons of the hippocampus and is localized in both dendrites and axons. It was recently shown that Neph2 is required for the formation of mossy fiber filopodia, the axon terminal structure of DG neurons forming synapses with GABAergic neurons of CA3. In contrast, however, it is unknown whether Neph2 also has any roles in the postsynaptic compartments of DG neurons. We here report that, through its C-terminal PDZ domain-binding motif, Neph2 directly interacts with postsynaptic density (PSD)-95, an abundant excitatory postsynaptic scaffolding protein. Moreover, Neph2 protein is detected in the brain PSD fraction and interacts with PSD-95 in synaptosomal lysates. Functionally, loss of Neph2 in mice leads to age-specific defects in the synaptic connectivity of DG neurons. Specifically, Neph2 -/- mice show significantly increased spontaneous excitatory synaptic events in DG neurons at postnatal week 2 when the endogenous Neph2 protein expression peaks, but show normal excitatory synaptic transmission at postnatal week 3. The evoked excitatory synaptic transmission and synaptic plasticity of medial perforant pathway (MPP)-DG synapses are also normal in Neph2 -/- mice at postnatal week 3, further confirming the age-specific synaptic defects. Together, our results provide some evidence for the postsynaptic function of Neph2 in DG neurons during the early postnatal period, which might be implicated in neurodevelopmental and cognitive disorders caused by NEPH2 mutations.

Developmental regulation of GABAergic signalling in the hippocampus of neuroligin 3 R451C knock-in mice: an animal model of Autism.

PubMed

Pizzarelli, Rocco; Cherubini, Enrico

2013-01-01

Autism Spectrum Disorders (ASDs) comprise an heterogeneous group of neuro-developmental abnormalities, mainly of genetic origin, characterized by impaired social interactions, communications deficits, and stereotyped behaviors. In a small percentage of cases, ASDs have been found to be associated with single mutations in genes involved in synaptic function. One of these involves the postsynaptic cell adhesion molecule neuroligin (NL) 3. NLs interact with presynaptic neurexins (Nrxs) to ensure a correct cross talk between post and presynaptic specializations. Here, transgenic mice carrying the human R451C mutation of Nlgn3, were used to study GABAergic signaling in the hippocampus early in postnatal life. Whole cell recordings from CA3 pyramidal neurons in slices from NL3(R451C) knock-in mice revealed an enhanced frequency of Giant Depolarizing Potentials (GDPs), as compared to controls. This effect was probably dependent on an increased GABAergic drive to principal cells as demonstrated by the enhanced frequency of miniature GABAA-mediated (GPSCs), but not AMPA-mediated postsynaptic currents (EPSCs). Changes in frequency of mGPSCs were associated with an acceleration of their decay kinetics, in the absence of any change in unitary synaptic conductance or in the number of GABAA receptor channels, as assessed by peak scaled non-stationary fluctuation analysis. The enhanced GABAergic but not glutamatergic transmission early in postnatal life may change the excitatory/inhibitory balance known to play a key role in the construction and refinement of neuronal circuits during postnatal development. This may lead to behavioral deficits reminiscent of those observed in ASDs patients.

Developmental regulation of GABAergic signalling in the hippocampus of neuroligin 3 R451C knock-in mice: an animal model of Autism

PubMed Central

Pizzarelli, Rocco; Cherubini, Enrico

2013-01-01

Autism Spectrum Disorders (ASDs) comprise an heterogeneous group of neuro-developmental abnormalities, mainly of genetic origin, characterized by impaired social interactions, communications deficits, and stereotyped behaviors. In a small percentage of cases, ASDs have been found to be associated with single mutations in genes involved in synaptic function. One of these involves the postsynaptic cell adhesion molecule neuroligin (NL) 3. NLs interact with presynaptic neurexins (Nrxs) to ensure a correct cross talk between post and presynaptic specializations. Here, transgenic mice carrying the human R451C mutation of Nlgn3, were used to study GABAergic signaling in the hippocampus early in postnatal life. Whole cell recordings from CA3 pyramidal neurons in slices from NL3R451C knock-in mice revealed an enhanced frequency of Giant Depolarizing Potentials (GDPs), as compared to controls. This effect was probably dependent on an increased GABAergic drive to principal cells as demonstrated by the enhanced frequency of miniature GABAA-mediated (GPSCs), but not AMPA-mediated postsynaptic currents (EPSCs). Changes in frequency of mGPSCs were associated with an acceleration of their decay kinetics, in the absence of any change in unitary synaptic conductance or in the number of GABAA receptor channels, as assessed by peak scaled non-stationary fluctuation analysis. The enhanced GABAergic but not glutamatergic transmission early in postnatal life may change the excitatory/inhibitory balance known to play a key role in the construction and refinement of neuronal circuits during postnatal development. This may lead to behavioral deficits reminiscent of those observed in ASDs patients. PMID:23761734

The Glutamatergic Aspects of Schizophrenia Molecular Pathophysiology: Role of the Postsynaptic Density, and Implications for Treatment

PubMed Central

Iasevoli, Felice; Tomasetti, Carmine; Buonaguro, Elisabetta F.; de Bartolomeis, Andrea

2014-01-01

Schizophrenia is one of the most debilitating psychiatric diseases with a lifetime prevalence of approximately 1%. Although the specific molecular underpinnings of schizophrenia are still unknown, evidence has long linked its pathophysiology to postsynaptic abnormalities. The postsynaptic density (PSD) is among the molecular structures suggested to be potentially involved in schizophrenia. More specifically, the PSD is an electron-dense thickening of glutamatergic synapses, including ionotropic and metabotropic glutamate receptors, cytoskeletal and scaffolding proteins, and adhesion and signaling molecules. Being implicated in the postsynaptic signaling of multiple neurotransmitter systems, mostly dopamine and glutamate, the PSD constitutes an ideal candidate for studying dopamine-glutamate disturbances in schizophrenia. Recent evidence suggests that some PSD proteins, such as PSD-95, Shank, and Homer are implicated in severe behavioral disorders, including schizophrenia. These findings, further corroborated by genetic and animal studies of schizophrenia, offer new insights for the development of pharmacological strategies able to overcome the limitations in terms of efficacy and side effects of current schizophrenia treatment. Indeed, PSD proteins are now being considered as potential molecular targets against this devastating illness. The current paper reviews the most recent hypotheses on the molecular mechanisms underlying schizophrenia pathophysiology. First, we review glutamatergic dysfunctions in schizophrenia and we provide an update on postsynaptic molecules involvement in schizophrenia pathophysiology by addressing both human and animal studies. Finally, the possibility that PSD proteins may represent potential targets for new molecular interventions in psychosis will be discussed. PMID:24851087

Reduced inhibitory gate in the barrel cortex of Neuroligin3R451C knock-in mice, an animal model of autism spectrum disorders.

PubMed

Cellot, Giada; Cherubini, Enrico

2014-07-16

Neuroligins are postsynaptic adhesion molecules that interacting with presynaptic neurexins ensure the cross-talk between pre- and postsynaptic specializations. Rare mutations in neurexin-neuroligin genes have been linked to autism spectrum disorders (ASDs). One of these, the R451C mutation of the gene encoding for Neuroligin3 (Nlgn3), has been found in patients with familial forms of ASDs. Animals carrying this mutation (NL3(R451C) knock-in mice) exhibit impaired social behaviors, reminiscent of those observed in ASD patients, associated with major alterations in both GABAergic and glutamatergic transmission, which vary among different brain regions and at different developmental stages. Here, pair recordings from parvalbumin- (PV) expressing basket cells and spiny neurons were used to study GABAergic synaptic signaling in layer IV barrel cortex of NL3(R451C) mutant mice. We found that the R451C mutation severely affects the probability of GABA release from PV-expressing basket cells, responsible for controlling via thalamo-cortical inputs the feed-forward inhibition. No changes in excitatory inputs to parvalbumin-positive basket cells or spiny neurons were detected. These data clearly show that primary targets of the NL3 mutation are PV-expressing basket cells, independently of the brain region where they are localized. Changes in the inhibitory gate of layer IV somatosensory cortex may alter sensory processing in ASD patients leading to misleading sensory representations with difficulties to combine pieces of information into a unified perceptual whole. © 2014 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Striatal-enriched Protein-tyrosine Phosphatase (STEP) Regulates Pyk2 Kinase Activity*

PubMed Central

Xu, Jian; Kurup, Pradeep; Bartos, Jason A.; Patriarchi, Tommaso; Hell, Johannes W.; Lombroso, Paul J.

2012-01-01

Proline-rich tyrosine kinase 2 (Pyk2) is a member of the focal adhesion kinase family and is highly expressed in brain and hematopoietic cells. Pyk2 plays diverse functions in cells, including the regulation of cell adhesion, migration, and cytoskeletal reorganization. In the brain, it is involved in the induction of long term potentiation through regulation of N-methyl-d-aspartate receptor trafficking. This occurs through the phosphorylation and activation of Src family tyrosine kinase members, such as Fyn, that phosphorylate GluN2B at Tyr1472. Phosphorylation at this site leads to exocytosis of GluN1-GluN2B receptors to synaptic membranes. Pyk2 activity is modulated by phosphorylation at several critical tyrosine sites, including Tyr402. In this study, we report that Pyk2 is a substrate of striatal-enriched protein-tyrosine phosphatase (STEP). STEP binds to and dephosphorylates Pyk2 at Tyr402. STEP KO mice showed enhanced phosphorylation of Pyk2 at Tyr402 and of the Pyk2 substrates paxillin and ASAP1. Functional studies indicated that STEP opposes Pyk2 activation after KCl depolarization of cortical slices and blocks Pyk2 translocation to postsynaptic densities, a key step required for Pyk2 activation and function. This is the first study to identify Pyk2 as a substrate for STEP. PMID:22544749

Effects of fluoride on synapse morphology and myelin damage in mouse hippocampus.

PubMed

Niu, Ruiyan; Chen, Huijuan; Manthari, Ram Kumar; Sun, Zilong; Wang, Jinming; Zhang, Jianhai; Wang, Jundong

2018-03-01

To investigate the fluoride-induced neurotoxicity on mice hippocampus, healthy adult mice were exposed to 25, 50, and 100 mg NaF/L for 60 days. The results showed that medium and high fluoride administration induced ultrastructural alterations in the structure of neuron synapse, including indistinct and short synaptic cleft, and thickened postsynaptic density (PSD). The significant reduced mRNA expressions of proteolipid protein (PLP) in medium and high fluoride groups suggested that myelin damage occurred in hippocampus. The myelin damage in turn was determined by the increased myelin-associated glycoprotein (MAG) level, which is naturally released by injured myelin, in high fluoride group, compared to the medium fluoride group. In addition, high fluoride exposure also reduced the mRNA and protein levels of cAMP response element-binding protein (CREB), brain-derived neurotrophic factor (BDNF), and neural cell adhesion molecule (NCAM). These findings suggested that the alteration in synaptic structure and myelin damage may partly be due to adverse effects of fluoride on the neurotrophy and neuron adhesion in mice hippocampus. Copyright © 2017 Elsevier Ltd. All rights reserved.

An autism-associated point mutation in the neuroligin cytoplasmic tail selectively impairs AMPA receptor-mediated synaptic transmission in hippocampus.

PubMed

Etherton, Mark R; Tabuchi, Katsuhiko; Sharma, Manu; Ko, Jaewon; Südhof, Thomas C

2011-06-03

Neuroligins are evolutionarily conserved postsynaptic cell-adhesion molecules that function, at least in part, by forming trans-synaptic complexes with presynaptic neurexins. Different neuroligin isoforms perform diverse functions and exhibit distinct intracellular localizations, but contain similar cytoplasmic sequences whose role remains largely unknown. Here, we analysed the effect of a single amino-acid substitution (R704C) that targets a conserved arginine residue in the cytoplasmic sequence of all neuroligins, and that was associated with autism in neuroligin-4. We introduced the R704C mutation into mouse neuroligin-3 by homologous recombination, and examined its effect on synapses in vitro and in vivo. Electrophysiological and morphological studies revealed that the neuroligin-3 R704C mutation did not significantly alter synapse formation, but dramatically impaired synapse function. Specifically, the R704C mutation caused a major and selective decrease in AMPA receptor-mediated synaptic transmission in pyramidal neurons of the hippocampus, without similarly changing NMDA or GABA receptor-mediated synaptic transmission, and without detectably altering presynaptic neurotransmitter release. Our results suggest that the cytoplasmic tail of neuroligin-3 has a central role in synaptic transmission by modulating the recruitment of AMPA receptors to postsynaptic sites at excitatory synapses.

Transsynaptic Coordination of Synaptic Growth, Function, and Stability by the L1-Type CAM Neuroglian

PubMed Central

Moreno, Eliza; Stephan, Raiko; Boerner, Jana; Godenschwege, Tanja A.; Pielage, Jan

2013-01-01

The precise control of synaptic connectivity is essential for the development and function of neuronal circuits. While there have been significant advances in our understanding how cell adhesion molecules mediate axon guidance and synapse formation, the mechanisms controlling synapse maintenance or plasticity in vivo remain largely uncharacterized. In an unbiased RNAi screen we identified the Drosophila L1-type CAM Neuroglian (Nrg) as a central coordinator of synapse growth, function, and stability. We demonstrate that the extracellular Ig-domains and the intracellular Ankyrin-interaction motif are essential for synapse development and stability. Nrg binds to Ankyrin2 in vivo and mutations reducing the binding affinities to Ankyrin2 cause an increase in Nrg mobility in motoneurons. We then demonstrate that the Nrg–Ank2 interaction controls the balance of synapse growth and stability at the neuromuscular junction. In contrast, at a central synapse, transsynaptic interactions of pre- and postsynaptic Nrg require a dynamic, temporal and spatial, regulation of the intracellular Ankyrin-binding motif to coordinate pre- and postsynaptic development. Our study at two complementary model synapses identifies the regulation of the interaction between the L1-type CAM and Ankyrin as an important novel module enabling local control of synaptic connectivity and function while maintaining general neuronal circuit architecture. PMID:23610557

Transsynaptic coordination of synaptic growth, function, and stability by the L1-type CAM Neuroglian.

PubMed

Enneking, Eva-Maria; Kudumala, Sirisha R; Moreno, Eliza; Stephan, Raiko; Boerner, Jana; Godenschwege, Tanja A; Pielage, Jan

2013-01-01

The precise control of synaptic connectivity is essential for the development and function of neuronal circuits. While there have been significant advances in our understanding how cell adhesion molecules mediate axon guidance and synapse formation, the mechanisms controlling synapse maintenance or plasticity in vivo remain largely uncharacterized. In an unbiased RNAi screen we identified the Drosophila L1-type CAM Neuroglian (Nrg) as a central coordinator of synapse growth, function, and stability. We demonstrate that the extracellular Ig-domains and the intracellular Ankyrin-interaction motif are essential for synapse development and stability. Nrg binds to Ankyrin2 in vivo and mutations reducing the binding affinities to Ankyrin2 cause an increase in Nrg mobility in motoneurons. We then demonstrate that the Nrg-Ank2 interaction controls the balance of synapse growth and stability at the neuromuscular junction. In contrast, at a central synapse, transsynaptic interactions of pre- and postsynaptic Nrg require a dynamic, temporal and spatial, regulation of the intracellular Ankyrin-binding motif to coordinate pre- and postsynaptic development. Our study at two complementary model synapses identifies the regulation of the interaction between the L1-type CAM and Ankyrin as an important novel module enabling local control of synaptic connectivity and function while maintaining general neuronal circuit architecture.

Nlgn4 knockout induces network hypo-excitability in juvenile mouse somatosensory cortex in vitro.

PubMed

Delattre, V; La Mendola, D; Meystre, J; Markram, H; Markram, K

2013-10-09

Neuroligins (Nlgns) are postsynaptic cell adhesion molecules that form transynaptic complexes with presynaptic neurexins and regulate synapse maturation and plasticity. We studied the impact of the loss of Nlgn4 on the excitatory and inhibitory circuits in somatosensory cortical slices of juvenile mice by electrically stimulating these circuits using a multi-electrode array and recording the synaptic input to single neurons using the patch-clamp technique. We detected a decreased network response to stimulation in both excitatory and inhibitory circuits of Nlgn4 knock-out animals as compared to wild-type controls, and a decreased excitation-inhibition ratio. These data indicate that Nlgn4 is involved in the regulation of excitatory and inhibitory circuits and contributes to a balanced circuit response to stimulation.

Involvement of pre- and postsynaptic NMDA receptors at local circuit interneuron connections in rat neocortex

PubMed Central

De-May, C.L.; Ali, A.B.

2013-01-01

To investigate the involvement of N-Methyl-D-aspartate (NMDA) receptors in local neocortical synaptic transmission, dual whole-cell recordings – combined with biocytin labelling – were obtained from bitufted adapting, multipolar adapting or multipolar non-adapting interneurons and pyramidal cells in layers II–V of rat (postnatal days 17–22) sensorimotor cortex. The voltage dependency of the amplitude of Excitatory postsynaptic potentials (EPSPs) received by the three types of interneuron appeared to coincide with the interneuron subclass; upon depolarisation, EPSPs received by multipolar non-adapting interneurons either decreased in amplitude or appeared insensitive, multipolar adapting interneuron EPSP amplitudes increased or appeared insensitive, whereas bitufted interneuron EPSP amplitudes increased or decreased. Connections were challenged with the NMDA receptor antagonist d-(−)-2-amino-5-phosphonopentanoic acid (d-AP5) (50 μM) revealing NMDA receptors to contribute to EPSPs received by all cell types, this also abolished the non-conventional voltage dependency. Reciprocal connections were frequent between pyramidal cells and multipolar interneurons, and inhibitory postsynaptic potentials (IPSPs) elicited in pyramidal cells by both multipolar adapting and multipolar non-adapting interneurons were sensitive to a significant reduction in amplitude by d-AP5. The involvement of presynaptic NMDA receptors was indicated by coefficient of variation analysis and an increase in the failures of transmission. Furthermore, by loading MK-801 into the pre- or postsynaptic neurons, we observed that a reduction in inhibition requires presynaptic and not postsynaptic NMDA receptors. These results suggest that NMDA receptors possess pre- and postsynaptic roles at selective neocortical synapses that are probably important in governing spike-timing and information flow. PMID:23079623

Dopamine synapse is a neuroligin-2–mediated contact between dopaminergic presynaptic and GABAergic postsynaptic structures

PubMed Central

Uchigashima, Motokazu; Ohtsuka, Toshihisa; Kobayashi, Kazuto; Watanabe, Masahiko

2016-01-01

Midbrain dopamine neurons project densely to the striatum and form so-called dopamine synapses on medium spiny neurons (MSNs), principal neurons in the striatum. Because dopamine receptors are widely expressed away from dopamine synapses, it remains unclear how dopamine synapses are involved in dopaminergic transmission. Here we demonstrate that dopamine synapses are contacts formed between dopaminergic presynaptic and GABAergic postsynaptic structures. The presynaptic structure expressed tyrosine hydroxylase, vesicular monoamine transporter-2, and plasmalemmal dopamine transporter, which are essential for dopamine synthesis, vesicular filling, and recycling, but was below the detection threshold for molecules involving GABA synthesis and vesicular filling or for GABA itself. In contrast, the postsynaptic structure of dopamine synapses expressed GABAergic molecules, including postsynaptic adhesion molecule neuroligin-2, postsynaptic scaffolding molecule gephyrin, and GABAA receptor α1, without any specific clustering of dopamine receptors. Of these, neuroligin-2 promoted presynaptic differentiation in axons of midbrain dopamine neurons and striatal GABAergic neurons in culture. After neuroligin-2 knockdown in the striatum, a significant decrease of dopamine synapses coupled with a reciprocal increase of GABAergic synapses was observed on MSN dendrites. This finding suggests that neuroligin-2 controls striatal synapse formation by giving competitive advantage to heterologous dopamine synapses over conventional GABAergic synapses. Considering that MSN dendrites are preferential targets of dopamine synapses and express high levels of dopamine receptors, dopamine synapse formation may serve to increase the specificity and potency of dopaminergic modulation of striatal outputs by anchoring dopamine release sites to dopamine-sensing targets. PMID:27035941

A ‘calcium capacitor’ shapes cholinergic inhibition of cochlear hair cells

PubMed Central

Fuchs, Paul Albert

2014-01-01

Efferent cholinergic neurons project from the brainstem to inhibit sensory hair cells of the vertebrate inner ear. This inhibitory synapse combines the activity of an unusual class of ionotropic cholinergic receptor with that of nearby calcium-dependent potassium channels to shunt and hyperpolarize the hair cell. Postsynaptic calcium signalling is constrained by a thin near-membrane cistern that is co-extensive with the efferent terminal contacts. The postsynaptic cistern may play an essential role in calcium homeostasis, serving as sink or source, depending on ongoing activity and the degree of buffer saturation. Release of calcium from postsynaptic stores leads to a process of retrograde facilitation via the synthesis of nitric oxide in the hair cell. Activity-dependent synaptic modification may contribute to changes in hair cell innervation that occur during development, and in the aged or damaged cochlea. PMID:24566542

Invaginating Structures in Mammalian Synapses

PubMed Central

Petralia, Ronald S.; Wang, Ya-Xian; Mattson, Mark P.; Yao, Pamela J.

2018-01-01

Invaginating structures at chemical synapses in the mammalian nervous system exist in presynaptic axon terminals, postsynaptic spines or dendrites, and glial processes. These invaginating structures can be divided into three categories. The first category includes slender protrusions invaginating into axonal terminals, postsynaptic spines, or glial processes. Best known examples of this category are spinules extending from postsynaptic spines into presynaptic terminals in forebrain synapses. Another example of this category are protrusions from inhibitory presynaptic terminals invaginating into postsynaptic neuronal somas. Regardless of the direction and location, the invaginating structures of the first category do not have synaptic active zones within the invagination. The second category includes postsynaptic spines invaginating into presynaptic terminals, whereas the third category includes presynaptic terminals invaginating into postsynaptic spines or dendrites. Unlike the first category, the second and third categories have active zones within the invagination. An example of the second category are mossy terminal synapses of the hippocampal CA3 region, in which enlarged spine-like structures invaginate partly or entirely into mossy terminals. An example of the third category is the neuromuscular junction (NMJ) where substantial invaginations of the presynaptic terminals invaginate into the muscle fibers. In the retina, rod and cone synapses have invaginating processes from horizontal and bipolar cells. Because horizontal cells act both as post and presynaptic structures, their invaginating processes represent both the second and third category. These invaginating structures likely play broad yet specialized roles in modulating neuronal cell signaling. PMID:29674962

Processing of the Synaptic Cell Adhesion Molecule Neurexin-3β by Alzheimer Disease α- and γ-Secretases*

PubMed Central

Bot, Nathalie; Schweizer, Claude; Ben Halima, Saoussen; Fraering, Patrick C.

2011-01-01

Neurexins (NRXNs) are synaptic cell adhesion molecules having essential roles in the assembly and maturation of synapses into fully functional units. Immunocytochemical and electrophysiological studies have shown that specific binding across the synaptic cleft of the ectodomains of presynaptic NRXNs and postsynaptic neuroligins have the potential to bidirectionally coordinate and trigger synapse formation. Moreover, in vivo studies as well as genome-wide association studies pointed out implication of NRXNs in the pathogenesis of cognitive disorders including autism spectrum disorders and different types of addictions including opioid and alcohol dependences, suggesting an important role in synaptic function. Despite extensive investigations, the mechanisms by which NRXNs modulate the properties of synapses remain largely unknown. We report here that α- and γ-secretases can sequentially process NRXN3β, leading to the formation of two final products, an ∼80-kDa N-terminal extracellular domain of Neurexin-3β (sNRXN3β) and an ∼12-kDa C-terminal intracellular NRXN3β domain (NRXN3β-ICD), both of them being potentially implicated in the regulation of NRXNs and neuroligins functions at the synapses or in yet unidentified signal transduction pathways. We further report that this processing is altered by several PS1 mutations in the catalytic subunit of the γ-secretase that cause early-onset familial Alzheimer disease. PMID:21084300

Coxsackievirus and adenovirus receptor (CAR) mediates trafficking of acid sensing ion channel 3 (ASIC3) via PSD-95.

PubMed

Excoffon, Katherine J D A; Kolawole, Abimbola O; Kusama, Nobuyoshi; Gansemer, Nicholas D; Sharma, Priyanka; Hruska-Hageman, Alesia M; Petroff, Elena; Benson, Christopher J

2012-08-17

We have previously shown that the Coxsackievirus and adenovirus receptor (CAR) can interact with post-synaptic density 95 (PSD-95) and localize PSD-95 to cell-cell junctions. We have also shown that activity of the acid sensing ion channel (ASIC3), a H(+)-gated cation channel that plays a role in mechanosensation and pain signaling, is negatively modulated by PSD-95 through a PDZ-based interaction. We asked whether CAR and ASIC3 simultaneously interact with PSD-95, and if so, whether co-expression of these proteins alters their cellular distribution and localization. Results indicate that CAR and ASIC3 co-immunoprecipitate only when co-expressed with PSD-95. CAR also brings both PSD-95 and ASIC3 to the junctions of heterologous cells. Moreover, CAR rescues PSD-95-mediated inhibition of ASIC3 currents. These data suggest that, in addition to activity as a viral receptor and adhesion molecule, CAR can play a role in trafficking proteins, including ion channels, in a PDZ-based scaffolding complex. Copyright © 2012 Elsevier Inc. All rights reserved.

Concurrent gradients of ribbon volume and AMPA-receptor patch volume in cochlear afferent synapses on gerbil inner hair cells.

PubMed

Zhang, Lichun; Engler, Sina; Koepcke, Lena; Steenken, Friederike; Köppl, Christine

2018-07-01

The Mongolian gerbil is a classic animal model for age-related hearing loss. As a prerequisite for studying age-related changes, we characterized cochlear afferent synaptic morphology in young adult gerbils, using immunolabeling and quantitative analysis of confocal microscopic images. Cochlear wholemounts were triple-labeled with a hair-cell marker, a marker of presynaptic ribbons, and a marker of postsynaptic AMPA-type glutamate receptors. Seven cochlear positions covering an equivalent frequency range from 0.5 - 32 kHz were evaluated. The spatial positions of synapses were determined in a coordinate system with reference to their individual inner hair cell. Synapse numbers confirmed previous reports for gerbils (on average, 20-22 afferents per inner hair cell). The volumes of presynaptic ribbons and postsynaptic glutamate receptor patches were positively correlated: larger ribbons associated with larger receptor patches and smaller ribbons with smaller patches. Furthermore, the volumes of both presynaptic ribbons and postsynaptic receptor patches co-varied along the modiolar-pillar and the longitudinal axes of their hair cell. The gradients in ribbon volume are consistent with previous findings in cat, guinea pig, mouse and rat and further support a role in differentiating the physiological properties of type I afferents. However, the positive correlation between the volumes of pre- and postsynaptic elements in the gerbil is different to the opposing gradients found in the mouse, suggesting species-specific differences in the postsynaptic AMPA receptors that are unrelated to the fundamental classes of type I afferents. Copyright © 2018 Elsevier B.V. All rights reserved.

Cell-type-specific role for nucleus accumbens neuroligin-2 in depression and stress susceptibility.

PubMed

Heshmati, Mitra; Aleyasin, Hossein; Menard, Caroline; Christoffel, Daniel J; Flanigan, Meghan E; Pfau, Madeline L; Hodes, Georgia E; Lepack, Ashley E; Bicks, Lucy K; Takahashi, Aki; Chandra, Ramesh; Turecki, Gustavo; Lobo, Mary Kay; Maze, Ian; Golden, Sam A; Russo, Scott J

2018-01-30

Behavioral coping strategies are critical for active resilience to stress and depression; here we describe a role for neuroligin-2 (NLGN-2) in the nucleus accumbens (NAc). Neuroligins (NLGN) are a family of neuronal postsynaptic cell adhesion proteins that are constituents of the excitatory and inhibitory synapse. Importantly, NLGN-3 and NLGN-4 mutations are strongly implicated as candidates underlying the development of neuropsychiatric disorders with social disturbances such as autism, but the role of NLGN-2 in neuropsychiatric disease states is unclear. Here we show a reduction in NLGN-2 gene expression in the NAc of patients with major depressive disorder. Chronic social defeat stress in mice also decreases NLGN-2 selectively in dopamine D1-positive cells, but not dopamine D2-positive cells, within the NAc of stress-susceptible mice. Functional NLGN-2 knockdown produces bidirectional, cell-type-specific effects: knockdown in dopamine D1-positive cells promotes subordination and stress susceptibility, whereas knockdown in dopamine D2-positive cells mediates active defensive behavior. These findings establish a behavioral role for NAc NLGN-2 in stress and depression; provide a basis for targeted, cell-type specific therapy; and highlight the role of active behavioral coping mechanisms in stress susceptibility.

Voltage-Gated Calcium Influx Modifies Cholinergic Inhibition of Inner Hair Cells in the Immature Rat Cochlea.

PubMed

Zachary, Stephen; Nowak, Nathaniel; Vyas, Pankhuri; Bonanni, Luke; Fuchs, Paul Albert

2018-06-20

Until postnatal day (P) 12, inner hair cells of the rat cochlea are invested with both afferent and efferent synaptic connections. With the onset of hearing at P12, the efferent synapses disappear, and afferent (ribbon) synapses operate with greater efficiency. This change coincides with increased expression of voltage-gated potassium channels, the loss of calcium-dependent electrogenesis, and the onset of graded receptor potentials driven by sound. The transient efferent synapses include near-membrane postsynaptic cisterns thought to regulate calcium influx through the hair cell's α9-containing and α10-containing nicotinic acetylcholine receptors. This influx activates small-conductance Ca 2+ -activated K + (SK) channels. Serial-section electron microscopy of inner hair cells from two 9-d-old (male) rat pups revealed many postsynaptic efferent cisterns and presynaptic afferent ribbons whose average minimal separation in five cells ranged from 1.1 to 1.7 μm. Efferent synaptic function was studied in rat pups (age, 7-9 d) of either sex. The duration of these SK channel-mediated IPSCs was increased by enhanced calcium influx through L-type voltage-gated channels, combined with ryanodine-sensitive release from internal stores-presumably the near-membrane postsynaptic cistern. These data support the possibility that inner hair cell calcium electrogenesis modulates the efficacy of efferent inhibition during the maturation of inner hair cell synapses. SIGNIFICANCE STATEMENT Strict calcium buffering is essential for cellular function. This problem is especially acute for compact hair cells where increasing cytoplasmic calcium promotes the opposing functions of closely adjoining afferent and efferent synapses. The near-membrane postsynaptic cistern at efferent synapses segregates synaptic calcium signals by acting as a dynamic calcium store. The hair cell serves as an informative model for synapses with postsynaptic cisterns (C synapses) found in central neurons. Copyright © 2018 the authors 0270-6474/18/385677-11$15.00/0.

Activation of postsynaptic GABAB receptors modulates the bursting pattern and synaptic activity of olfactory bulb juxtaglomerular neurons.

PubMed

Karpuk, Nikolay; Hayar, Abdallah

2008-01-01

Olfactory bulb glomeruli are formed by a network of three major types of neurons collectively called juxtaglomerular (JG) cells, which include external tufted (ET), periglomerular (PG), and short axon (SA) cells. There is solid evidence that gamma-aminobutyric acid (GABA) released from PG neurons presynaptically inhibits glutamate release from olfactory nerve terminals via activation of GABA(B) receptors (GABA(B)-Rs). However, it is still unclear whether ET cells have GABA(B)-Rs. We have investigated whether ET cells have functional postsynaptic GABA(B)-Rs using extracellular and whole cell recordings in olfactory bulb slices. In the presence of fast synaptic blockers (CNQX, APV, and gabazine), the GABA(B)-R agonist baclofen either completely inhibited the bursting or reduced the bursting frequency and increased the burst duration and the number of spikes/burst in ET cells. In the presence of fast synaptic blockers and tetrodotoxin, baclofen induced an outward current in ET cells, suggesting a direct postsynaptic effect. Baclofen reduced the frequency and amplitude of spontaneous EPSCs in PG and SA cells. In the presence of sodium and potassium channel blockers, baclofen reduced the frequency of miniature EPSCs, which were inhibited by the calcium channel blocker cadmium. All baclofen effects were reversed by application of the GABA(B)-R antagonist CGP55845. We suggest that activation of GABA(B)-Rs directly inhibits ET cell bursting and decreases excitatory dendrodendritic transmission from ET to PG and SA cells. Thus the postsynaptic GABA(B)-Rs on ET cells may play an important role in shaping the activation pattern of the glomeruli during olfactory coding.

A network of autism linked genes stabilizes two pools of synaptic GABAA receptors

PubMed Central

Tong, Xia-Jing; Hu, Zhitao; Liu, Yu; Anderson, Dorian; Kaplan, Joshua M

2015-01-01

Changing receptor abundance at synapses is an important mechanism for regulating synaptic strength. Synapses contain two pools of receptors, immobilized and diffusing receptors, both of which are confined to post-synaptic elements. Here we show that immobile and diffusing GABAA receptors are stabilized by distinct synaptic scaffolds at C. elegans neuromuscular junctions. Immobilized GABAA receptors are stabilized by binding to FRM-3/EPB4.1 and LIN-2A/CASK. Diffusing GABAA receptors are stabilized by the synaptic adhesion molecules Neurexin and Neuroligin. Inhibitory post-synaptic currents are eliminated in double mutants lacking both scaffolds. Neurexin, Neuroligin, and CASK mutations are all linked to Autism Spectrum Disorders (ASD). Our results suggest that these mutations may directly alter inhibitory transmission, which could contribute to the developmental and cognitive deficits observed in ASD. DOI: http://dx.doi.org/10.7554/eLife.09648.001 PMID:26575289

Dynamical model of long-term synaptic plasticity

PubMed Central

Abarbanel, Henry D. I.; Huerta, R.; Rabinovich, M. I.

2002-01-01

Long-term synaptic plasticity leading to enhancement in synaptic efficacy (long-term potentiation, LTP) or decrease in synaptic efficacy (long-term depression, LTD) is widely regarded as underlying learning and memory in nervous systems. LTP and LTD at excitatory neuronal synapses are observed to be induced by precise timing of pre- and postsynaptic events. Modification of synaptic transmission in long-term plasticity is a complex process involving many pathways; for example, it is also known that both forms of synaptic plasticity can be induced by various time courses of Ca2+ introduction into the postsynaptic cell. We present a phenomenological description of a two-component process for synaptic plasticity. Our dynamical model reproduces the spike time-dependent plasticity of excitatory synapses as a function of relative timing between pre- and postsynaptic events, as observed in recent experiments. The model accounts for LTP and LTD when the postsynaptic cell is voltage clamped and depolarized (LTP) or hyperpolarized (LTD) and no postsynaptic action potentials are evoked. We are also able to connect our model with the Bienenstock, Cooper, and Munro rule. We give model predictions for changes in synaptic strength when periodic spike trains of varying frequency and Poisson distributed spike trains with varying average frequency are presented pre- and postsynaptically. When the frequency of spike presentation exceeds ≈30–40 Hz, only LTP is induced. PMID:12114531

Analysis of the genes encoding neuroligins NLGN3 and NLGN4 in Bulgarian patients with autism.

PubMed

Avdjieva-Tzavella, D M; Todorov, T P; Todorova, A P; Kirov, A V; Hadjidekova, S P; Rukova, B B; Litvinenko, I O; Hristova-Naydenova, D N; Tincheva, R S; Toncheva, D I

2012-01-01

Many studies have supported a genetic aetiology for autism. Neuroligins are postsynaptically located cell-adhesion molecules. Mutations in two X-linked neuroligin genes, NLGN3 and NLGN4, have been implicated in pathogenesis of autism. In order to confirm these causative mutations in our autistic population and to determine their frequency we screened 20 individuals affected with autism. We identified one patient with a point mutation in NLGN4 gene that substituted a Met for Thr 787 - c.2360C > T, p.(Thr787Met) and three patients with identical polymorphisms in the same gene: c.933C > T, p.(Thr311Thr) in combination with c.[1777C > T+1779C > G, p.(Leu593Leu)]. All patients tested for NLGN3 mutations were negative. These results indicate that mutations in these genes are responsible for at most a small fraction of autism cases.

Selective Localization of Shanks to VGLUT1-Positive Excitatory Synapses in the Mouse Hippocampus

PubMed Central

Heise, Christopher; Schroeder, Jan C.; Schoen, Michael; Halbedl, Sonja; Reim, Dominik; Woelfle, Sarah; Kreutz, Michael R.; Schmeisser, Michael J.; Boeckers, Tobias M.

2016-01-01

Members of the Shank family of multidomain proteins (Shank1, Shank2, and Shank3) are core components of the postsynaptic density (PSD) of excitatory synapses. At synaptic sites Shanks serve as scaffolding molecules that cluster neurotransmitter receptors as well as cell adhesion molecules attaching them to the actin cytoskeleton. In this study we investigated the synapse specific localization of Shank1-3 and focused on well-defined synaptic contacts within the hippocampal formation. We found that all three family members are present only at VGLUT1-positive synapses, which is particularly visible at mossy fiber contacts. No costaining was found at VGLUT2-positive contacts indicating that the molecular organization of VGLUT2-associated PSDs diverges from classical VGLUT1-positive excitatory contacts in the hippocampus. In light of SHANK mutations in neuropsychiatric disorders, this study indicates which glutamatergic networks within the hippocampus will be primarily affected by shankopathies. PMID:27199660

Selective Localization of Shanks to VGLUT1-Positive Excitatory Synapses in the Mouse Hippocampus.

PubMed

Heise, Christopher; Schroeder, Jan C; Schoen, Michael; Halbedl, Sonja; Reim, Dominik; Woelfle, Sarah; Kreutz, Michael R; Schmeisser, Michael J; Boeckers, Tobias M

2016-01-01

Members of the Shank family of multidomain proteins (Shank1, Shank2, and Shank3) are core components of the postsynaptic density (PSD) of excitatory synapses. At synaptic sites Shanks serve as scaffolding molecules that cluster neurotransmitter receptors as well as cell adhesion molecules attaching them to the actin cytoskeleton. In this study we investigated the synapse specific localization of Shank1-3 and focused on well-defined synaptic contacts within the hippocampal formation. We found that all three family members are present only at VGLUT1-positive synapses, which is particularly visible at mossy fiber contacts. No costaining was found at VGLUT2-positive contacts indicating that the molecular organization of VGLUT2-associated PSDs diverges from classical VGLUT1-positive excitatory contacts in the hippocampus. In light of SHANK mutations in neuropsychiatric disorders, this study indicates which glutamatergic networks within the hippocampus will be primarily affected by shankopathies.

Patient autoantibodies deplete postsynaptic muscle-specific kinase leading to disassembly of the ACh receptor scaffold and myasthenia gravis in mice

PubMed Central

Cole, R N; Ghazanfari, N; Ngo, S T; Gervásio, O L; Reddel, S W; Phillips, W D

2010-01-01

The postsynaptic muscle-specific kinase (MuSK) coordinates formation of the neuromuscular junction (NMJ) during embryonic development. Here we have studied the effects of MuSK autoantibodies upon the NMJ in adult mice. Daily injections of IgG from four MuSK autoantibody-positive myasthenia gravis patients (MuSK IgG; 45 mg day−1i.p. for 14 days) caused reductions in postsynaptic ACh receptor (AChR) packing as assessed by fluorescence resonance energy transfer (FRET). IgG from the patients with the highest titres of MuSK autoantibodies caused large (51–73%) reductions in postsynaptic MuSK staining (cf. control mice; P < 0.01) and muscle weakness. Among mice injected for 14 days with control and MuSK patient IgGs, the residual level of MuSK correlated with the degree of impairment of postsynaptic AChR packing. However, the loss of postsynaptic MuSK preceded this impairment of postsynaptic AChR. When added to cultured C2 muscle cells the MuSK autoantibodies caused tyrosine phosphorylation of MuSK and the AChR β-subunit, and internalization of MuSK from the plasma membrane. The results suggest a pathogenic mechanism in which MuSK autoantibodies rapidly deplete MuSK from the postsynaptic membrane leading to progressive dispersal of postsynaptic AChRs. Moreover, maintenance of postsynaptic AChR packing at the adult NMJ would appear to depend upon physical engagement of MuSK with the AChR scaffold, notwithstanding activation of the MuSK-rapsyn system of AChR clustering. PMID:20603331

Molecular organization of excitatory chemical synapses in the mammalian brain

NASA Astrophysics Data System (ADS)

Gundelfinger, E. D.; tom Dieck, S.

Chemical synapses are highly specialized cell-cell junctions designed for efficient signaling between nerve cells. Distinct cytoskeletal matrices are assembled at either side of the synaptic junction. The presynaptic cytomatrix at the active zone (CAZ) defines and organizes the site of neurotransmitter release from presynaptic nerve terminals. The postsynaptic density (PSD) tethers neurotransmitter receptors and the postsynaptic signal transduction machinery. Recent progress in the identification and characterization of novel CAZ and PSD components has revealed new insights into the molecular organization and assembly mechanisms of the synaptic neurotransmission apparatus. On the presynaptic side, Bassoon and Piccolo, two related giant proteins, are crucially involved in scaffolding the CAZ. On the postsynaptic side, two families of multi-domain adaptor proteins, the MAGuKs (membrane-associated guanylate kinase homologs) and the ProSAP (proline-rich synapse-associated protein, also termed Shank) family members are thought to be major organizing molecules of the PSD.

Neurobeachin is required postsynaptically for electrical and chemical synapse formation

PubMed Central

Miller, Adam C.; Voelker, Lisa H.; Shah, Arish N.; Moens, Cecilia B.

2014-01-01

Summary Background Neural networks and their function are defined by synapses, which are adhesions specialized for intercellular communication that can be either chemical or electrical. At chemical synapses transmission between neurons is mediated by neurotransmitters, while at electrical synapses direct ionic and metabolic coupling occurs via gap junctions between neurons. The molecular pathways required for electrical synaptogenesis are not well understood and whether they share mechanisms of formation with chemical synapses is not clear. Results Here, using a forward genetic screen in zebrafish we find that the autism-associated gene neurobeachin (nbea), which encodes a BEACH-domain containing protein implicated in endomembrane trafficking, is required for both electrical and chemical synapse formation. Additionally, we find that nbea is dispensable for axonal formation and early dendritic outgrowth, but is required to maintain dendritic complexity. These synaptic and morphological defects correlate with deficiencies in behavioral performance. Using chimeric animals in which individually identifiable neurons are either mutant or wildtype we find that Nbea is necessary and sufficient autonomously in the postsynaptic neuron for both synapse formation and dendritic arborization. Conclusions Our data identify a surprising link between electrical and chemical synapse formation and show that Nbea acts as a critical regulator in the postsynaptic neuron for the coordination of dendritic morphology with synaptogenesis. PMID:25484298

Focal adhesion kinase regulates neuronal growth, synaptic plasticity and hippocampus-dependent spatial learning and memory.

PubMed

Monje, Francisco J; Kim, Eun-Jung; Pollak, Daniela D; Cabatic, Maureen; Li, Lin; Baston, Arthur; Lubec, Gert

2012-01-01

The focal adhesion kinase (FAK) is a non-receptor tyrosine kinase abundantly expressed in the mammalian brain and highly enriched in neuronal growth cones. Inhibitory and facilitatory activities of FAK on neuronal growth have been reported and its role in neuritic outgrowth remains controversial. Unlike other tyrosine kinases, such as the neurotrophin receptors regulating neuronal growth and plasticity, the relevance of FAK for learning and memory in vivo has not been clearly defined yet. A comprehensive study aimed at determining the role of FAK in neuronal growth, neurotransmitter release and synaptic plasticity in hippocampal neurons and in hippocampus-dependent learning and memory was therefore undertaken using the mouse model. Gain- and loss-of-function experiments indicated that FAK is a critical regulator of hippocampal cell morphology. FAK mediated neurotrophin-induced neuritic outgrowth and FAK inhibition affected both miniature excitatory postsynaptic potentials and activity-dependent hippocampal long-term potentiation prompting us to explore the possible role of FAK in spatial learning and memory in vivo. Our data indicate that FAK has a growth-promoting effect, is importantly involved in the regulation of the synaptic function and mediates in vivo hippocampus-dependent spatial learning and memory. Copyright © 2011 S. Karger AG, Basel.

Nucleus reticularis neurons mediate diverse inhibitory effects in thalamus.

PubMed

Cox, C L; Huguenard, J R; Prince, D A

1997-08-05

Detailed information regarding the contribution of individual gamma-aminobutyric acid (GABA)-containing inhibitory neurons to the overall synaptic activity of single postsynaptic cells is essential to our understanding of fundamental elements of synaptic integration and operation of neuronal circuits. For example, GABA-containing cells in the thalamic reticular nucleus (nRt) provide major inhibitory innervation of thalamic relay nuclei that is critical to thalamocortical rhythm generation. To investigate the contribution of individual nRt neurons to the strength of this internuclear inhibition, we obtained whole-cell recordings of unitary inhibitory postsynaptic currents (IPSCs) evoked in ventrobasal thalamocortical (VB) neurons by stimulation of single nRt cells in rat thalamic slices, in conjunction with intracellular biocytin labeling. Two types of monosynaptic IPSCs could be distinguished. "Weak" inhibitory connections were characterized by a significant number of postsynaptic failures in response to presynaptic nRt action potentials and relatively small IPSCs. In contrast, "strong" inhibition was characterized by the absence of postsynaptic failures and significantly larger unitary IPSCs. By using miniature IPSC amplitudes to infer quantal size, we estimated that unitary IPSCs associated with weak inhibition resulted from activation of 1-3 release sites, whereas stronger inhibition would require simultaneous activation of 5-70 release sites. The inhibitory strengths were positively correlated with the density of axonal swellings of the presynaptic nRt neurons, an indicator that characterizes different nRt axonal arborization patterns. These results demonstrate that there is a heterogeneity of inhibitory interactions between nRt and VB neurons, and that variations in gross morphological features of axonal arbors in the central nervous system can be associated with significant differences in postsynaptic response characteristics.

The quantal release at a neuro-neuronal synapse is regulated by the content of acetylcholine in the presynaptic cell.

PubMed

Poulain, B; Baux, G; Tauc, L

1986-01-01

Transmitter release was studied with respect to the presynaptic acetylcholine (ACh) content at a central identified inhibitory synapse (Cl- conductance) of Aplysia californica. Statistical analysis of the synaptic noise evoked by sustained depolarization of the presynaptic neuron allowed us to calculate the quantal parameters of the postsynaptic responses. Loading of the presynaptic neurone with injected ACh led to an increase in the postsynaptic responses whereas the calculated miniature postsynaptic current (MPSC) was unmodified. Destruction of choline by choline oxidase either applied extracellularly and coupled to intense stimulations of the presynaptic cell or injected into the presynaptic neuron induced a depression of the postsynaptic response although the amplitude of the calculated MPSC remained constant. As the size of the MPSC, i.e. the size of the quantum, did not change in these experiments, it was concluded that the presynaptic ACh content controls the number of quanta released by a given presynaptic depolarization. As additional evidence, effects of abrupt increase in tonicity of the external medium were studied. The observed transient enhancement of the quantal content of the postsynaptic response could be attributed to an increase in the presynaptic concentration of ACh, resulting from the reduction in cellular volume.

Not all neuroligin 3 and 4X missense variants lead to significant functional inactivation.

PubMed

Xu, Xiaojuan; Hu, Zhengmao; Zhang, Lusi; Liu, Hongfang; Cheng, Yuemei; Xia, Kun; Zhang, Xuehong

2017-09-01

Neuroligins are postsynaptic cell adhesion molecules that interact with neurexins to regulate the fine balance between excitation and inhibition of synapses. Recently, accumulating evidence, involving mutation analysis, cellular assays, and mouse models, has suggested that neuroligin (NLGN) mutations affect synapse maturation and function. Previously, four missense variations [p.G426S (NLGN3), p.G84R (NLGN4X), p.Q162K (NLGN4X), and p.A283T (NLGN4X)] in four different unrelated patients have been identified by PCR and direct sequencing. In this study, we analyzed the functional effect of these missense variations by in vitro experiment via the stable HEK293 cells expressing wild-type and mutant neuroligin. We found that the four mutations did not significantly impair the expression of neuroligin 3 and neuroligin 4X, and also did not measurably inhibit the neurexin 1-neuroligin interaction. These variants might play a modest role in the pathogenesis of autism or might simply be unreported infrequent polymorphisms. Our data suggest that these four previously described neuroligin mutations are not primary risk factors for autism.

Isolation of Synaptosomes, Synaptic Plasma Membranes, and Synaptic Junctional Complexes.

PubMed

Michaelis, Mary L; Jiang, Lei; Michaelis, Elias K

2017-01-01

Isolation of synaptic nerve terminals or synaptosomes provides an opportunity to study the process of neurotransmission at many levels and with a variety of approaches. For example, structural features of the synaptic terminals and the organelles within them, such as synaptic vesicles and mitochondria, have been elucidated with electron microscopy. The postsynaptic membranes are joined to the presynaptic "active zone" of transmitter release through cell adhesion molecules and remain attached throughout the isolation of synaptosomes. These "post synaptic densities" or "PSDs" contain the receptors for the transmitters released from the nerve terminals and can easily be seen with electron microscopy. Biochemical and cell biological studies with synaptosomes have revealed which proteins and lipids are most actively involved in synaptic release of neurotransmitters. The functional properties of the nerve terminals, such as responses to depolarization and the uptake or release of signaling molecules, have also been characterized through the use of fluorescent dyes, tagged transmitters, and transporter substrates. In addition, isolated synaptosomes can serve as the starting material for the isolation of relatively pure synaptic plasma membranes (SPMs) that are devoid of organelles from the internal environment of the nerve terminal, such as mitochondria and synaptic vesicles. The isolated SPMs can reseal and form vesicular structures in which transport of ions such as sodium and calcium, as well as solutes such as neurotransmitters can be studied. The PSDs also remain associated with the presynaptic membranes during isolation of SPM fractions, making it possible to isolate the synaptic junctional complexes (SJCs) devoid of the rest of the plasma membranes of the nerve terminals and postsynaptic membrane components. Isolated SJCs can be used to identify the proteins that constitute this highly specialized region of neurons. In this chapter, we describe the steps involved in isolating synaptosomes, SPMs, and SJCs from brain so that these preparations can be used with new technological advances to address many as yet unanswered questions about the synapse and its remarkable activities in neuronal cell communication.

Unitary IPSPs evoked by interneurons at the stratum radiatum-stratum lacunosum-moleculare border in the CA1 area of the rat hippocampus in vitro

PubMed Central

Vida, Imre; Halasy, Katalin; Szinyei, Csaba; Somogyi, Peter; Buhl, Eberhard H

1998-01-01

Hippocampal non-principal neurons at the stratum radiatum-stratum lacunosum-moleculare border (R-LM interneurons) of the CA1 area may constitute several cell classes and have been implicated in the generation of GABAergic unitary IPSPs. Using biocytin-filled electrodes we recorded R-LM interneurons intracellularly in vitro and determined their postsynaptic effects in concomitantly recorded pyramidal cells. Light microscopic analysis revealed four populations of R-LM interneurons with distinct axons: (1) basket cells (n= 4) with axons predominantly ramifying in the pyramidal cell layer; (2) Schaffer collateral/commissural pathway-associated interneurons (n= 10) stratifying in stratum radiatum and, to a lesser extent, stratum oriens; (3) perforant pathway-associated interneurons (n= 6) innervating the perforant path termination zone in stratum lacunosum-moleculare of the CA1 area as well as equivalent portions of the dentate gyrus and subiculum; and (4) neurogliaform interneurons (n= 2) characterized by their dense, compact axonal and dendritic arbour. Random electron microscopic sampling of synaptic targets revealed a preponderance of pyramidal neurons as postsynaptic elements. Basket cells had a synaptic target preference for somata and proximal dendrites, whereas the remainder of R-LM interneurons innervated dendritic shafts and spines. The axon of dendrite-targeting cells formed up to six putative contacts with individual postsynaptic pyramidal cells. Anatomically recovered R-LM interneurons (n= 22) had a mean resting membrane potential of -56.7 ± 3.6 mV, a membrane time constant of 12.9 ± 7.7 ms and an input resistance of 86.4 ± 29.2 MΩ. Depolarizing current pulses generally elicited overshooting action potentials (70.8 ± 6.9 mV) which had a mean duration, when measured at half-amplitude, of 0.7 ± 0.1 ms. In response to prolonged (> 200 ms) depolarizing current pulses all R-LM interneurons displayed (a varying degree of) spike frequency adaptation. Basket cells, Schaffer-associated and neurogliaform interneurons elicited small-amplitude (< 2 mV), short-latency IPSPs in postsynaptic pyramids (n= 5, 13 and 1, respectively). Those interactions in which an effect was elicited with the repetitive activation of the presynaptic neuron (n= 13) showed a substantial degree of postsynaptic response summation. Unitary IPSPs had fast kinetics and, whenever tested (n= 5; 1 basket cell and 4 Schaffer-associated interneurons), were abolished by the GABAA receptor antagonist bicuculline. Thus, R-LM interneurons comprise several distinct populations which evoke fast GABAA receptor-mediated IPSPs. The domain-specific innervation of postsynaptic pyramidal cells suggests functionally diverse effects on the integration of afferent information in functionally non-equivalent compartments of pyramidal cells. PMID:9503336

Synaptic hyperpolarization and inhibition of turtle cochlear hair cells.

PubMed

Art, J J; Fettiplace, R; Fuchs, P A

1984-11-01

Intracellular recordings were made from turtle cochlear hair cells in order to examine the properties of the post-synaptic potentials evoked by electrical stimulation of the efferent axons. Single shocks to the efferents generated a hair cell membrane hyperpolarization with an average amplitude generally less than 1 mV and lasting for about 100 ms. With short trains of shocks, the size of the post-synaptic potential grew markedly to a maximum of 20-30 mV. The interaction between pairs of shocks separated by a varying interval was studied. For an interval of 4 ms, the response to the second shock was increased on average by a factor of 3 and the conditioning effect of the first shock decayed with a time constant of about 100 ms. We suggest the augmentation in response to trains of shocks may be partly due to facilitation of efferent transmitter release. The efferent post-synaptic potentials could be reversibly abolished by perfusion with perilymphs containing 3 microM-curare or atropine, and infusion of acetylcholine gave a transient membrane hyperpolarization. These observations are consistent with efferent action being mediated via a cholinergic synapse onto the hair cells. The post-synaptic potentials could be reversed in polarity by injection of hyperpolarizing currents through the recording electrode. The reversal potential was estimated as about -80 mV, 30 mV negative to the resting potential. Near reversal, a small brief depolarization was evident and may constitute a minor component of the synaptic response. The value of the reversal potential was unaffected by substitution of the perilymphatic chloride, but was altered in a predictable manner by changes in extracellular potassium concentration indicating that the post-synaptic potentials arise mainly by an increase in the permeability of the hair cell membrane to potassium ions. Throughout the post-synaptic hyperpolarization there was a reduction in the sensitivity of the hair cell to tones at its characteristic frequency. The desensitization, maximal for low sound pressures, varied in different cells from a factor of 1.6 to 28. At the peak of the largest synaptic potentials, the receptor potential remained negative to the resting potential with all but the loudest characteristic frequency tone s. We suggest that there are two factors in efferent inhibition; one a r duction in the receptor potential at the hair cell's characteristic frequency and the other a hyperpolarization of its membrane potential which should reduce the release of excitatory transmitter onto the afferent terminals.

Nongenomic Glucocorticoid Suppression of a Postsynaptic Potassium Current via Emergent Autocrine Endocannabinoid Signaling in Hypothalamic Neuroendocrine Cells following Chronic Dehydration

PubMed Central

Wu, Ning

2017-01-01

Glucocorticoids rapidly stimulate endocannabinoid synthesis and modulation of synaptic transmission in hypothalamic neuroendocrine cells via a nongenomic signaling mechanism. The endocannabinoid actions are synapse-constrained by astrocyte restriction of extracellular spatial domains. Exogenous cannabinoids have been shown to modulate postsynaptic potassium currents, including the A-type potassium current (IA), in different cell types. The activity of magnocellular neuroendocrine cells is shaped by a prominent IA. We tested for a rapid glucocorticoid modulation of the postsynaptic IK and IA in magnocellular neuroendocrine cells of the hypothalamic paraventricular nucleus (PVN) using whole-cell recordings in rat brain slices. Application of the synthetic glucocorticoid dexamethasone (Dex) had no rapid effect on the IK or IA amplitude, voltage dependence, or kinetics in magnocellular neurons in slices from untreated rats. In magnocellular neurons from salt-loaded rats, however, Dex application caused a rapid suppression of the IA and a depolarizing shift in IA voltage dependence. Exogenously applied endocannabinoids mimicked the rapid Dex modulation of the IA, and CB1 receptor antagonists and agonists blocked and occluded the Dex-induced changes in the IA, respectively, suggesting an endocannabinoid dependence of the rapid glucocorticoid effect. Preincubation of control slices in a gliotoxin resulted in the partial recapitulation of the glucocorticoid-induced rapid suppression of the IA. These findings demonstrate a glucocorticoid suppression of the postsynaptic IA in PVN magnocellular neurons via an autocrine endocannabinoid-dependent mechanism following chronic dehydration, and suggest a possible role for astrocytes in the control of the autocrine endocannabinoid actions. PMID:28966975

High-frequency stimulation of the temporoammonic pathway induces input-specific long-term potentiation in subicular bursting cells.

PubMed

Fidzinski, Pawel; Wawra, Matthias; Bartsch, Julia; Heinemann, Uwe; Behr, Joachim

2012-01-09

The subiculum (Sub) as a part of the hippocampal formation is thought to play a functional role in learning and memory. In addition to its major input from CA1 pyramidal cells, the subiculum receives input from the entorhinal cortex (EC) via the temporoammonic pathway. Thus far, synaptic plasticity in the subiculum was mainly investigated at CA1-Sub synapses. According to their spiking pattern, pyramidal cells in the subiculum were classified as bursting cells and non-bursting cells. In the present study, we demonstrate that subicular bursting cells show input-specific forms of long-term potentiation (LTP). At CA1-Sub synapses, bursting cells have been shown to express a presynaptic NMDA receptor-dependent LTP that depends on the activation of a cAMP-PKA cascade (Wozny et al., Journal of Physiology 2008). In contrast, at EC-Sub synapses the induction of LTP in bursting cells shows a high induction-threshold and relies on the activation of postsynaptic NMDA receptors, postsynaptic depolarization and postsynaptic Ca(2+) influx. Each form of LTP is input-specific and fails to induce heterosynaptic plasticity. Taken together, our data suggest that distinct, input-specific mechanisms govern high frequency-induced LTP at subicular bursting cells' synapses. Copyright © 2011 Elsevier B.V. All rights reserved.

Modulation of Central Synapses by Astrocyte-Released ATP and Postsynaptic P2X Receptors

PubMed Central

Pankratov, Yuriy

2017-01-01

Communication between neuronal and glial cells is important for neural plasticity. P2X receptors are ATP-gated cation channels widely expressed in the brain where they mediate action of extracellular ATP released by neurons and/or glia. Recent data show that postsynaptic P2X receptors underlie slow neuromodulatory actions rather than fast synaptic transmission at brain synapses. Here, we review these findings with a particular focus on the release of ATP by astrocytes and the diversity of postsynaptic P2X-mediated modulation of synaptic strength and plasticity in the CNS. PMID:28845311

Transmission, Development, and Plasticity of Synapses

PubMed Central

Harris, Kathryn P.

2015-01-01

Chemical synapses are sites of contact and information transfer between a neuron and its partner cell. Each synapse is a specialized junction, where the presynaptic cell assembles machinery for the release of neurotransmitter, and the postsynaptic cell assembles components to receive and integrate this signal. Synapses also exhibit plasticity, during which synaptic function and/or structure are modified in response to activity. With a robust panel of genetic, imaging, and electrophysiology approaches, and strong evolutionary conservation of molecular components, Drosophila has emerged as an essential model system for investigating the mechanisms underlying synaptic assembly, function, and plasticity. We will discuss techniques for studying synapses in Drosophila, with a focus on the larval neuromuscular junction (NMJ), a well-established model glutamatergic synapse. Vesicle fusion, which underlies synaptic release of neurotransmitters, has been well characterized at this synapse. In addition, studies of synaptic assembly and organization of active zones and postsynaptic densities have revealed pathways that coordinate those events across the synaptic cleft. We will also review modes of synaptic growth and plasticity at the fly NMJ, and discuss how pre- and postsynaptic cells communicate to regulate plasticity in response to activity. PMID:26447126

Excitatory Synaptic Drive and Feedforward Inhibition in the Hippocampal CA3 Circuit Are Regulated by SynCAM 1.

PubMed

Park, Kellie A; Ribic, Adema; Laage Gaupp, Fabian M; Coman, Daniel; Huang, Yuegao; Dulla, Chris G; Hyder, Fahmeed; Biederer, Thomas

2016-07-13

Select adhesion proteins control the development of synapses and modulate their structural and functional properties. Despite these important roles, the extent to which different synapse-organizing mechanisms act across brain regions to establish connectivity and regulate network properties is incompletely understood. Further, their functional roles in different neuronal populations remain to be defined. Here, we applied diffusion tensor imaging (DTI), a modality of magnetic resonance imaging (MRI), to map connectivity changes in knock-out (KO) mice lacking the synaptogenic cell adhesion protein SynCAM 1. This identified reduced fractional anisotropy in the hippocampal CA3 area in absence of SynCAM 1. In agreement, mossy fiber refinement in CA3 was impaired in SynCAM 1 KO mice. Mossy fibers make excitatory inputs onto postsynaptic specializations of CA3 pyramidal neurons termed thorny excrescences and these structures were smaller in the absence of SynCAM 1. However, the most prevalent targets of mossy fibers are GABAergic interneurons and SynCAM 1 loss unexpectedly reduced the number of excitatory terminals onto parvalbumin (PV)-positive interneurons in CA3. SynCAM 1 KO mice additionally exhibited lower postsynaptic GluA1 expression in these PV-positive interneurons. These synaptic imbalances in SynCAM 1 KO mice resulted in CA3 disinhibition, in agreement with reduced feedforward inhibition in this network in the absence of SynCAM 1-dependent excitatory drive onto interneurons. In turn, mice lacking SynCAM 1 were impaired in memory tasks involving CA3. Our results support that SynCAM 1 modulates excitatory mossy fiber inputs onto both interneurons and principal neurons in the hippocampal CA3 area to balance network excitability. This study advances our understanding of synapse-organizing mechanisms on two levels. First, the data support that synaptogenic proteins guide connectivity and can function in distinct brain regions even if they are expressed broadly. Second, the results demonstrate that a synaptogenic process that controls excitatory inputs to both pyramidal neurons and interneurons can balance excitation and inhibition. Specifically, the study reveals that hippocampal CA3 connectivity is modulated by the synapse-organizing adhesion protein SynCAM 1 and identifies a novel, SynCAM 1-dependent mechanism that controls excitatory inputs onto parvalbumin-positive interneurons. This enables SynCAM 1 to regulate feedforward inhibition and set network excitability. Further, we show that diffusion tensor imaging is sensitive to these cellular refinements affecting neuronal connectivity. Copyright © 2016 the authors 0270-6474/16/367465-12$15.00/0.

GABAB receptor-mediated responses in GABAergic projection neurones of rat nucleus reticularis thalami in vitro.

PubMed

Ulrich, D; Huguenard, J R

1996-06-15

1. Whole-cell voltage-clamp recordings were obtained from GABAergic neurones of rat nucleus reticularis thalami (NRT) in vitro to assess pre- and postsynaptic GABAB receptor-mediated responses. Presynaptic inhibition of GABA release was studied at terminals on local axon collaterals within NRT as well as on projection fibres in the somatosensory relay nuclei. 2. The GABAB receptor agonist (R)-baclofen (10 microM) reduced monosynaptically evoked GABAA-mediated inhibitory postsynaptic currents (IPSCs) in NRT and somatosensory relay cells to 11 and 12% of control, respectively. 3. Action potential-independent miniature IPSCs (mIPSCs) were observed in both cell types. Mean mIPSC amplitude was 20 pA in both NRT and relay cells at a holding potential of 0 mV. The mean mIPSC frequencies were 0.83 and 2.2 Hz in NRT and relay cells, respectively. Baclofen decreased mIPSP frequency by about half in each cell type without affecting amplitude. 4. Paired-burst inhibition of evoked IPSCs was studied in relay and NRT cells by applying pairs of 100 Hz stimulus bursts separated by 600 ms. The mean ratio of second to first peak IPSC amplitudes was 0.77. 5. In NRT cells baclofen induced a linear postsynaptic conductance increase of 0.82 nS with an associated reversal potential of -121 mV. A small (0.14 nS) GABAB component of the evoked IPSC was detected in only a minority of NRT cells (3 of 18). 6. All pre- and postsynaptic effects of baclofen, as well as PBI, were largely reversed by the specific GABAB receptor antagonist CGP 35348 (0.5 mM). 7. We conclude that activation of GABAB receptors in NRT leads to presynaptic autoinhibition of IPSCs in both NRT and relay cells, and to direct activation of a small linear K+ conductance. In addition our experiments suggest that reciprocal connectivity within NRT can be partially mediated by a small GABAB inhibitory event.

Relationships between morphology and physiology of pyramid-pyramid single axon connections in rat neocortex in vitro.

PubMed Central

Deuchars, J; West, D C; Thomson, A M

1994-01-01

1. Double intracellular recordings were made from 1163 pairs of pyramidal neurones in layer V-VI of the rat somatomotor cortex in vitro using sharp electrodes filled with biocytin. Monosynaptically connected pairs of cells were identified when an action potential in one could elicit a constant latency excitatory postsynaptic potential (EPSP) in the other and the cells were filled with biocytin. Labelled cells were subsequently identified histologically with avidin-horseradish peroxidase. 2. Thirty-four pairs of cells were found to be monosynaptically connected. Fifteen of these pairs were sufficiently stable for electrophysiological recordings and three of these were recovered sufficiently to permit full morphological reconstruction. 3. The EPSP recorded between the first pair of pyramids varied in amplitude between 0 and 3 mV (mean 1.33 +/- 1.06 mV) and fluctuated considerably (coefficient of variation, 0.796). This was largely due to a high incidence of apparent failures of transmission. On reconstruction two boutons from the presynaptic pyramid axon were in close apposition to the proximal portions of basal dendrites of the postsynaptic cell. 4. In the second pair of pyramids the EPSP had a mean amplitude of 1.06 mV, and displayed a 10-90% rise time of 2.8 ms and a width at half-amplitude of 23 ms. This EPSP did not alter significantly with changes in membrane potential at the soma. The presynaptic axon closely apposed the distal apical dendrite of the postsynaptic cell in eight places. 5. In the third pair of pyramids, the EPSPs, recorded at a relatively depolarized membrane potential, were long lasting and could elicit slow dendritic spikes with long and variable latencies. These slow spikes suggested that the postsynaptic recording site was dendritic and on reconstruction a possible location was identified on the apical dendrite. A total of five presynaptic boutons closely apposed three separate, proximal branches of the postsynaptic apical dendrite. 6. These results provide the first illustration of a morphological basis for variations in functional properties of pyramid-pyramid connections in the neocortex. Images Figure 1 Figure 3 Figure 5 PMID:7965856

Calcium currents and graded synaptic transmission between heart interneurons of the leech.

PubMed

Angstadt, J D; Calabrese, R L

1991-03-01

Synaptic transmission between reciprocally inhibitory heart interneurons (HN cells) of the medicinal leech was examined in the absence of Na-mediated action potentials. Under voltage clamp, depolarizing steps from a holding potential of -60 mV elicited 2 kinetically distinct components of inward current in the presynaptic HN cell: an early transient current that inactivates within 200 msec and a persistent current that only partially decays over several seconds. Both currents begin to activate near -60 mV. Steady-state inactivation occurs over the voltage range between -70 and -45 mV and is completely removed by 1-2-sec hyperpolarizing voltage steps to -80 mV. The inward currents are carried by Ca2+, Ba2+, or Sr2+ ions, but not by Co2+, Mn2+, or Ni2+. These same inward currents underlie the burst-generating plateau potentials previously described in HN cells (Arbas and Calabrese, 1987a,b). With a presynaptic holding potential of -60 mV, the threshold for transmitter release is near -45 mV. Postsynaptic currents in the contralateral HN cell have a reversal potential near -60 mV. The largest postsynaptic currents (300-400 pA) exhibit an initial peak response that is followed by a more slowly decaying component. The persistent component of Ca2+ current in the presynaptic neuron is strongly correlated with the prolonged component of the postsynaptic current, while the transient presynaptic Ca2+ current appears to correspond to the early peak of postsynaptic current. These data are consistent with the hypothesis that voltage-dependent calcium currents contribute to the oscillatory capability of reciprocally inhibitory HN cells by (1) generating the plateau potential that drives the burst of action potentials and (2) underlying the release of inhibitory transmitter onto the contralateral cell.

Cell surface domain specific postsynaptic currents evoked by identified GABAergic neurones in rat hippocampus in vitro

PubMed Central

Maccaferri, Gianmaria; David, J; Roberts, B; Szucs, Peter; Cottingham, Carol A; Somogyi, Peter

2000-01-01

Inhibitory postsynaptic currents (IPSCs) evoked in CA1 pyramidal cells (n = 46) by identified interneurones (n = 43) located in str. oriens were recorded in order to compare their functional properties and to determine the effect of synapse location on the apparent IPSC kinetics as recorded using somatic voltage clamp at −70 mV and nearly symmetrical [Cl−]. Five types of visualised presynaptic interneurone, oriens-lacunosum moleculare (O-LMC), basket (BC), axo-axonic (AAC), bistratified (BiC) and oriens-bistratified (O-BiC) cells, were distinguished by immunocytochemistry and/or synapse location using light and electron microscopy. Somatostatin immunoreactive O-LMCs, innervating the most distal dendritic shafts and spines, evoked the smallest amplitude (26 ± 10 pA, s.e.m., n = 8) and slowest IPSCs (10–90 % rise time, 6.2 ± 0.6 ms; decay, 20.8 ± 1.7 ms, n = 8), with no paired-pulse modulation of the second IPSC (93 ± 4 %) at 100 ms interspike interval. In contrast, parvalbumin-positive AACs evoked larger amplitude (308 ± 103 pA, n = 7) and kinetically faster (rise time, 0.8 ± 0.1 ms; decay 11.2 ± 0.9 ms, n = 7) IPSCs showing paired-pulse depression (to 68 ± 5 %, n = 6). Parvalbumin- or CCK-positive BCs (n = 9) terminating on soma/dendrites, BiCs (n = 4) and O-BiCs (n = 7) innervating dendrites evoked IPSCs with intermediate kinetic parameters. The properties of IPSCs and sensitivity to bicuculline indicated that they were mediated by GABAA receptors. In three cases, kinetically complex, multiphasic IPSCs, evoked by an action potential in the recorded basket cells, suggested that coupled interneurones, possibly through electrotonic junctions, converged on the same postsynaptic neurone. The population of O-BiCs (4 of 4 somatostatin positive) characterised in this study had horizontal dendrites restricted to str. oriens/alveus and innervated stratum radiatum and oriens. Other BiCs had radial dendrites as described earlier. The parameters of IPSCs evoked by BiCs and O-BiCs showed the largest cell to cell variation, and a single interneurone could evoke both small and slow as well as large and relatively fast IPSCs. The kinetic properties of the somatically recorded postsynaptic current are correlated with the innervated cell surface domain. A significant correlation of rise and decay times for the overall population of unitary IPSCs suggests that electrotonic filtering of distal responses is a major factor for the location and cell type specific differences of unitary IPSCs, but molecular heterogeneity of postsynaptic GABAA receptors may also contribute to the observed kinetic differences. Furthermore, domain specific differences in the short-term plasticity of the postsynaptic response indicate a differentiation of interneurones in activity-dependent responses. PMID:10747186

Endogenous cannabinoids mediate retrograde signals from depolarized postsynaptic neurons to presynaptic terminals.

PubMed

Ohno-Shosaku, T; Maejima, T; Kano, M

2001-03-01

Endogenous cannabinoids are considered to function as diffusible and short-lived modulators that may transmit signals retrogradely from postsynaptic to presynaptic neurons. To evaluate this possibility, we have made a paired whole-cell recording from cultured hippocampal neurons with inhibitory synaptic connections. In about 60% of pairs, a cannabinoid agonist greatly reduced the release of the inhibitory neurotransmitter GABA from presynaptic terminals. In most of such pairs but not in those insensitive to the agonist, depolarization of postsynaptic neurons and the resultant elevation of intracellular Ca2+ concentration caused transient suppression of inhibitory synaptic currents, which is mainly due to reduction of GABA release. This depolarization-induced suppression was completely blocked by selective cannabinoid antagonists. Our results reveal that endogenous cannabinoids mediate retrograde signals from depolarized postsynaptic neurons to presynaptic terminals to cause the reduction of transmitter release.

Parkin Deficiency Reduces Hippocampal Glutamatergic Neurotransmission by Impairing AMPA Receptor Endocytosis.

PubMed

Cortese, Giuseppe P; Zhu, Mei; Williams, Damian; Heath, Sarah; Waites, Clarissa L

2016-11-30

Mutations in the gene encoding Parkin, an E3 ubiquitin ligase, lead to juvenile-onset Parkinson's disease by inducing the selective death of midbrain dopaminergic neurons. Accumulating evidence indicates that Parkin also has an important role in excitatory glutamatergic neurotransmission, although its precise mechanism of action remains unclear. Here, we investigate Parkin's role at glutamatergic synapses of rat hippocampal neurons. We find that Parkin-deficient neurons exhibit significantly reduced AMPA receptor (AMPAR)-mediated currents and cell-surface expression, and that these phenotypes result from decreased postsynaptic expression of the adaptor protein Homer1, which is necessary for coupling AMPAR endocytic zones with the postsynaptic density. Accordingly, Parkin loss of function leads to the reduced density of postsynaptic endocytic zones and to impaired AMPAR internalization. These findings demonstrate a novel and essential role for Parkin in glutamatergic neurotransmission, as a stabilizer of postsynaptic Homer1 and the Homer1-linked endocytic machinery necessary for maintaining normal cell-surface AMPAR levels. Mutations in Parkin, a ubiquitinating enzyme, lead to the selective loss of midbrain dopaminergic neurons and juvenile-onset Parkinson's disease (PD). Parkin loss of function has also been shown to alter hippocampal glutamatergic neurotransmission, providing a potential explanation for PD-associated cognitive impairment. However, very little is known about Parkin's specific sites or mechanisms of action at glutamatergic synapses. Here, we show that Parkin deficiency leads to decreased AMPA receptor-mediated activity due to disruption of the postsynaptic endocytic zones required for maintaining proper cell-surface AMPA receptor levels. These findings demonstrate a novel role for Parkin in synaptic AMPA receptor internalization and suggest a Parkin-dependent mechanism for hippocampal dysfunction that may explain cognitive deficits associated with some forms of PD. Copyright © 2016 the authors 0270-6474/16/3612243-16$15.00/0.

Functional compatibility between Purkinje cell axon branches and their target neurons in the cerebellum.

PubMed

Yang, Zhilai; Chen, Na; Ge, Rongjing; Qian, Hao; Wang, Jin-Hui

2017-09-22

A neuron sprouts an axon, and its branches to innervate many target neurons that are divergent in their functions. In order to efficiently regulate the diversified cells, the axon branches should differentiate functionally to be compatible with their target neurons, i.e., a function compatibility between presynaptic and postsynaptic partners. We have examined this hypothesis by using electrophysiological method in the cerebellum, in which the main axon of Purkinje cell projected to deep nucleus cells and the recurrent axons innervated the adjacent Purkinje cells. The fidelity of spike propagation is superior in the recurrent branches than the main axon. The capabilities of encoding spikes and processing GABAergic inputs are advanced in Purkinje cells versus deep nucleus cells. The functional differences among Purkinje's axonal branches and their postsynaptic neurons are preset by the variable dynamics of their voltage-gated sodium channels. In addition, activity strengths between presynaptic and postsynaptic partners are proportionally correlated, i.e., active axonal branches innervate active target neurons, or vice versa. The physiological impact of the functional compatibility is to make the neurons in their circuits to be activated appropriately. In conclusion, each cerebellar Purkinje cell sprouts the differentiated axon branches to be compatible with the diversified target cells in their functions, in order to construct the homeostatic and efficient units for their coordinated activity in neural circuits.

Functional compatibility between Purkinje cell axon branches and their target neurons in the cerebellum

PubMed Central

Qian, Hao; Wang, Jin-Hui

2017-01-01

A neuron sprouts an axon, and its branches to innervate many target neurons that are divergent in their functions. In order to efficiently regulate the diversified cells, the axon branches should differentiate functionally to be compatible with their target neurons, i.e., a function compatibility between presynaptic and postsynaptic partners. We have examined this hypothesis by using electrophysiological method in the cerebellum, in which the main axon of Purkinje cell projected to deep nucleus cells and the recurrent axons innervated the adjacent Purkinje cells. The fidelity of spike propagation is superior in the recurrent branches than the main axon. The capabilities of encoding spikes and processing GABAergic inputs are advanced in Purkinje cells versus deep nucleus cells. The functional differences among Purkinje's axonal branches and their postsynaptic neurons are preset by the variable dynamics of their voltage-gated sodium channels. In addition, activity strengths between presynaptic and postsynaptic partners are proportionally correlated, i.e., active axonal branches innervate active target neurons, or vice versa. The physiological impact of the functional compatibility is to make the neurons in their circuits to be activated appropriately. In conclusion, each cerebellar Purkinje cell sprouts the differentiated axon branches to be compatible with the diversified target cells in their functions, in order to construct the homeostatic and efficient units for their coordinated activity in neural circuits. PMID:29069799

CDKL5 ensures excitatory synapse stability by reinforcing NGL-1-PSD95 interaction in the postsynaptic compartment and is impaired in patient iPSC-derived neurons.

PubMed

Ricciardi, Sara; Ungaro, Federica; Hambrock, Melanie; Rademacher, Nils; Stefanelli, Gilda; Brambilla, Dario; Sessa, Alessandro; Magagnotti, Cinzia; Bachi, Angela; Giarda, Elisa; Verpelli, Chiara; Kilstrup-Nielsen, Charlotte; Sala, Carlo; Kalscheuer, Vera M; Broccoli, Vania

2012-09-01

Mutations of the cyclin-dependent kinase-like 5 (CDKL5) and netrin-G1 (NTNG1) genes cause a severe neurodevelopmental disorder with clinical features that are closely related to Rett syndrome, including intellectual disability, early-onset intractable epilepsy and autism. We report here that CDKL5 is localized at excitatory synapses and contributes to correct dendritic spine structure and synapse activity. To exert this role, CDKL5 binds and phosphorylates the cell adhesion molecule NGL-1. This phosphorylation event ensures a stable association between NGL-1 and PSD95. Accordingly, phospho-mutant NGL-1 is unable to induce synaptic contacts whereas its phospho-mimetic form binds PSD95 more efficiently and partially rescues the CDKL5-specific spine defects. Interestingly, similarly to rodent neurons, iPSC-derived neurons from patients with CDKL5 mutations exhibit aberrant dendritic spines, thus suggesting a common function of CDKL5 in mice and humans.

Fine-scale topography in sensory systems: insights from Drosophila and vertebrates

PubMed Central

Kaneko, Takuya; Ye, Bing

2015-01-01

To encode the positions of sensory stimuli, sensory circuits form topographic maps in the central nervous system through specific point-to-point connections between pre- and post-synaptic neurons. In vertebrate visual systems, the establishment of topographic maps involves the formation of a coarse topography followed by that of fine-scale topography that distinguishes the axon terminals of neighboring neurons. It is known that intrinsic differences in the form of broad gradients of guidance molecules instruct coarse topography while neuronal activity is required for fine-scale topography. On the other hand, studies in the Drosophila visual system have shown that intrinsic differences in cell adhesion among the axon terminals of neighboring neurons instruct the fine-scale topography. Recent studies on activity-dependent topography in the Drosophila somatosensory system have revealed a role of neuronal activity in creating molecular differences among sensory neurons for establishing fine-scale topography, implicating a conserved principle. Here we review the findings in both Drosophila and vertebrates and propose an integrated model for fine-scale topography. PMID:26091779

Fine-scale topography in sensory systems: insights from Drosophila and vertebrates.

PubMed

Kaneko, Takuya; Ye, Bing

2015-09-01

To encode the positions of sensory stimuli, sensory circuits form topographic maps in the central nervous system through specific point-to-point connections between pre- and postsynaptic neurons. In vertebrate visual systems, the establishment of topographic maps involves the formation of a coarse topography followed by that of fine-scale topography that distinguishes the axon terminals of neighboring neurons. It is known that intrinsic differences in the form of broad gradients of guidance molecules instruct coarse topography while neuronal activity is required for fine-scale topography. On the other hand, studies in the Drosophila visual system have shown that intrinsic differences in cell adhesion among the axon terminals of neighboring neurons instruct the fine-scale topography. Recent studies on activity-dependent topography in the Drosophila somatosensory system have revealed a role of neuronal activity in creating molecular differences among sensory neurons for establishing fine-scale topography, implicating a conserved principle. Here we review the findings in both Drosophila and vertebrates and propose an integrated model for fine-scale topography.

Molecular Dissection of Neuroligin 2 and Slitrk3 Reveals an Essential Framework for GABAergic Synapse Development.

PubMed

Li, Jun; Han, Wenyan; Pelkey, Kenneth A; Duan, Jingjing; Mao, Xia; Wang, Ya-Xian; Craig, Michael T; Dong, Lijin; Petralia, Ronald S; McBain, Chris J; Lu, Wei

2017-11-15

In the brain, many types of interneurons make functionally diverse inhibitory synapses onto principal neurons. Although numerous molecules have been identified to function in inhibitory synapse development, it remains unknown whether there is a unifying mechanism for development of diverse inhibitory synapses. Here we report a general molecular mechanism underlying hippocampal inhibitory synapse development. In developing neurons, the establishment of GABAergic transmission depends on Neuroligin 2 (NL2), a synaptic cell adhesion molecule (CAM). During maturation, inhibitory synapse development requires both NL2 and Slitrk3 (ST3), another CAM. Importantly, NL2 and ST3 interact with nanomolar affinity through their extracellular domains to synergistically promote synapse development. Selective perturbation of the NL2-ST3 interaction impairs inhibitory synapse development with consequent disruptions in hippocampal network activity and increased seizure susceptibility. Our findings reveal how unique postsynaptic CAMs work in concert to control synaptogenesis and establish a general framework for GABAergic synapse development. Published by Elsevier Inc.

Distinct roles for extracellular and intracellular domains in neuroligin function at inhibitory synapses.

PubMed

Nguyen, Quynh-Anh; Horn, Meryl E; Nicoll, Roger A

2016-11-02

Neuroligins (NLGNs) are postsynaptic cell adhesion molecules that interact trans-synaptically with neurexins to mediate synapse development and function. NLGN2 is only at inhibitory synapses while NLGN3 is at both excitatory and inhibitory synapses. We found that NLGN3 function at inhibitory synapses in rat CA1 depends on the presence of NLGN2 and identified a domain in the extracellular region that accounted for this functional difference between NLGN2 and 3 specifically at inhibitory synapses. We further show that the presence of a cytoplasmic tail (c-tail) is indispensible, and identified two domains in the c-tail that are necessary for NLGN function at inhibitory synapses. These domains point to a gephyrin-dependent mechanism that is disrupted by an autism-associated mutation at R705 and a gephyrin-independent mechanism reliant on a putative phosphorylation site at S714. Our work highlights unique and separate roles for the extracellular and intracellular regions in specifying and carrying out NLGN function respectively.

Balance within the Neurexin Trans-Synaptic Connexus Stabilizes Behavioral Control

PubMed Central

Clarke, Raymond A.; Eapen, Valsamma

2014-01-01

Autism spectrum disorder (ASD) is characterized by a broad spectrum of behavioral deficits of unknown etiology. ASD associated mutations implicate numerous neurological pathways including a common association with the neurexin trans-synaptic connexus (NTSC) which regulates neuronal cell-adhesion, neuronal circuitry, and neurotransmission. Comparable DNA lesions affecting the NTSC, however, associate with a diversity of behavioral deficits within and without the autism spectrum including a very strong association with Tourette syndrome. The NTSC is comprised of numerous post-synaptic ligands competing for trans-synaptic connection with one of the many different neurexin receptors yet no apparent association exists between specific NTSC molecules/complexes and specific behavioral deficits. Together these findings indicate a fundamental role for NTSC-balance in stabilizing pre-behavioral control. Further molecular and clinical characterization and stratification of ASD and TS on the basis of NTSC status will help elucidate the molecular basis of behavior – and define how the NTSC functions in combination with other molecular determinates to strengthen behavioral control and specify behavioral deficits. PMID:24578685

Synaptically evoked Ca2+ release from intracellular stores is not influenced by vesicular zinc in CA3 hippocampal pyramidal neurones.

PubMed

Evstratova, Alesya; Tóth, Katalin

2011-12-01

The co-release of neuromodulatory substances in combination with classic neurotransmitters such as glutamate and GABA from individual presynaptic nerve terminals has the capacity to dramatically influence synaptic efficacy and plasticity. At hippocampal mossy fibre synapses vesicular zinc is suggested to serve as a cotransmitter capable of regulating calcium release from internal stores in postsynaptic CA3 pyramidal cells. Here we investigated this possibility using combined intracellular ratiometric calcium imaging and patch-clamp recording techniques. In acute hippocampal slices a brief train of mossy fibre stimulation produced a large, delayed postsynaptic Ca(2+) wave that was spatially restricted to the proximal apical dendrites of CA3 pyramidal cells within stratum lucidum. This calcium increase was sensitive to intracellularly applied heparin indicating reliance upon release from internal stores and was triggered by activation of both group I metabotropic glutamate and NMDA receptors. Importantly, treatment of slices with the membrane-impermeant zinc chelator CaEDTA did not influence the synaptically evoked postsynaptic Ca(2+) waves. Moreover, mossy fibre stimulus evoked postsynaptic Ca(2+) signals were not significantly different between wild-type and zinc transporter 3 (ZnT3) knock-out animals. Considered together our data do not support a role for vesicular zinc in regulating mossy fibre evoked Ca(2+) release from CA3 pyramidal cell internal stores.

Synaptically evoked Ca2+ release from intracellular stores is not influenced by vesicular zinc in CA3 hippocampal pyramidal neurones

PubMed Central

Evstratova, Alesya; Tóth, Katalin

2011-01-01

Abstract The co-release of neuromodulatory substances in combination with classic neurotransmitters such as glutamate and GABA from individual presynaptic nerve terminals has the capacity to dramatically influence synaptic efficacy and plasticity. At hippocampal mossy fibre synapses vesicular zinc is suggested to serve as a cotransmitter capable of regulating calcium release from internal stores in postsynaptic CA3 pyramidal cells. Here we investigated this possibility using combined intracellular ratiometric calcium imaging and patch-clamp recording techniques. In acute hippocampal slices a brief train of mossy fibre stimulation produced a large, delayed postsynaptic Ca2+ wave that was spatially restricted to the proximal apical dendrites of CA3 pyramidal cells within stratum lucidum. This calcium increase was sensitive to intracellularly applied heparin indicating reliance upon release from internal stores and was triggered by activation of both group I metabotropic glutamate and NMDA receptors. Importantly, treatment of slices with the membrane-impermeant zinc chelator CaEDTA did not influence the synaptically evoked postsynaptic Ca2+ waves. Moreover, mossy fibre stimulus evoked postsynaptic Ca2+ signals were not significantly different between wild-type and zinc transporter 3 (ZnT3) knock-out animals. Considered together our data do not support a role for vesicular zinc in regulating mossy fibre evoked Ca2+ release from CA3 pyramidal cell internal stores. PMID:21986206

Synaptic plasticity in a cerebellum-like structure depends on temporal order

NASA Astrophysics Data System (ADS)

Bell, Curtis C.; Han, Victor Z.; Sugawara, Yoshiko; Grant, Kirsty

1997-05-01

Cerebellum-like structures in fish appear to act as adaptive sensory processors, in which learned predictions about sensory input are generated and subtracted from actual sensory input, allowing unpredicted inputs to stand out1-3. Pairing sensory input with centrally originating predictive signals, such as corollary discharge signals linked to motor commands, results in neural responses to the predictive signals alone that are Negative images' of the previously paired sensory responses. Adding these 'negative images' to actual sensory inputs minimizes the neural response to predictable sensory features. At the cellular level, sensory input is relayed to the basal region of Purkinje-like cells, whereas predictive signals are relayed by parallel fibres to the apical dendrites of the same cells4. The generation of negative images could be explained by plasticity at parallel fibre synapses5-7. We show here that such plasticity exists in the electrosensory lobe of mormyrid electric fish and that it has the necessary properties for such a model: it is reversible, anti-hebbian (excitatory postsynaptic potentials (EPSPs) are depressed after pairing with a postsynaptic spike) and tightly dependent on the sequence of pre- and postsynaptic events, with depression occurring only if the postsynaptic spike follows EPSP onset within 60 ms.

Postsynaptic activity reverses the sign of the acetylcholine-induced long-term plasticity of GABAA inhibition

PubMed Central

Domínguez, Soledad; Fernández de Sevilla, David; Buño, Washington

2014-01-01

Acetylcholine (ACh) regulates forms of plasticity that control cognitive functions but the underlying mechanisms remain largely unknown. ACh controls the intrinsic excitability, as well as the synaptic excitation and inhibition of CA1 hippocampal pyramidal cells (PCs), cells known to participate in circuits involved in cognition and spatial navigation. However, how ACh regulates inhibition in function of postsynaptic activity has not been well studied. Here we show that in rat PCs, a brief pulse of ACh or a brief stimulation of cholinergic septal fibers combined with repeated depolarization induces strong long-term enhancement of GABAA inhibition (GABAA-LTP). Indeed, this enhanced inhibition is due to the increased activation of α5βγ2 subunit-containing GABAA receptors by the GABA released. GABAA-LTP requires the activation of M1-muscarinic receptors and an increase in cytosolic Ca2+. In the absence of PC depolarization ACh triggered a presynaptic depolarization-induced suppression of inhibition (DSI), revealing that postsynaptic activity gates the effects of ACh from presynaptic DSI to postsynaptic LTP. These results provide key insights into mechanisms potentially linked with cognitive functions, spatial navigation, and the homeostatic control of abnormal hyperexcitable states. PMID:24938789

Parvalbumin-expressing interneurons can act solo while somatostatin-expressing interneurons act in chorus in most cases on cortical pyramidal cells.

PubMed

Safari, Mir-Shahram; Mirnajafi-Zadeh, Javad; Hioki, Hiroyuki; Tsumoto, Tadaharu

2017-10-06

Neural circuits in the cerebral cortex consist primarily of excitatory pyramidal (Pyr) cells and inhibitory interneurons. Interneurons are divided into several subtypes, in which the two major groups are those expressing parvalbumin (PV) or somatostatin (SOM). These subtypes of interneurons are reported to play distinct roles in tuning and/or gain of visual response of pyramidal cells in the visual cortex. It remains unclear whether there is any quantitative and functional difference between the PV → Pyr and SOM → Pyr connections. We compared unitary inhibitory postsynaptic currents (uIPSCs) evoked by electrophysiological activation of single presynaptic interneurons with population IPSCs evoked by photo-activation of a mass of interneurons in vivo and in vitro in transgenic mice in which PV or SOM neurons expressed channelrhodopsin-2, and found that at least about 14 PV neurons made strong connections with a postsynaptic Pyr cell while a much larger number of SOM neurons made weak connections. Activation or suppression of single PV neurons modified visual responses of postsynaptic Pyr cells in 6 of 7 pairs whereas that of single SOM neurons showed no significant modification in 8 of 11 pairs, suggesting that PV neurons can act solo whereas most of SOM neurons may act in chorus on Pyr cells.

Postsynaptic elevation of calcium induces persistent depression of developing neuromuscular synapses.

PubMed

Cash, S; Dan, Y; Poo, M M; Zucker, R

1996-04-01

Synaptic activity is known to modulate neuronal connectivity in the nervous system. At developing Xenopus neuromuscular synapses in culture, repetitive postsynaptic application of ACh near the synapse leads to immediate and persistent synaptic depression, which was shown to be caused by reduction of presynaptic evoked transmitter release. However, little depression was found when ACh was applied to the muscle 20 microns or further from the synapse. Fluorescence imaging of cytosolic Ca2+ ([Ca2+]i) showed that each ACh pulse induced a transient elevation of myocyte [Ca2+]i that spread approximately 20 microns. Local photoactivated release of Ca2+ from the caged Ca2+ chelators nitr-5 or nitrophen in the postsynaptic cell was sufficient to induce persistent synaptic depression. These results support a model in which localized Ca2+ influx into the postsynaptic myocyte initiates transsynaptic retrograde modulation of presynaptic secretion mechanisms.

Not GABA but glycine mediates segmental, propriospinal, and bulbospinal postsynaptic inhibition in adult mouse spinal forelimb motor neurons.

PubMed

Jiang, Juan; Alstermark, Bror

2015-02-04

The general view is that both glycine (Eccles, 1964) and GABA (Curtis and Felix, 1971) evoke postsynaptic inhibition in spinal motor neurons. In newborn or juvenile animals, there are conflicting results showing postsynaptic inhibition in motor neurons by corelease of GABA and glycine (Jonas et al., 1998) or by glycine alone (Bhumbra et al., 2012). To resolve the relative contributions of GABA and glycine to postsynaptic inhibition, we performed in vivo intracellular recordings from forelimb motor neurons in adult mice. Postsynaptic potentials evoked from segmental, propriospinal, and bulbospinal systems in motor neurons were compared across four different conditions: control, after gabazine, gabazine followed by strychnine, and strychnine alone. No significant differences were observed in the proportion of IPSPs and EPSPs between control and gabazine conditions. In contrast, EPSPs but not IPSPs were recorded after adding strychnine with gabazine or administering strychnine alone, suggesting an exclusive role for glycine in postsynaptic inhibition. To test whether the injected (intraperitoneal) dose of gabazine blocked GABAergic inhibitory transmission, we evoked GABAA receptor-mediated monosynaptic IPSPs in deep cerebellar nuclei neurons by stimulation of Purkinje cell fibers. No monosynaptic IPSPs could be recorded in the presence of gabazine, showing the efficacy of gabazine treatment. Our results demonstrate that, in the intact adult mouse, the postsynaptic inhibitory effects in spinal motor neurons exerted by three different systems, intrasegmental and intersegmental as well as supraspinal, are exclusively glycinergic. These findings emphasize the importance of glycinergic postsynaptic inhibition in motor neurons and challenge the view that GABA also contributes. Copyright © 2015 the authors 0270-6474/15/351991-08$15.00/0.

Neuromuscular synapse integrity requires linkage of acetylcholine receptors to postsynaptic intermediate filament networks via rapsyn–plectin 1f complexes

PubMed Central

Mihailovska, Eva; Raith, Marianne; Valencia, Rocio G.; Fischer, Irmgard; Banchaabouchi, Mumna Al; Herbst, Ruth; Wiche, Gerhard

2014-01-01

Mutations in the cytolinker protein plectin lead to grossly distorted morphology of neuromuscular junctions (NMJs) in patients suffering from epidermolysis bullosa simplex (EBS)-muscular dystrophy (MS) with myasthenic syndrome (MyS). Here we investigated whether plectin contributes to the structural integrity of NMJs by linking them to the postsynaptic intermediate filament (IF) network. Live imaging of acetylcholine receptors (AChRs) in cultured myotubes differentiated ex vivo from immortalized plectin-deficient myoblasts revealed them to be highly mobile and unable to coalesce into stable clusters, in contrast to wild-type cells. We found plectin isoform 1f (P1f) to bridge AChRs and IFs via direct interaction with the AChR-scaffolding protein rapsyn in an isoform-specific manner; forced expression of P1f in plectin-deficient cells rescued both compromised AChR clustering and IF network anchoring. In conditional plectin knockout mice with gene disruption in muscle precursor/satellite cells (Pax7-Cre/cKO), uncoupling of AChRs from IFs was shown to lead to loss of postsynaptic membrane infoldings and disorganization of the NMJ microenvironment, including its invasion by microtubules. In their phenotypic behavior, mutant mice closely mimicked EBS-MD-MyS patients, including impaired body balance, severe muscle weakness, and reduced life span. Our study demonstrates that linkage to desmin IF networks via plectin is crucial for formation and maintenance of AChR clusters, postsynaptic NMJ organization, and body locomotion. PMID:25318670

Cell type dependence and variability in the short-term plasticity of EPSCs in identified mouse hippocampal interneurones

PubMed Central

Losonczy, Attila; Zhang, Limei; Shigemoto, Ryuichi; Somogyi, Peter; Nusser, Zoltan

2002-01-01

Synapses exhibit different short-term plasticity patterns and this behaviour influences information processing in neuronal networks. We tested how the short-term plasticity of excitatory postsynaptic currents (EPSCs) depends on the postsynaptic cell type, identified by axonal arborizations and molecular markers in the hippocampal CA1 area. Three distinct types of short-term synaptic behaviour (facilitating, depressing and combined facilitating–depressing) were defined by fitting a dynamic neurotransmission model to the data. Approximately 75 % of the oriens-lacunosum-moleculare (O-LM) interneurones received facilitating EPSCs, but in three of 12 O-LM cells EPSCs also showed significant depression. Over 90 % of the O-LM cells were immunopositive for somatostatin and mGluR1α and all tested cells were decorated by strongly mGluR7a positive axon terminals. Responses in eight of 12 basket cells were described well with a model involving only depression, but the other cells displayed combined facilitating–depressing EPSCs. No apparent difference was found between the plasticity of EPSCs in cholecystokinin- or parvalbumin-containing basket cells. In oriens-bistratified cells (O-Bi), two of nine cells showed facilitating EPSCs, another two depressing, and the remaining five cells combined facilitating–depressing EPSCs. Seven of 10 cells tested for somatostatin were immunopositive, but mGluR1α was detectable only in two of 11 tested cells. Furthermore, most O-Bi cells projected to the CA3 area and the subiculum, as well as outside the hippocampal formation. Postsynaptic responses to action potentials recorded in vivo from a CA1 place cell were modelled, and revealed great differences between and within cell types. Our results demonstrate that the short-term plasticity of EPSCs is cell type dependent, but with significant heterogeneity within all three interneurone populations. PMID:12096061

[Postsynaptic reactions of cerebral cortex neurons, activated by nociceptive afferents during stimulation of the Raphe nuclei].

PubMed

Labakhua, T Sh; Dzhanashiia, T K; Gedevanishvili, G I; Dzhokhadze, L D; Tkemaladze, T T; Abzianidze, I V

2012-01-01

On cats, we studied the influence of stimulation of the Raphe nuclei (RN) on postsynaptic processes evoked in neurons of the somatosensory cortex by stimulation of nociceptive (intensive stimulation of the tooth pulp) and non-nociceptive (moderate stimulation of the ventroposteromedial--VPN--nucleus of the thalamus) afferent inputs. 6 cells, selectively excited by stimulation of nocciceptors and 9 cells, activated by both the above nociceptive and non-nociceptive influences (nociceptive and convergent neurons, respectively) were recorded intracellular. In neurons of both groups, responses to nociceptive stimulation (of sufficient intensity) looked like an EPSP-spike-IPSP (the letter of significant duration, up to 200-300 ms) compleх. Conditioning stimulation of the RN which preceded test stimulus applied to the tooth pulp or VPM nucleus by 100 to 800 ms, induced 40-60 % decrease of the IPSP amplitude only, while maхimal effect of influence, in both cases, was noted within intervals of 300-800 ms between conditioning and test stimulus. During stimulation of the RN, serotonin released via receptor and second messengers, provides postsynaptic modulation of GABAergic system, decreasing the IPSP amplitude which occurs after stimulation of both the tooth pulp and VPM thalamic nucleus. This process may be realized trough either pre- or postsynaptic mechanisms.

The potential role of postsynaptic phospholipase C activity in synaptic facilitation and behavioral sensitization in Aplysia.

PubMed

Fulton, Daniel; Condro, Michael C; Pearce, Kaycey; Glanzman, David L

2008-07-01

Previous findings indicate that synaptic facilitation, a cellular mechanism underlying sensitization of the siphon withdrawal response (SWR) in Aplysia, depends on a cascade of postsynaptic events, including activation of inositol triphosphate (IP3) receptors and release of Ca2+ from postsynaptic intracellular stores. These findings suggest that phospholipase C (PLC), the enzyme that catalyzes IP3 formation, may play an important role in postsynaptic signaling during facilitation and learning in Aplysia. Using the PLC inhibitor U73122, we found that PLC activity is required for synaptic facilitation following a 10-min treatment with 5-HT, as measured at 20 min after 5-HT washout. Prior work has indicated that facilitation at this time is supported primarily by postsynaptic processes. To determine whether postsynaptic PLC activity is involved in 5-HT-mediated facilitatory actions, we examined the effect of U73122 on enhancement of the response of motor neurons isolated in cell culture to glutamate, the sensory neuron transmitter. A 10-min application of 5-HT induced persistent (>40 min) enhancement of glutamate-evoked potentials (Glu-EPs) recorded from isolated motor neurons, and this enhancement was blocked by U73122. Finally, we showed that injecting U73122 into intact animals before behavioral training impaired intermediate-term sensitization, indicating that PLC activity contributes to this form of nonassociative learning.

AMPA receptor activation controls type I metabotropic glutamate receptor signalling via a tyrosine kinase at parallel fibre-Purkinje cell synapses.

PubMed

Auger, Céline; Ogden, David

2010-08-15

Metabotropic glutamate receptors type 1 (mGluR1s) and ionotropic AMPA receptors (AMPARs) are colocalized at parallel fibre (PF) to Purkinje cell synapses of the cerebellum. Single stimulation of PFs activates fast AMPAR excitatory postsynaptic currents, whereas the activation of mGluR1s requires burst stimulation. mGluR1s signal through several pathways in Purkinje cells and the most prominent is the activation of a slow EPSC (sEPSC). To separate the two synaptic currents, studies of the sEPSC have commonly been performed in the presence of AMPA/KA receptor antagonists. We show here in rat cerebellar slices that inhibition of the fast EPSC by AMPAR antagonists strongly and selectively potentiates the mGluR1 sEPSC, showing a negative regulation of mGluR1 by AMPAR. This effect is observed with low concentrations of NBQX (300 nM to 1 microM), with the selective AMPAR antagonist GYKI 53655 and also with gamma-DGG, a low affinity glutamate receptor antagonist. When photorelease of glutamate from MNI-glutamate was used to study the postsynaptic responses in isolation, AMPAR inhibition produced a similar potentiation of the mGluR1 sEPSC, showing that the interaction is postsynaptic. Finally, perfusion of the postsynaptic cell with PP1, an inhibitor of src-family tyrosine kinase, increased the amplitude of the mGluR1 sEPSC and occluded the effect of AMPAR inhibition. Thus, at PF to Purkinje cell synapses, AMPAR activation inhibits the mGluR1 sEPSC via activation of a src-family tyrosine kinase. Consequently mGluR1 signalling will be more sensitive to spillover of glutamate than to local synaptic release. Furthermore, it will be enhanced at silent PF synapses which are the majority in Purkinje cells.

Cell Shape Change by Drebrin.

PubMed

Hayashi, Kensuke

2017-01-01

Drebrin is localized in actin-rich regions of neuronal and non-neuronal cells. In mature neurons, its localization is strictly restricted to the postsynaptic sites. In order to understand the function of drebrin in cells, many studies have been performed to examine the effect of overexpression or knocking down of drebrin in various cell types, including neurons, myoblasts, kidney cells, and intestinal epithelial cells. In most cases alteration of cell shape and impairment or facilitation of actin-based activities of these cells were observed. Interestingly, overexpression of drebrin in matured neurons results in the alteration in dendritic spine morphology. Further studies have shown alteration in the localization of postsynaptic receptors and even changes in synaptic transmission caused by drebrin overexpression or depletion in neurons. These drebrin's effects are thought to come from drebrin's actin-cross-linking activity or competitive binding to actin against tropomyosin, fascin, and α-actinin. Furthermore, drebrin binds to various molecules, such as homer, EB3, and cell-cell junctional proteins, indicating that drebrin is a multifunctional cytoskeletal regulator.

The Septate Junction Protein Caspr is Required for Structural Support and Retention of KCNQ4 at Calyceal Synapses of Vestibular Hair Cells

PubMed Central

Sousa, Aurea D.; Andrade, Leonardo R.; Salles, Felipe T.; Pillai, Anilkumar M.; Buttermore, Elizabeth; Bhat, Manzoor A.; Kachar, Bechara

2009-01-01

The afferent innervation contacting the type I hair cells of the vestibular sensory epithelia form distinct calyceal synapses. The apposed pre- and post-synaptic membranes at this large area of synaptic contact are kept at a remarkably regular distance. Here, we show by freeze-fracture electron microscopy that a patterned alignment of proteins at the calyceal membrane resembles a type of intercellular junction that is rare in vertebrates, the septate junction (SJ). We found that a core molecular component of SJs, Caspr, colocalizes with the K+ channel KCNQ4 at the post-synaptic membranes of these calyceal synapses. Immunolabeling and ultrastructural analyses of Caspr knockout mice reveal that, in the absence of Caspr, the separation between the membranes of the hair cells and the afferent neurons is conspicuously irregular and often increased by an order of magnitude. In these mutants, KCNQ4 fails to cluster at the post-synaptic membrane and appears diffused along the entire calyceal membrane. Our results indicate that a septate-like junction provides structural support to calyceal synaptic contact with the vestibular hair cell, and that Caspr is required for the recruitment or retention of KCNQ4 at these synapses. PMID:19279247

Innervation regulates synaptic ribbons in lateral line mechanosensory hair cells

PubMed Central

Pujol, Remy; Cunningham, Dale E.; Hailey, Dale W.; Prendergast, Andrew; Rubel, Edwin W.; Raible, David W.

2016-01-01

ABSTRACT Failure to form proper synapses in mechanosensory hair cells, the sensory cells responsible for hearing and balance, leads to deafness and balance disorders. Ribbons are electron-dense structures that tether synaptic vesicles to the presynaptic zone of mechanosensory hair cells where they are juxtaposed with the post-synaptic endings of afferent fibers. They are initially formed throughout the cytoplasm, and, as cells mature, ribbons translocate to the basolateral membrane of hair cells to form functional synapses. We have examined the effect of post-synaptic elements on ribbon formation and maintenance in the zebrafish lateral line system by observing mutants that lack hair cell innervation, wild-type larvae whose nerves have been transected and ribbons in regenerating hair cells. Our results demonstrate that innervation is not required for initial ribbon formation but suggest that it is crucial for regulating the number, size and localization of ribbons in maturing hair cells, and for ribbon maintenance at the mature synapse. PMID:27103160

Innervation regulates synaptic ribbons in lateral line mechanosensory hair cells.

PubMed

Suli, Arminda; Pujol, Remy; Cunningham, Dale E; Hailey, Dale W; Prendergast, Andrew; Rubel, Edwin W; Raible, David W

2016-06-01

Failure to form proper synapses in mechanosensory hair cells, the sensory cells responsible for hearing and balance, leads to deafness and balance disorders. Ribbons are electron-dense structures that tether synaptic vesicles to the presynaptic zone of mechanosensory hair cells where they are juxtaposed with the post-synaptic endings of afferent fibers. They are initially formed throughout the cytoplasm, and, as cells mature, ribbons translocate to the basolateral membrane of hair cells to form functional synapses. We have examined the effect of post-synaptic elements on ribbon formation and maintenance in the zebrafish lateral line system by observing mutants that lack hair cell innervation, wild-type larvae whose nerves have been transected and ribbons in regenerating hair cells. Our results demonstrate that innervation is not required for initial ribbon formation but suggest that it is crucial for regulating the number, size and localization of ribbons in maturing hair cells, and for ribbon maintenance at the mature synapse. © 2016. Published by The Company of Biologists Ltd.

Communication networks in the brain: neurons, receptors, neurotransmitters, and alcohol.

PubMed

Lovinger, David M

2008-01-01

Nerve cells (i.e., neurons) communicate via a combination of electrical and chemical signals. Within the neuron, electrical signals driven by charged particles allow rapid conduction from one end of the cell to the other. Communication between neurons occurs at tiny gaps called synapses, where specialized parts of the two cells (i.e., the presynaptic and postsynaptic neurons) come within nanometers of one another to allow for chemical transmission. The presynaptic neuron releases a chemical (i.e., a neurotransmitter) that is received by the postsynaptic neuron's specialized proteins called neurotransmitter receptors. The neurotransmitter molecules bind to the receptor proteins and alter postsynaptic neuronal function. Two types of neurotransmitter receptors exist-ligand-gated ion channels, which permit rapid ion flow directly across the outer cell membrane, and G-protein-coupled receptors, which set into motion chemical signaling events within the cell. Hundreds of molecules are known to act as neurotransmitters in the brain. Neuronal development and function also are affected by peptides known as neurotrophins and by steroid hormones. This article reviews the chemical nature, neuronal actions, receptor subtypes, and therapeutic roles of several transmitters, neurotrophins, and hormones. It focuses on neurotransmitters with important roles in acute and chronic alcohol effects on the brain, such as those that contribute to intoxication, tolerance, dependence, and neurotoxicity, as well as maintained alcohol drinking and addiction.

Role of GABAA-Mediated Inhibition and Functional Assortment of Synapses onto Individual Layer 4 Neurons in Regulating Plasticity Expression in Visual Cortex.

PubMed

Saez, Ignacio; Friedlander, Michael J

2016-01-01

Layer 4 (L4) of primary visual cortex (V1) is the main recipient of thalamocortical fibers from the dorsal lateral geniculate nucleus (LGNd). Thus, it is considered the main entry point of visual information into the neocortex and the first anatomical opportunity for intracortical visual processing before information leaves L4 and reaches supra- and infragranular cortical layers. The strength of monosynaptic connections from individual L4 excitatory cells onto adjacent L4 cells (unitary connections) is highly malleable, demonstrating that the initial stage of intracortical synaptic transmission of thalamocortical information can be altered by previous activity. However, the inhibitory network within L4 of V1 may act as an internal gate for induction of excitatory synaptic plasticity, thus providing either high fidelity throughput to supragranular layers or transmittal of a modified signal subject to recent activity-dependent plasticity. To evaluate this possibility, we compared the induction of synaptic plasticity using classical extracellular stimulation protocols that recruit a combination of excitatory and inhibitory synapses with stimulation of a single excitatory neuron onto a L4 cell. In order to induce plasticity, we paired pre- and postsynaptic activity (with the onset of postsynaptic spiking leading the presynaptic activation by 10ms) using extracellular stimulation (ECS) in acute slices of primary visual cortex and comparing the outcomes with our previously published results in which an identical protocol was used to induce synaptic plasticity between individual pre- and postsynaptic L4 excitatory neurons. Our results indicate that pairing of ECS with spiking in a L4 neuron fails to induce plasticity in L4-L4 connections if synaptic inhibition is intact. However, application of a similar pairing protocol under GABAARs inhibition by bath application of 2μM bicuculline does induce robust synaptic plasticity, long term potentiation (LTP) or long term depression (LTD), similar to our results with pairing of pre- and postsynaptic activation between individual excitatory L4 neurons in which inhibitory connections are not activated. These results are consistent with the well-established observation that inhibition limits the capacity for induction of plasticity at excitatory synapses and that pre- and postsynaptic activation at a fixed time interval can result in a variable range of plasticity outcomes. However, in the current study by virtue of having two sets of experimental data, we have provided a new insight into these processes. By randomly mixing the assorting of individual L4 neurons according to the frequency distribution of the experimentally determined plasticity outcome distribution based on the calculated convergence of multiple individual L4 neurons onto a single postsynaptic L4 neuron, we were able to compare then actual ECS plasticity outcomes to those predicted by randomly mixing individual pairs of neurons. Interestingly, the observed plasticity profiles with ECS cannot account for the random assortment of plasticity behaviors of synaptic connections between individual cell pairs. These results suggest that connections impinging onto a single postsynaptic cell may be grouped according to plasticity states.

Expression of the postsynaptic scaffold PSD-95 and development of synaptic physiology during giant terminal formation in the auditory brainstem of the chicken.

PubMed

Goyer, David; Fensky, Luisa; Hilverling, Anna Maria; Kurth, Stefanie; Kuenzel, Thomas

2015-05-01

In the avian nucleus magnocellularis (NM) endbulb of Held giant synapses develop from temporary bouton terminals. The molecular regulation of this process is not well understood. Furthermore, it is unknown how the postsynaptic specialization of the endbulb synapses develops. We therefore analysed expression of the postsynaptic scaffold protein PSD-95 during the transition from bouton-to-endbulb synapses. PSD-95 has been implicated in the regulation of the strength of glutamatergic synapses and could accordingly be of functional relevance for giant synapse formation. PSD-95 protein was expressed at synaptic sites in embryonic chicken auditory brainstem and upregulated between embryonic days (E)12 and E16. We applied immunofluorescence staining and confocal microscopy to quantify pre-and postsynaptic protein signals during bouton-to-endbulb transition. Giant terminal formation progressed along the tonotopic axis in NM, but was absent in low-frequency NM. We found a tonotopic gradient of postsynaptic PSD-95 signals in NM. Furthermore, PSD-95 immunosignals showed the greatest increase between E12 and E15, temporally preceding the bouton-to-endbulb transition. We then applied whole-cell electrophysiology to measure synaptic currents elicited by synaptic terminals during bouton-to-endbulb transition. With progressing endbulb formation postsynaptic currents rose more rapidly and synapses were less susceptible to short-term depression, but currents were not different in amplitude or decay-time constant. We conclude that development of presynaptic specializations follows postsynaptic development and speculate that the early PSD-95 increase could play a functional role in endbulb formation. © 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Homeostatic plasticity shapes cell-type-specific wiring in the retina

PubMed Central

Tien, Nai-Wen; Soto, Florentina; Kerschensteiner, Daniel

2017-01-01

SUMMARY Convergent input from different presynaptic partners shapes the responses of postsynaptic neurons. Whether developing postsynaptic neurons establish connections with each presynaptic partner independently, or balance inputs to attain specific responses is unclear. Retinal ganglion cells (RGCs) receive convergent input from bipolar cell types with different contrast responses and temporal tuning. Here, using optogenetic activation and pharmacogenetic silencing, we found that type 6 bipolar cells (B6) dominate excitatory input to ONα-RGCs. We generated mice in which B6 cells were selectively removed from developing circuits (B6-DTA). In B6-DTA mice, ONα-RGCs adjusted connectivity with other bipolar cells in a cell-type-specific manner. They recruited new partners, increased synapses with some existing partners, and maintained constant input from others. Patch clamp recordings revealed that anatomical rewiring precisely preserved contrast- and temporal frequency response functions of ONα-RGCs, indicating that homeostatic plasticity shapes cell-type-specific wiring in the developing retina to stabilize visual information sent to the brain. PMID:28457596

Molecular Mechanism of Active Zone Organization at Vertebrate Neuromuscular Junctions

PubMed Central

Nishimune, Hiroshi

2013-01-01

Organization of presynaptic active zones is essential for development, plasticity, and pathology of the nervous system. Recent studies indicate a trans-synaptic molecular mechanism that organizes the active zones by connecting the pre- and the postsynaptic specialization. The presynaptic component of this trans-synaptic mechanism is comprised of cytosolic active zone proteins bound to the cytosolic domains of voltage-dependent calcium channels (P/Q-, N-, and L-type) on the presynaptic membrane. The postsynaptic component of this mechanism is the synapse organizer (laminin β2) that is expressed by the postsynaptic cell and accumulates specifically on top of the postsynaptic specialization. The pre- and the postsynaptic components interact directly between the extracellular domains of calcium channels and laminin β2 to anchor the presynaptic protein complex in front of the postsynaptic specialization. Hence, the presynaptic calcium channel functions as a scaffolding protein for active zone organization and as an ion-conducting channel for synaptic transmission. In contrast to the requirement of calcium influx for synaptic transmission, the formation of the active zone does not require the calcium influx through the calcium channels. Importantly, the active zones of adult synapses are not stable structures and require maintenance for their integrity. Furthermore, aging or diseases of the central and peripheral nervous system impair the active zones. This review will focus on the molecular mechanisms that organize the presynaptic active zones and summarize recent findings at the neuromuscular junctions and other synapses. PMID:22135013

Mapping cell surface adhesion by rotation tracking and adhesion footprinting

NASA Astrophysics Data System (ADS)

Li, Isaac T. S.; Ha, Taekjip; Chemla, Yann R.

2017-03-01

Rolling adhesion, in which cells passively roll along surfaces under shear flow, is a critical process involved in inflammatory responses and cancer metastasis. Surface adhesion properties regulated by adhesion receptors and membrane tethers are critical in understanding cell rolling behavior. Locally, adhesion molecules are distributed at the tips of membrane tethers. However, how functional adhesion properties are globally distributed on the individual cell’s surface is unknown. Here, we developed a label-free technique to determine the spatial distribution of adhesive properties on rolling cell surfaces. Using dark-field imaging and particle tracking, we extract the rotational motion of individual rolling cells. The rotational information allows us to construct an adhesion map along the contact circumference of a single cell. To complement this approach, we also developed a fluorescent adhesion footprint assay to record the molecular adhesion events from cell rolling. Applying the combination of the two methods on human promyelocytic leukemia cells, our results surprisingly reveal that adhesion is non-uniformly distributed in patches on the cell surfaces. Our label-free adhesion mapping methods are applicable to the variety of cell types that undergo rolling adhesion and provide a quantitative picture of cell surface adhesion at the functional and molecular level.

Postsynaptic FMRP Promotes the Pruning of Cell-to-Cell Connections among Pyramidal Neurons in the L5A Neocortical Network

PubMed Central

Patel, Ankur B.; Loerwald, Kristofer W.; Huber, Kimberly M.

2014-01-01

Pruning of structural synapses occurs with development and learning. A deficit in pruning of cortical excitatory synapses and the resulting hyperconnectivity is hypothesized to underlie the etiology of fragile X syndrome (FXS) and related autistic disorders. However, clear evidence for pruning in neocortex and its impairment in FXS remains elusive. Using simultaneous recordings of pyramidal neurons in the layer 5A neocortical network of the wild-type (WT) mouse to observe cell-to-cell connections in isolation, we demonstrate here a specific form of “connection pruning.” Connection frequency among pyramidal neurons decreases between the third and fifth postnatal weeks, indicating a period of connection pruning. Over the same interval in the FXS model mouse, the Fmr1 knock-out (KO), connection frequency does not decrease. Therefore, connection frequency in the fifth week is higher in the Fmr1 KO compared with WT, indicating a state of hyperconnectivity. These alterations are due to postsynaptic deletion of Fmr1. At early ages (2 weeks), postsynaptic Fmr1 promoted the maturation of cell-to-cell connections, but not their number. These findings indicate that impaired connection pruning at later ages, and not an excess of connection formation, underlies the hyperconnectivity in the Fmr1 KO mouse. FMRP did not appear to regulate synapses individually, but instead regulated cell-to-cell connectivity in which groups of synapses mediating a single cell-to-cell connection are uniformly removed, retained, and matured. Although we do not link connection pruning directly to the pruning of structurally defined synapses, this study nevertheless provides an important model system for studying altered pruning in FXS. PMID:24573297

Postsynaptic FMRP promotes the pruning of cell-to-cell connections among pyramidal neurons in the L5A neocortical network.

PubMed

Patel, Ankur B; Loerwald, Kristofer W; Huber, Kimberly M; Gibson, Jay R

2014-02-26

Pruning of structural synapses occurs with development and learning. A deficit in pruning of cortical excitatory synapses and the resulting hyperconnectivity is hypothesized to underlie the etiology of fragile X syndrome (FXS) and related autistic disorders. However, clear evidence for pruning in neocortex and its impairment in FXS remains elusive. Using simultaneous recordings of pyramidal neurons in the layer 5A neocortical network of the wild-type (WT) mouse to observe cell-to-cell connections in isolation, we demonstrate here a specific form of "connection pruning." Connection frequency among pyramidal neurons decreases between the third and fifth postnatal weeks, indicating a period of connection pruning. Over the same interval in the FXS model mouse, the Fmr1 knock-out (KO), connection frequency does not decrease. Therefore, connection frequency in the fifth week is higher in the Fmr1 KO compared with WT, indicating a state of hyperconnectivity. These alterations are due to postsynaptic deletion of Fmr1. At early ages (2 weeks), postsynaptic Fmr1 promoted the maturation of cell-to-cell connections, but not their number. These findings indicate that impaired connection pruning at later ages, and not an excess of connection formation, underlies the hyperconnectivity in the Fmr1 KO mouse. FMRP did not appear to regulate synapses individually, but instead regulated cell-to-cell connectivity in which groups of synapses mediating a single cell-to-cell connection are uniformly removed, retained, and matured. Although we do not link connection pruning directly to the pruning of structurally defined synapses, this study nevertheless provides an important model system for studying altered pruning in FXS.

Alteration of the fast excitatory postsynaptic current by barium in voltage-clamped amphibian sympathetic ganglion cells.

PubMed Central

Connor, E. A.; Parsons, R. L.

1984-01-01

Barium-induced alterations in fast excitatory postsynaptic currents (e.p.s.cs) have been studied in voltage-clamped bullfrog sympathetic ganglion B cells. In the presence of 2-8 mM barium, e.p.s.c. decay was prolonged and in many cells the e.p.s.c. decay phase deviated from a single exponential function. The decay phase in these cases was more accurately described as the sum of two exponential functions. The frequency of occurrence of a complex decay increased both with increasing barium concentration and with hyperpolarization. Miniature e.p.s.c. decay also was prolonged in barium-treated cells. E.p.s.c. amplitude was not markedly affected by barium (2-8 mM) in cells voltage-clamped to -50 mV whereas at -90 mV there was a progressive increase in peak size with increasing barium concentration. In control cells the e.p.s.c.-voltage relationship was linear between -20 and -100 mV; however, this relationship became progressively non-linear with membrane hyperpolarization in barium-treated cells. The e.p.s.c. reversal potential was shifted to a more negative value in the presence of barium. There was a voltage-dependent increase in charge movement during the e.p.s.c. in barium-treated cells which was not present in control cells. We conclude that the voltage-dependent alteration in e.p.s.c. decay time course, peak amplitude and charge movement in barium-treated cells is due to a direct postsynaptic action of barium on the kinetics of receptor-channel gating in postganglionic sympathetic neurones. PMID:6333261

Distinct roles for extracellular and intracellular domains in neuroligin function at inhibitory synapses

PubMed Central

Nguyen, Quynh-Anh; Horn, Meryl E; Nicoll, Roger A

2016-01-01

Neuroligins (NLGNs) are postsynaptic cell adhesion molecules that interact trans-synaptically with neurexins to mediate synapse development and function. NLGN2 is only at inhibitory synapses while NLGN3 is at both excitatory and inhibitory synapses. We found that NLGN3 function at inhibitory synapses in rat CA1 depends on the presence of NLGN2 and identified a domain in the extracellular region that accounted for this functional difference between NLGN2 and 3 specifically at inhibitory synapses. We further show that the presence of a cytoplasmic tail (c-tail) is indispensible, and identified two domains in the c-tail that are necessary for NLGN function at inhibitory synapses. These domains point to a gephyrin-dependent mechanism that is disrupted by an autism-associated mutation at R705 and a gephyrin-independent mechanism reliant on a putative phosphorylation site at S714. Our work highlights unique and separate roles for the extracellular and intracellular regions in specifying and carrying out NLGN function respectively. DOI: http://dx.doi.org/10.7554/eLife.19236.001 PMID:27805570

Synaptic genes are extensively downregulated across multiple brain regions in normal human aging and Alzheimer’s disease

PubMed Central

Berchtold, Nicole C.; Coleman, Paul D.; Cribbs, David H.; Rogers, Joseph; Gillen, Daniel L.; Cotman, Carl W.

2014-01-01

Synapses are essential for transmitting, processing, and storing information, all of which decline in aging and Alzheimer’s disease (AD). Because synapse loss only partially accounts for the cognitive declines seen in aging and AD, we hypothesized that existing synapses might undergo molecular changes that reduce their functional capacity. Microarrays were used to evaluate expression profiles of 340 synaptic genes in aging (20–99 years) and AD across 4 brain regions from 81 cases. The analysis revealed an unexpectedly large number of significant expression changes in synapse-related genes in aging, with many undergoing progressive downregulation across aging and AD. Functional classification of the genes showing altered expression revealed that multiple aspects of synaptic function are affected, notably synaptic vesicle trafficking and release, neurotransmitter receptors and receptor trafficking, postsynaptic density scaffolding, cell adhesion regulating synaptic stability, and neuromodulatory systems. The widespread declines in synaptic gene expression in normal aging suggests that function of existing synapses might be impaired, and that a common set of synaptic genes are vulnerable to change in aging and AD. PMID:23273601

Developmental switch in the contribution of presynaptic and postsynaptic NMDA receptors to long-term depression

PubMed Central

Corlew, Rebekah; Wang, Yun; Ghermazien, Haben; Erisir, Alev; Philpot, Benjamin D.

2010-01-01

NMDA receptor (NMDAR) activation is required for many forms of learning and memory as well as sensory system receptive field plasticity, yet the relative contribution of pre- and postsynaptic NMDARs over cortical development remains unknown. Here we demonstrate a rapid developmental loss of functional presynaptic NMDARs in the neocortex. Presynaptic NMDARs enhance neurotransmitter release at synapses onto visual cortex pyramidal cells in young mice (< postnatal day 20; P20), but they have no apparent effect after the onset of the critical period for receptive field plasticity (>P21). Immuno-electron microscopy revealed that the loss of presynaptic NMDAR function is likely due in part to a 50% reduction in the prevalence of presynaptic NMDARs. Coincident with the observed loss of presynaptic NMDAR function, there is an abrupt change in the mechanisms of timing-dependent long-term depression (tLTD). Induction of tLTD before the onset of the critical period requires activation of pre- but not postsynaptic NMDARs, while the induction of tLTD in older mice requires activation of postsynaptic NMDARs. By demonstrating that both pre- and postsynaptic NMDARs contribute to the induction of synaptic plasticity, and that their relative roles shift over development, our findings define a novel, and perhaps general, property of synaptic plasticity in emerging cortical circuits. PMID:17855598

Cell Adhesions: Actin-Based Modules that Mediate Cell-Extracellular Matrix and Cell-Cell Interactions

PubMed Central

Bachir, Alexia; Horwitz, Alan Rick; Nelson, W. James; Bianchini, Julie M.

2018-01-01

Cell adhesions link cells to the extracellular matrix (ECM) and to each other, and depend on interactions with the actin cytoskeleton. Both cell-ECM and cell-cell adhesion sites contain discrete, yet overlapping functional modules. These modules establish physical association with the actin cytoskeleton, locally modulate actin organization and dynamics, and trigger intracellular signaling pathways. Interplay between these modules generates distinct actin architectures that underlie different stages, types, and functions of cell-ECM and cell-cell adhesions. Actomyosin contractility is required to generate mature, stable adhesions, as well as sense and translate the mechanical properties of the cellular environment to changes in cell organization and behavior. In this chapter we discuss the organization and function of different adhesion modules and how they interact with the actin cytoskeleton. We highlight the molecular mechanisms of mechanotransduction in adhesions, and how adhesion molecules mediate crosstalk between cell-ECM and cell-cell adhesion sites. PMID:28679638

Target-specific expression of presynaptic NMDA receptors in neocortical microcircuits.

PubMed

Buchanan, Katherine A; Blackman, Arne V; Moreau, Alexandre W; Elgar, Dale; Costa, Rui P; Lalanne, Txomin; Tudor Jones, Adam A; Oyrer, Julia; Sjöström, P Jesper

2012-08-09

Traditionally, NMDA receptors are located postsynaptically; yet, putatively presynaptic NMDA receptors (preNMDARs) have been reported. Although implicated in controlling synaptic plasticity, their function is not well understood and their expression patterns are debated. We demonstrate that, in layer 5 of developing mouse visual cortex, preNMDARs specifically control synaptic transmission at pyramidal cell inputs to other pyramidal cells and to Martinotti cells, while leaving those to basket cells unaffected. We also reveal a type of interneuron that mediates ascending inhibition. In agreement with synapse-specific expression, we find preNMDAR-mediated calcium signals in a subset of pyramidal cell terminals. A tuned network model predicts that preNMDARs specifically reroute information flow in local circuits during high-frequency firing, in particular by impacting frequency-dependent disynaptic inhibition mediated by Martinotti cells, a finding that we experimentally verify. We conclude that postsynaptic cell type determines presynaptic terminal molecular identity and that preNMDARs govern information processing in neocortical columns. Copyright © 2012 Elsevier Inc. All rights reserved.

A Review of Cell Adhesion Studies for Biomedical and Biological Applications.

PubMed

Khalili, Amelia Ahmad; Ahmad, Mohd Ridzuan

2015-08-05

Cell adhesion is essential in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. The mechanical interactions between a cell and its extracellular matrix (ECM) can influence and control cell behavior and function. The essential function of cell adhesion has created tremendous interests in developing methods for measuring and studying cell adhesion properties. The study of cell adhesion could be categorized into cell adhesion attachment and detachment events. The study of cell adhesion has been widely explored via both events for many important purposes in cellular biology, biomedical, and engineering fields. Cell adhesion attachment and detachment events could be further grouped into the cell population and single cell approach. Various techniques to measure cell adhesion have been applied to many fields of study in order to gain understanding of cell signaling pathways, biomaterial studies for implantable sensors, artificial bone and tooth replacement, the development of tissue-on-a-chip and organ-on-a-chip in tissue engineering, the effects of biochemical treatments and environmental stimuli to the cell adhesion, the potential of drug treatments, cancer metastasis study, and the determination of the adhesion properties of normal and cancerous cells. This review discussed the overview of the available methods to study cell adhesion through attachment and detachment events.

A Review of Cell Adhesion Studies for Biomedical and Biological Applications

PubMed Central

Ahmad Khalili, Amelia; Ahmad, Mohd Ridzuan

2015-01-01

Cell adhesion is essential in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. The mechanical interactions between a cell and its extracellular matrix (ECM) can influence and control cell behavior and function. The essential function of cell adhesion has created tremendous interests in developing methods for measuring and studying cell adhesion properties. The study of cell adhesion could be categorized into cell adhesion attachment and detachment events. The study of cell adhesion has been widely explored via both events for many important purposes in cellular biology, biomedical, and engineering fields. Cell adhesion attachment and detachment events could be further grouped into the cell population and single cell approach. Various techniques to measure cell adhesion have been applied to many fields of study in order to gain understanding of cell signaling pathways, biomaterial studies for implantable sensors, artificial bone and tooth replacement, the development of tissue-on-a-chip and organ-on-a-chip in tissue engineering, the effects of biochemical treatments and environmental stimuli to the cell adhesion, the potential of drug treatments, cancer metastasis study, and the determination of the adhesion properties of normal and cancerous cells. This review discussed the overview of the available methods to study cell adhesion through attachment and detachment events. PMID:26251901

In vitro Flow Adhesion Assay for Analyzing Shear-resistant Adhesion of Metastatic Cancer Cells to Endothelial Cells.

PubMed

Kang, Shin-Ae; Bajana, Sandra; Tanaka, Takemi

2016-02-20

Hematogenous metastasis is a primary cause of mortality from metastatic cancer. The shear-resistant adhesion of circulating tumor cells to the vascular endothelial cell surface under blood flow is an essential step in cell extravasation and further tissue invasion. This is similar to a process exploited by leukocytes for adhesion to inflamed blood vessels (leukocyte mimicry). The shear resistant adhesion is mediated by high affinity interactions between endothelial adhesion molecules and their counter receptor ligand expressed on circulating cells. Thus, weak interaction results in a rapid detachment of circulating cells from endothelium. Despite the critical role of vascular adhesion of cancer cells in hematogenous metastasis, our knowledge regarding this process has been limited due to the difficulty of mimicking dynamic flow conditions in vitro . In order to gain better insight into the shear-resistant adhesion of cancer cells to the endothelium, we developed a protocol for measuring the shear resistant adhesion of circulating tumor cells to endothelial cells under physiologic flow conditions by adapting a well established flow adhesion assay for inflammatory cells. This technique is useful to evaluate 1) the shear resistant adhesion competency of cancer cells and 2) the endothelial adhesion molecules necessary to support cancer cell adhesion (Kang et al. , 2015).

Synaptic noise is an information bottleneck in the inner retina during dynamic visual stimulation

PubMed Central

Freed, Michael A; Liang, Zhiyin

2014-01-01

In daylight, noise generated by cones determines the fidelity with which visual signals are initially encoded. Subsequent stages of visual processing require synapses from bipolar cells to ganglion cells, but whether these synapses generate a significant amount of noise was unknown. To characterize noise generated by these synapses, we recorded excitatory postsynaptic currents from mammalian retinal ganglion cells and subjected them to a computational noise analysis. The release of transmitter quanta at bipolar cell synapses contributed substantially to the noise variance found in the ganglion cell, causing a significant loss of fidelity from bipolar cell array to postsynaptic ganglion cell. Virtually all the remaining noise variance originated in the presynaptic circuit. Circuit noise had a frequency content similar to noise shared by ganglion cells but a very different frequency content from noise from bipolar cell synapses, indicating that these synapses constitute a source of independent noise not shared by ganglion cells. These findings contribute a picture of daylight retinal circuits where noise from cones and noise generated by synaptic transmission of cone signals significantly limit visual fidelity. PMID:24297850

Soluble adhesion molecules in human cancers: sources and fates.

PubMed

van Kilsdonk, Jeroen W J; van Kempen, Léon C L T; van Muijen, Goos N P; Ruiter, Dirk J; Swart, Guido W M

2010-06-01

Adhesion molecules endow tumor cells with the necessary cell-cell contacts and cell-matrix interactions. As such, adhesion molecules are involved in cell signalling, proliferation and tumor growth. Rearrangements in the adhesion repertoire allow tumor cells to migrate, invade and form metastases. Besides these membrane-bound adhesion molecules several soluble adhesion molecules are detected in the supernatant of tumor cell lines and patient body fluids. Truncated soluble adhesion molecules can be generated by several conventional mechanisms, including alternative splicing of mRNA transcripts, chromosomal translocation, and extracellular proteolytic ectodomain shedding. Secretion of vesicles (ectosomes and exosomes) is an alternative mechanism mediating the release of full-length adhesion molecules. Soluble adhesion molecules function as modulators of cell adhesion, induce proteolytic activity and facilitate cell signalling. Additionally, adhesion molecules present on secreted vesicles might be involved in the vesicle-target cell interaction. Based on currently available data, released soluble adhesion molecules contribute to cancer progression and therefore should not be regarded as unrelated and non-functional side products of tumor progression. 2010 Elsevier GmbH. All rights reserved.

The roles of cell adhesion molecules in tumor suppression and cell migration: a new paradox.

PubMed

Moh, Mei Chung; Shen, Shali

2009-01-01

In addition to mediating cell adhesion, many cell adhesion molecules act as tumor suppressors. These proteins are capable of restricting cell growth mainly through contact inhibition. Alterations of these cell adhesion molecules are a common event in cancer. The resulting loss of cell-cell and/or cell-extracellular matrix adhesion promotes cell growth as well as tumor dissemination. Therefore, it is conventionally accepted that cell adhesion molecules that function as tumor suppressors are also involved in limiting tumor cell migration. Paradoxically, in 2005, we identified an immunoglobulin superfamily cell adhesion molecule hepaCAM that is able to suppress cancer cell growth and yet induce migration. Almost concurrently, CEACAM1 was verified to co-function as a tumor suppressor and invasion promoter. To date, the reason and mechanism responsible for this exceptional phenomenon remain unclear. Nevertheless, the emergence of these intriguing cell adhesion molecules with conflicting roles may open a new chapter to the biological significance of cell adhesion molecules.

Suppression of lysyl-tRNA synthetase, KRS, causes incomplete epithelial-mesenchymal transition and ineffective cell‑extracellular matrix adhesion for migration.

PubMed

Nam, Seo Hee; Kang, Minkyung; Ryu, Jihye; Kim, Hye-Jin; Kim, Doyeun; Kim, Dae Gyu; Kwon, Nam Hoon; Kim, Sunghoon; Lee, Jung Weon

2016-04-01

The cell-adhesion properties of cancer cells can be targeted to block cancer metastasis. Although cytosolic lysyl-tRNA synthetase (KRS) functions in protein synthesis, KRS on the plasma membrane is involved in cancer metastasis. We hypothesized that KRS is involved in cell adhesion-related signal transduction for cellular migration. To test this hypothesis, colon cancer cells with modulated KRS protein levels were analyzed for cell-cell contact and cell-substrate adhesion properties and cellular behavior. Although KRS suppression decreased expression of cell-cell adhesion molecules, cells still formed colonies without being scattered, supporting an incomplete epithelial mesenchymal transition. Noteworthy, KRS-suppressed cells still exhibited focal adhesions on laminin, with Tyr397-phopshorylated focal adhesion kinase (FAK), but they lacked laminin-adhesion-mediated extracellular signal-regulated kinase (ERK) and paxillin activation. KRS, p67LR and integrin α6β1 were found to interact, presumably to activate ERK for paxillin expression and Tyr118 phosphorylation even without involvement of FAK, so that specific inhibition of ERK or KRS in parental HCT116 cells blocked cell-cell adhesion and cell-substrate properties for focal adhesion formation and signaling activity. Together, these results indicate that KRS can promote cell-cell and cell-ECM adhesion for migration.

Cell adhesion heterogeneity reinforces tumour cell dissemination: novel insights from a mathematical model.

PubMed

Reher, David; Klink, Barbara; Deutsch, Andreas; Voss-Böhme, Anja

2017-08-11

Cancer cell invasion, dissemination, and metastasis have been linked to an epithelial-mesenchymal transition (EMT) of individual tumour cells. During EMT, adhesion molecules like E-cadherin are downregulated and the decrease of cell-cell adhesion allows tumour cells to dissociate from the primary tumour mass. This complex process depends on intracellular cues that are subject to genetic and epigenetic variability, as well as extrinsic cues from the local environment resulting in a spatial heterogeneity in the adhesive phenotype of individual tumour cells. Here, we use a novel mathematical model to study how adhesion heterogeneity, influenced by intrinsic and extrinsic factors, affects the dissemination of tumour cells from an epithelial cell population. The model is a multiscale cellular automaton that couples intracellular adhesion receptor regulation with cell-cell adhesion. Simulations of our mathematical model indicate profound effects of adhesion heterogeneity on tumour cell dissemination. In particular, we show that a large variation of intracellular adhesion receptor concentrations in a cell population reinforces cell dissemination, regardless of extrinsic cues mediated through the local cell density. However, additional control of adhesion receptor concentration through the local cell density, which can be assumed in healthy cells, weakens the effect. Furthermore, we provide evidence that adhesion heterogeneity can explain the remarkable differences in adhesion receptor concentrations of epithelial and mesenchymal phenotypes observed during EMT and might drive early dissemination of tumour cells. Our results suggest that adhesion heterogeneity may be a universal trigger to reinforce cell dissemination in epithelial cell populations. This effect can be at least partially compensated by a control of adhesion receptor regulation through neighbouring cells. Accordingly, our findings explain how both an increase in intra-tumour adhesion heterogeneity and the loss of control through the local environment can promote tumour cell dissemination. This article was reviewed by Hanspeter Herzel, Thomas Dandekar and Marek Kimmel.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tao-Cheng, Jung-Hwa; Yang, Yijung; Bayer, K. Ulrich

Highlights: • NMDA-induces accumulation of Shank at the postsynaptic density. • Shank accumulation is preferential to the distal region of the postsynaptic density. • Shank accumulation is mediated by CaMKII. - Abstract: Shank is a specialized scaffold protein present in high abundance at the postsynaptic density (PSD). Using pre-embedding immunogold electron microscopy on cultured hippocampal neurons, we had previously demonstrated further accumulation of Shank at the PSD under excitatory conditions. Here, using the same experimental protocol, we demonstrate that a cell permeable CaMKII inhibitor, tatCN21, blocks NMDA-induced accumulation of Shank at the PSD. Furthermore we show that NMDA application changesmore » the distribution pattern of Shank at the PSD, promoting a 7–10 nm shift in the median distance of Shank labels away from the postsynaptic membrane. Inhibition of CaMKII with tatCN21 also blocks this shift in the distribution of Shank. Altogether these results imply that upon activation of NMDA receptors, CaMKII mediates accumulation of Shank, preferentially at the distal regions of the PSD complex extending toward the cytoplasm.« less

Cell-cell and cell-extracellular matrix adhesions cooperate to organize actomyosin networks and maintain force transmission during dorsal closure.

PubMed

Goodwin, Katharine; Lostchuck, Emily E; Cramb, Kaitlyn M L; Zulueta-Coarasa, Teresa; Fernandez-Gonzalez, Rodrigo; Tanentzapf, Guy

2017-05-15

Tissue morphogenesis relies on the coordinated action of actin networks, cell-cell adhesions, and cell-extracellular matrix (ECM) adhesions. Such coordination can be achieved through cross-talk between cell-cell and cell-ECM adhesions. Drosophila dorsal closure (DC), a morphogenetic process in which an extraembryonic tissue called the amnioserosa contracts and ingresses to close a discontinuity in the dorsal epidermis of the embryo, requires both cell-cell and cell-ECM adhesions. However, whether the functions of these two types of adhesions are coordinated during DC is not known. Here we analyzed possible interdependence between cell-cell and cell-ECM adhesions during DC and its effect on the actomyosin network. We find that loss of cell-ECM adhesion results in aberrant distributions of cadherin-mediated adhesions and actin networks in the amnioserosa and subsequent disruption of myosin recruitment and dynamics. Moreover, loss of cell-cell adhesion caused up-regulation of cell-ECM adhesion, leading to reduced cell deformation and force transmission across amnioserosa cells. Our results show how interdependence between cell-cell and cell-ECM adhesions is important in regulating cell behaviors, force generation, and force transmission critical for tissue morphogenesis. © 2017 Goodwin, Lostchuck, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Activation-Dependent Rapid Postsynaptic Clustering of Glycine Receptors in Mature Spinal Cord Neurons

PubMed Central

Eto, Kei; Murakoshi, Hideji; Watanabe, Miho; Hirata, Hiromi; Moorhouse, Andrew J.; Ishibashi, Hitoshi

2017-01-01

Abstract Inhibitory synapses are established during development but continue to be generated and modulated in strength in the mature nervous system. In the spinal cord and brainstem, presynaptically released inhibitory neurotransmitter dominantly switches from GABA to glycine during normal development in vivo. While presynaptic mechanisms of the shift of inhibitory neurotransmission are well investigated, the contribution of postsynaptic neurotransmitter receptors to this shift is not fully elucidated. Synaptic clustering of glycine receptors (GlyRs) is regulated by activation-dependent depolarization in early development. However, GlyR activation induces hyperpolarization after the first postnatal week, and little is known whether and how presynaptically released glycine regulates postsynaptic receptors in a depolarization-independent manner in mature developmental stage. Here we developed spinal cord neuronal culture of rodents using chronic strychnine application to investigate whether initial activation of GlyRs in mature stage could change postsynaptic localization of GlyRs. Immunocytochemical analyses demonstrate that chronic blockade of GlyR activation until mature developmental stage resulted in smaller clusters of postsynaptic GlyRs that could be enlarged upon receptor activation for 1 h in the mature stage. Furthermore, live cell-imaging techniques show that GlyR activation decreases its lateral diffusion at synapses, and this phenomenon is dependent on PKC, but neither Ca2+ nor CaMKII activity. These results suggest that the GlyR activation can regulate receptor diffusion and cluster size at inhibitory synapses in mature stage, providing not only new insights into the postsynaptic mechanism of shifting inhibitory neurotransmission but also the inhibitory synaptic plasticity in mature nervous system. PMID:28197549

Activation-Dependent Rapid Postsynaptic Clustering of Glycine Receptors in Mature Spinal Cord Neurons.

PubMed

Nakahata, Yoshihisa; Eto, Kei; Murakoshi, Hideji; Watanabe, Miho; Kuriu, Toshihiko; Hirata, Hiromi; Moorhouse, Andrew J; Ishibashi, Hitoshi; Nabekura, Junichi

2017-01-01

Inhibitory synapses are established during development but continue to be generated and modulated in strength in the mature nervous system. In the spinal cord and brainstem, presynaptically released inhibitory neurotransmitter dominantly switches from GABA to glycine during normal development in vivo . While presynaptic mechanisms of the shift of inhibitory neurotransmission are well investigated, the contribution of postsynaptic neurotransmitter receptors to this shift is not fully elucidated. Synaptic clustering of glycine receptors (GlyRs) is regulated by activation-dependent depolarization in early development. However, GlyR activation induces hyperpolarization after the first postnatal week, and little is known whether and how presynaptically released glycine regulates postsynaptic receptors in a depolarization-independent manner in mature developmental stage. Here we developed spinal cord neuronal culture of rodents using chronic strychnine application to investigate whether initial activation of GlyRs in mature stage could change postsynaptic localization of GlyRs. Immunocytochemical analyses demonstrate that chronic blockade of GlyR activation until mature developmental stage resulted in smaller clusters of postsynaptic GlyRs that could be enlarged upon receptor activation for 1 h in the mature stage. Furthermore, live cell-imaging techniques show that GlyR activation decreases its lateral diffusion at synapses, and this phenomenon is dependent on PKC, but neither Ca 2+ nor CaMKII activity. These results suggest that the GlyR activation can regulate receptor diffusion and cluster size at inhibitory synapses in mature stage, providing not only new insights into the postsynaptic mechanism of shifting inhibitory neurotransmission but also the inhibitory synaptic plasticity in mature nervous system.

Specialized postsynaptic morphology enhances neurotransmitter dilution and high-frequency signaling at an auditory synapse.

PubMed

Graydon, Cole W; Cho, Soyoun; Diamond, Jeffrey S; Kachar, Bechara; von Gersdorff, Henrique; Grimes, William N

2014-06-11

Sensory processing in the auditory system requires that synapses, neurons, and circuits encode information with particularly high temporal and spectral precision. In the amphibian papillia, sound frequencies up to 1 kHz are encoded along a tonotopic array of hair cells and transmitted to afferent fibers via fast, repetitive synaptic transmission, thereby promoting phase locking between the presynaptic and postsynaptic cells. Here, we have combined serial section electron microscopy, paired electrophysiological recordings, and Monte Carlo diffusion simulations to examine novel mechanisms that facilitate fast synaptic transmission in the inner ear of frogs (Rana catesbeiana and Rana pipiens). Three-dimensional anatomical reconstructions reveal specialized spine-like contacts between individual afferent fibers and hair cells that are surrounded by large, open regions of extracellular space. Morphologically realistic diffusion simulations suggest that these local enlargements in extracellular space speed transmitter clearance and reduce spillover between neighboring synapses, thereby minimizing postsynaptic receptor desensitization and improving sensitivity during prolonged signal transmission. Additionally, evoked EPSCs in afferent fibers are unaffected by glutamate transporter blockade, suggesting that transmitter diffusion and dilution, and not uptake, play a primary role in speeding neurotransmission and ensuring fidelity at these synapses. Copyright © 2014 the authors 0270-6474/14/348358-15$15.00/0.

Actin-Based Adhesion Modules Mediate Cell Interactions with the Extracellular Matrix and Neighboring Cells.

PubMed

Bachir, Alexia I; Horwitz, Alan Rick; Nelson, W James; Bianchini, Julie M

2017-07-05

Cell adhesions link cells to the extracellular matrix (ECM) and to each other and depend on interactions with the actin cytoskeleton. Both cell-ECM and cell-cell adhesion sites contain discrete, yet overlapping, functional modules. These modules establish physical associations with the actin cytoskeleton, locally modulate actin organization and dynamics, and trigger intracellular signaling pathways. Interplay between these modules generates distinct actin architectures that underlie different stages, types, and functions of cell-ECM and cell-cell adhesions. Actomyosin contractility is required to generate mature, stable adhesions, as well as to sense and translate the mechanical properties of the cellular environment into changes in cell organization and behavior. Here, we review the organization and function of different adhesion modules and how they interact with the actin cytoskeleton. We highlight the molecular mechanisms of mechanotransduction in adhesions and how adhesion molecules mediate cross talk between cell-ECM and cell-cell adhesion sites. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.

Just-in-time connectivity for large spiking networks.

PubMed

Lytton, William W; Omurtag, Ahmet; Neymotin, Samuel A; Hines, Michael L

2008-11-01

The scale of large neuronal network simulations is memory limited due to the need to store connectivity information: connectivity storage grows as the square of neuron number up to anatomically relevant limits. Using the NEURON simulator as a discrete-event simulator (no integration), we explored the consequences of avoiding the space costs of connectivity through regenerating connectivity parameters when needed: just in time after a presynaptic cell fires. We explored various strategies for automated generation of one or more of the basic static connectivity parameters: delays, postsynaptic cell identities, and weights, as well as run-time connectivity state: the event queue. Comparison of the JitCon implementation to NEURON's standard NetCon connectivity method showed substantial space savings, with associated run-time penalty. Although JitCon saved space by eliminating connectivity parameters, larger simulations were still memory limited due to growth of the synaptic event queue. We therefore designed a JitEvent algorithm that added items to the queue only when required: instead of alerting multiple postsynaptic cells, a spiking presynaptic cell posted a callback event at the shortest synaptic delay time. At the time of the callback, this same presynaptic cell directly notified the first postsynaptic cell and generated another self-callback for the next delay time. The JitEvent implementation yielded substantial additional time and space savings. We conclude that just-in-time strategies are necessary for very large network simulations but that a variety of alternative strategies should be considered whose optimality will depend on the characteristics of the simulation to be run.

Just in time connectivity for large spiking networks

PubMed Central

Lytton, William W.; Omurtag, Ahmet; Neymotin, Samuel A; Hines, Michael L

2008-01-01

The scale of large neuronal network simulations is memory-limited due to the need to store connectivity information: connectivity storage grows as the square of neuron number up to anatomically-relevant limits. Using the NEURON simulator as a discrete-event simulator (no integration), we explored the consequences of avoiding the space costs of connectivity through regenerating connectivity parameters when needed – just-in-time after a presynaptic cell fires. We explored various strategies for automated generation of one or more of the basic static connectivity parameters: delays, postsynaptic cell identities and weights, as well as run-time connectivity state: the event queue. Comparison of the JitCon implementation to NEURON’s standard NetCon connectivity method showed substantial space savings, with associated run-time penalty. Although JitCon saved space by eliminating connectivity parameters, larger simulations were still memory-limited due to growth of the synaptic event queue. We therefore designed a JitEvent algorithm that only added items to the queue when required: instead of alerting multiple postsynaptic cells, a spiking presynaptic cell posted a callback event at the shortest synaptic delay time. At the time of the callback, this same presynaptic cell directly notified the first postsynaptic cell and generated another self-callback for the next delay time. The JitEvent implementation yielded substantial additional time and space savings. We conclude that just-in-time strategies are necessary for very large network simulations but that a variety of alternative strategies should be considered whose optimality will depend on the characteristics of the simulation to be run. PMID:18533821

Analysis of slow depolarizing potential in frog taste cell induced by parasympathetic efferent stimulation under hypoxia.

PubMed

Sato, Toshihide; Nishishita, Kazushisa; Okada, Yukio; Toda, Kazuo

2007-05-01

Strong electrical stimulation (ES) of the frog glossopharyngeal (GP) efferent nerve induced slow depolarizing potentials (DPs) in taste cells under hypoxia. This study aimed to elucidate whether the slow DPs were postsynaptically induced in taste cells. After a block of parasympathetic nerve (PSN) ganglia by tubocurarine, ES of GP nerve never induced slow DPs in the taste cells, so slow DPs were induced by PSN. When Ca(2+) in the blood plasma under hypoxia was decreased to approximately 0.5 mM, the slow DPs reduced in amplitude and lengthened in latency. Increasing the normal Ca(2+) to approximately 20 mM increased the amplitude of slow DPs and shortened the latency. Addition of Cd(2+) to the plasma greatly reduced the amplitude of slow DPs and lengthened the latency. These data suggest that the slow DPs depend on Ca(2+) and Cd(2+) concentration at the presynaptic PSN terminals of taste disk. Antagonists, [D-Arg(1), D-Trp(7,9), Leu(11)]-substance P and L-703 606, of neurotransmitter substance P neurokinin(1) receptor completely blocked the slow DPs. Intravenous application of substance P induced a DP of approximately 7 mV and a reduction of membrane resistance of approximately 48% in taste cells. A nonselective cation channel antagonist, flufenamic acid, completely blocked the slow DPs. These findings suggest that the slow DPs are postsynaptically initiated in frog taste cells under hypoxia by opening nonselective cation channels on the postsynaptic membrane after substance P is probably released from the presynaptic PSN axon terminals.

Autophagy promotes degradation of internalized collagen and regulates distribution of focal adhesions to suppress cell adhesion

PubMed Central

Kawano, Shinichi; Esaki, Motohiro; Torisu, Kumiko; Matsuno, Yuichi; Kitazono, Takanari

2017-01-01

ABSTRACT Adhesion of cells to the extracellular matrix (ECM) via focal adhesions (FAs) is crucial for cell survival, migration, and differentiation. Although the regulation of FAs, including by integrins and the ECM, is important to cell behavior, how FAs are regulated is not well known. Autophagy is induced by both cell adhesion and cell detachment. Here, we showed that autophagosomes are located close to internalized collagen and paxillin, which is a well-known marker of FAs. Autophagy-deficient cells showed increased levels of internalized collagen compared with control cells. Moreover, paxillin exhibited a more peripheral distribution and the area of paxillin was increased, and adhesion-induced focal adhesion kinase signaling was impaired and adhesion was enhanced, in autophagy-deficient cells. These results suggest that autophagy suppressed cell adhesion by regulating internalized ECM and FAs. PMID:28970230

Analytical cell adhesion chromatography reveals impaired persistence of metastatic cell rolling adhesion to P-selectin

PubMed Central

Oh, Jaeho; Edwards, Erin E.; McClatchey, P. Mason; Thomas, Susan N.

2015-01-01

ABSTRACT Selectins facilitate the recruitment of circulating cells from the bloodstream by mediating rolling adhesion, which initiates the cell–cell signaling that directs extravasation into surrounding tissues. To measure the relative efficiency of cell adhesion in shear flow for in vitro drug screening, we designed and implemented a microfluidic-based analytical cell adhesion chromatography system. The juxtaposition of instantaneous rolling velocities with elution times revealed that human metastatic cancer cells, but not human leukocytes, had a reduced capacity to sustain rolling adhesion with P-selectin. We define a new parameter, termed adhesion persistence, which is conceptually similar to migration persistence in the context of chemotaxis, but instead describes the capacity of cells to resist the influence of shear flow and sustain rolling interactions with an adhesive substrate that might modulate the probability of extravasation. Among cell types assayed, adhesion persistence to P-selectin was specifically reduced in metastatic but not leukocyte-like cells in response to a low dose of heparin. In conclusion, we demonstrate this as an effective methodology to identify selectin adhesion antagonist doses that modulate homing cell adhesion and engraftment in a cell-subtype-selective manner. PMID:26349809

Human embryonic stem cell-derived neurons adopt and regulate the activity of an established neural network

PubMed Central

Weick, Jason P.; Liu, Yan; Zhang, Su-Chun

2011-01-01

Whether hESC-derived neurons can fully integrate with and functionally regulate an existing neural network remains unknown. Here, we demonstrate that hESC-derived neurons receive unitary postsynaptic currents both in vitro and in vivo and adopt the rhythmic firing behavior of mouse cortical networks via synaptic integration. Optical stimulation of hESC-derived neurons expressing Channelrhodopsin-2 elicited both inhibitory and excitatory postsynaptic currents and triggered network bursting in mouse neurons. Furthermore, light stimulation of hESC-derived neurons transplanted to the hippocampus of adult mice triggered postsynaptic currents in host pyramidal neurons in acute slice preparations. Thus, hESC-derived neurons can participate in and modulate neural network activity through functional synaptic integration, suggesting they are capable of contributing to neural network information processing both in vitro and in vivo. PMID:22106298

Spike Train Auto-Structure Impacts Post-Synaptic Firing and Timing-Based Plasticity

PubMed Central

Scheller, Bertram; Castellano, Marta; Vicente, Raul; Pipa, Gordon

2011-01-01

Cortical neurons are typically driven by several thousand synapses. The precise spatiotemporal pattern formed by these inputs can modulate the response of a post-synaptic cell. In this work, we explore how the temporal structure of pre-synaptic inhibitory and excitatory inputs impact the post-synaptic firing of a conductance-based integrate and fire neuron. Both the excitatory and inhibitory input was modeled by renewal gamma processes with varying shape factors for modeling regular and temporally random Poisson activity. We demonstrate that the temporal structure of mutually independent inputs affects the post-synaptic firing, while the strength of the effect depends on the firing rates of both the excitatory and inhibitory inputs. In a second step, we explore the effect of temporal structure of mutually independent inputs on a simple version of Hebbian learning, i.e., hard bound spike-timing-dependent plasticity. We explore both the equilibrium weight distribution and the speed of the transient weight dynamics for different mutually independent gamma processes. We find that both the equilibrium distribution of the synaptic weights and the speed of synaptic changes are modulated by the temporal structure of the input. Finally, we highlight that the sensitivity of both the post-synaptic firing as well as the spike-timing-dependent plasticity on the auto-structure of the input of a neuron could be used to modulate the learning rate of synaptic modification. PMID:22203800

Changes in E-cadherin rigidity sensing regulate cell adhesion.

PubMed

Collins, Caitlin; Denisin, Aleksandra K; Pruitt, Beth L; Nelson, W James

2017-07-18

Mechanical cues are sensed and transduced by cell adhesion complexes to regulate diverse cell behaviors. Extracellular matrix (ECM) rigidity sensing by integrin adhesions has been well studied, but rigidity sensing by cadherins during cell adhesion is largely unexplored. Using mechanically tunable polyacrylamide (PA) gels functionalized with the extracellular domain of E-cadherin (Ecad-Fc), we showed that E-cadherin-dependent epithelial cell adhesion was sensitive to changes in PA gel elastic modulus that produced striking differences in cell morphology, actin organization, and membrane dynamics. Traction force microscopy (TFM) revealed that cells produced the greatest tractions at the cell periphery, where distinct types of actin-based membrane protrusions formed. Cells responded to substrate rigidity by reorganizing the distribution and size of high-traction-stress regions at the cell periphery. Differences in adhesion and protrusion dynamics were mediated by balancing the activities of specific signaling molecules. Cell adhesion to a 30-kPa Ecad-Fc PA gel required Cdc42- and formin-dependent filopodia formation, whereas adhesion to a 60-kPa Ecad-Fc PA gel induced Arp2/3-dependent lamellipodial protrusions. A quantitative 3D cell-cell adhesion assay and live cell imaging of cell-cell contact formation revealed that inhibition of Cdc42, formin, and Arp2/3 activities blocked the initiation, but not the maintenance of established cell-cell adhesions. These results indicate that the same signaling molecules activated by E-cadherin rigidity sensing on PA gels contribute to actin organization and membrane dynamics during cell-cell adhesion. We hypothesize that a transition in the stiffness of E-cadherin homotypic interactions regulates actin and membrane dynamics during initial stages of cell-cell adhesion.

There are four dynamically and functionally distinct populations of E-cadherin in cell junctions

PubMed Central

Erami, Zahra; Timpson, Paul; Yao, Wu; Zaidel-Bar, Ronen; Anderson, Kurt I.

2015-01-01

ABSTRACT E-cadherin is a trans-membrane tumor suppressor responsible for epithelial cell adhesion. E-cadherin forms adhesive clusters through combined extra-cellular cis- and trans-interactions and intracellular interaction with the actin cytoskeleton. Here we identify four populations of E-cadherin within cell junctions based on the molecular interactions which determine their mobility and adhesive properties. Adhesive and non-adhesive populations of E-cadherin each consist of mobile and immobile fractions. Up to half of the E-cadherin immobilized in cell junctions is non-adhesive. Incorporation of E-cadherin into functional adhesions require all three adhesive interactions, with deletion of any one resulting in loss of effective cell-cell adhesion. Interestingly, the only interaction which could independently slow the diffusion of E-cadherin was the tail-mediated intra-cellular interaction. The adhesive and non-adhesive mobile fractions of E-cadherin can be distinguished by their sensitivity to chemical cross-linking with adhesive clusters. Our data define the size, mobility, and adhesive properties of four distinct populations of E-cadherin within cell junctions, and support association with the actin cytoskeleton as the first step in adhesion formation. PMID:26471767

Prenatal exposure to valproic acid leads to reduced expression of synaptic adhesion molecule neuroligin 3 in mice.

PubMed

Kolozsi, E; Mackenzie, R N; Roullet, F I; deCatanzaro, D; Foster, J A

2009-11-10

In rodents, a single administration of valproic acid (VPA) in utero leads to developmental delays and lifelong deficits in motor performance, social behavior, and anxiety-like behavior in the offspring. Recently, we have demonstrated that VPA mice show alterations in postnatal growth and development, and deficits in olfactory discrimination and social behavior early in development. Based on behavioral and molecular parallels between VPA rodents and individuals with autism, maternal challenge with VPA has been suggested to be a good animal model of autism. Neuroligins (NLGN) are a family of postsynaptic cell-adhesion molecules that play a role in synaptic maturation through association with their presynaptic partners, the neurexins (NRXN). Both NLGNs and NRXN members have been implicated in genetic studies of autism. In the present study, we examined changes at the level of expression of NLGN and NRXN mRNAs in the adult brain from mice exposed in utero to VPA. Mouse brain tissue was processed using in situ hybridization and analyzed with densitometry to examine expression of three NLGN genes (NLGN1, NLGN2, and NLGN3) and three NRXN genes (NRXN1, NRXN2, and NRXN3). Expression levels of NLGN1, NLGN2, NRXN1, NRXN2, and NRXN3 were observed to be similar in VPA and control mice. NLGN3 mRNA expression was found to be significantly lower in the VPA mice relative to control animals in hippocampal subregions, cornu ammonis (CA1) and dentate gyrus, and somatosensory cortex. This lowered expression may be linked to autistic-like behavioral phenotype observed in the VPA mice.

Heterogeneity of Focal Adhesions and Focal Contacts in Motile Fibroblasts.

PubMed

Gladkikh, Aleena; Kovaleva, Anastasia; Tvorogova, Anna; Vorobjev, Ivan A

2018-01-01

Cell-extracellular matrix (ECM) adhesion is an important property of virtually all cells in multicellular organisms. Cell-ECM adhesion studies, therefore, are very significant both for biology and medicine. Over the last three decades, biomedical studies resulted in a tremendous advance in our understanding of the molecular basis and functions of cell-ECM adhesion. Based on morphological and molecular criteria, several different types of model cell-ECM adhesion structures including focal adhesions, focal complexes, fibrillar adhesions, podosomes, and three-dimensional matrix adhesions have been described. All the subcellular structures that mediate cell-ECM adhesion are quite heterogeneous, often varying in size, shape, distribution, dynamics, and, to a certain extent, molecular constituents. The morphological "plasticity" of cell-ECM adhesion perhaps reflects the needs of cells to sense, adapt, and respond to a variety of extracellular environments. In addition, cell type (e.g., differentiation status, oncogenic transformation, etc.) often exerts marked influence on the structure of cell-ECM adhesions. Although molecular, genetic, biochemical, and structural studies provide important maps or "snapshots" of cell-ECM adhesions, the area of research that is equally valuable is to study the heterogeneity of FA subpopulations within cells. Recently time-lapse observations on the FA dynamics become feasible, and behavior of individual FA gives additional information on cell-ECM interactions. Here we describe a robust method of labeling of FA using plasmids with fluorescent markers for paxillin and vinculin and quantifying the morphological and dynamical parameters of FA.

Accumulation of K+ in the synaptic cleft modulates activity by influencing both vestibular hair cell and calyx afferent in the turtle

PubMed Central

Contini, Donatella; Price, Steven D.

2016-01-01

Key points In the synaptic cleft between type I hair cells and calyceal afferents, K+ ions accumulate as a function of activity, dynamically altering the driving force and permeation through ion channels facing the synaptic cleft.High‐fidelity synaptic transmission is possible due to large conductances that minimize hair cell and afferent time constants in the presence of significant membrane capacitance.Elevated potassium maintains hair cells near a potential where transduction currents are sufficient to depolarize them to voltages necessary for calcium influx and synaptic vesicle fusion.Elevated potassium depolarizes the postsynaptic afferent by altering ion permeation through hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels, and contributes to depolarizing the afferent to potentials where a single EPSP (quantum) can generate an action potential.With increased stimulation, hair cell depolarization increases the frequency of quanta released, elevates [K+]cleft and depolarizes the afferent to potentials at which smaller and smaller EPSPs would be sufficient to trigger APs. Abstract Fast neurotransmitters act in conjunction with slower modulatory effectors that accumulate in restricted synaptic spaces found at giant synapses such as the calyceal endings in the auditory and vestibular systems. Here, we used dual patch‐clamp recordings from turtle vestibular hair cells and their afferent neurons to show that potassium ions accumulating in the synaptic cleft modulated membrane potentials and extended the range of information transfer. High‐fidelity synaptic transmission was possible due to large conductances that minimized hair cell and afferent time constants in the presence of significant membrane capacitance. Increased potassium concentration in the cleft maintained the hair cell near potentials that promoted the influx of calcium necessary for synaptic vesicle fusion. The elevated potassium concentration also depolarized the postsynaptic neuron by altering ion permeation through hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels. This depolarization enabled the afferent to reliably generate action potentials evoked by single AMPA‐dependent EPSPs. Depolarization of the postsynaptic afferent could also elevate potassium in the synaptic cleft, and would depolarize other hair cells enveloped by the same neuritic process increasing the fidelity of neurotransmission at those synapses as well. Collectively, these data demonstrate that neuronal activity gives rise to potassium accumulation, and suggest that potassium ion action on HCN channels can modulate neurotransmission, preserving the fidelity of high‐speed synaptic transmission by dynamically shifting the resting potentials of both presynaptic and postsynaptic cells. PMID:27633787

Rac-WAVE-mediated actin reorganization is required for organization and maintenance of cell-cell adhesion.

PubMed

Yamazaki, Daisuke; Oikawa, Tsukasa; Takenawa, Tadaomi

2007-01-01

During cadherin-dependent cell-cell adhesion, the actin cytoskeleton undergoes dynamic reorganization in epithelial cells. Rho-family small GTPases, which regulate actin dynamics, play pivotal roles in cadherin-dependent cell-cell adhesion; however, the precise molecular mechanisms that underlie cell-cell adhesion formation remain unclear. Here we show that Wiskott-Aldrich syndrome protein family verprolin-homologous protein (WAVE)-mediated reorganization of actin, downstream of Rac plays an important role in normal development of cadherin-dependent cell-cell adhesions in MDCK cells. Rac-induced development of cadherin-dependent adhesions required WAVE2-dependent actin reorganization. The process of cell-cell adhesion is divided into three steps: formation of new cell-cell contacts, stabilization of these new contacts and junction maturation. WAVE1 and WAVE2 were expressed in MDCK cells. The functions of WAVE1 and WAVE2 were redundant in this system but WAVE2 appeared to play a more significant role. During the first step, WAVE2-dependent lamellipodial protrusions facilitated formation of cell-cell contacts. During the second step, WAVE2 recruited actin filaments to new cell-cell contacts and stabilized newly formed cadherin clusters. During the third step, WAVE2-dependent actin reorganization was required for organization and maintenance of mature cell-cell adhesions. Thus, Rac-WAVE-dependent actin reorganization is not only involved in formation of cell-cell adhesions but is also required for their maintenance.

Effect of flagella expression on adhesion of Achromobacter piechaudii to chalk surfaces.

PubMed

Nejidat, A; Saadi, I; Ronen, Z

2008-12-01

To examine flagella role and cell motility in adhesion of Achromobacter piechaudii to chalk. Transmission electron microscopy revealed that stationary cells have thicker and longer flagella than logarithmic cells. SDS-PAGE analysis showed that flagellin was more abundant in stationary cells than logarithmic ones. Sonication or inhibition of flagellin synthesis caused a 30% reduction in adhesion to chalk. Preincubation of chalk with flagella extracts reduced adhesion, by 50%. Three motility mutants were isolated. Mutants 94 and 153 were nonmotile, expressed normal levels of flagellin, have regular flagella and exhibited reduced adhesion. Mutant 208 expressed low levels of flagellin, no flagella and a spherical cell shape but with normal adhesion capacity. Multiple cell surface factors affect the adhesion efficiency to chalk. Flagella per se through physical interaction and through cell motility contribute to the adhesion process. The adhesion behaviour of mutant 208 suggests that cell shape can compensate for flagellar removal and motility. Physiological status affects bacterial cell surface properties and hence adhesion efficiency to chalk. This interaction is essential to sustain biodegradation activities and thus, remediation of contaminated chalk aquifers.

Burst-induced anti-Hebbian depression acts through short-term synaptic dynamics to cancel redundant sensory signals.

PubMed

Harvey-Girard, Erik; Lewis, John; Maler, Leonard

2010-04-28

Weakly electric fish can enhance the detection and localization of important signals such as those of prey in part by cancellation of redundant spatially diffuse electric signals due to, e.g., their tail bending. The cancellation mechanism is based on descending input, conveyed by parallel fibers emanating from cerebellar granule cells, that produces a negative image of the global low-frequency signals in pyramidal cells within the first-order electrosensory region, the electrosensory lateral line lobe (ELL). Here we demonstrate that the parallel fiber synaptic input to ELL pyramidal cell undergoes long-term depression (LTD) whenever both parallel fiber afferents and their target cells are stimulated to produce paired burst discharges. Paired large bursts (4-4) induce robust LTD over pre-post delays of up to +/-50 ms, whereas smaller bursts (2-2) induce weaker LTD. Single spikes (either presynaptic or postsynaptic) paired with bursts did not induce LTD. Tetanic presynaptic stimulation was also ineffective in inducing LTD. Thus, we have demonstrated a form of anti-Hebbian LTD that depends on the temporal correlation of burst discharge. We then demonstrated that the burst-induced LTD is postsynaptic and requires the NR2B subunit of the NMDA receptor, elevation of postsynaptic Ca(2+), and activation of CaMKIIbeta. A model incorporating local inhibitory circuitry and previously identified short-term presynaptic potentiation of the parallel fiber synapses further suggests that the combination of burst-induced LTD, presynaptic potentiation, and local inhibition may be sufficient to explain the generation of the negative image and cancellation of redundant sensory input by ELL pyramidal cells.

Roles of cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1.

PubMed

Shinohara, M; Kodama, A; Matozaki, T; Fukuhara, A; Tachibana, K; Nakanishi, H; Takai, Y

2001-06-01

Gab-1 is a multiple docking protein that is tyrosine phosphorylated by receptor tyrosine kinases such as c-Met, hepatocyte growth factor/scatter factor receptor, and epidermal growth factor receptor. We have now demonstrated that cell-cell adhesion also induces marked tyrosine phosphorylation of Gab-1 and that disruption of cell-cell adhesion results in its dephosphorylation. An anti-E-cadherin antibody decreased cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas the expression of E-cadherin specifically induced tyrosine phosphorylation of Gab-1. A relatively selective inhibitor of Src family kinases reduced cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas expression of a dominant-negative mutant of Csk increased it. Disruption of cell-cell adhesion, which reduced tyrosine phosphorylation of Gab-1, also reduced the activation of mitogen-activated protein kinase and Akt in response to cell-cell adhesion. These results indicate that E-cadherin-mediated cell-cell adhesion induces tyrosine phosphorylation by a Src family kinase of Gab-1, thereby regulating the activation of Ras/MAP kinase and phosphatidylinositol 3-kinase/Akt cascades.

Changes in E-cadherin rigidity sensing regulate cell adhesion

PubMed Central

Collins, Caitlin; Pruitt, Beth L.; Nelson, W. James

2017-01-01

Mechanical cues are sensed and transduced by cell adhesion complexes to regulate diverse cell behaviors. Extracellular matrix (ECM) rigidity sensing by integrin adhesions has been well studied, but rigidity sensing by cadherins during cell adhesion is largely unexplored. Using mechanically tunable polyacrylamide (PA) gels functionalized with the extracellular domain of E-cadherin (Ecad-Fc), we showed that E-cadherin–dependent epithelial cell adhesion was sensitive to changes in PA gel elastic modulus that produced striking differences in cell morphology, actin organization, and membrane dynamics. Traction force microscopy (TFM) revealed that cells produced the greatest tractions at the cell periphery, where distinct types of actin-based membrane protrusions formed. Cells responded to substrate rigidity by reorganizing the distribution and size of high-traction-stress regions at the cell periphery. Differences in adhesion and protrusion dynamics were mediated by balancing the activities of specific signaling molecules. Cell adhesion to a 30-kPa Ecad-Fc PA gel required Cdc42- and formin-dependent filopodia formation, whereas adhesion to a 60-kPa Ecad-Fc PA gel induced Arp2/3-dependent lamellipodial protrusions. A quantitative 3D cell–cell adhesion assay and live cell imaging of cell–cell contact formation revealed that inhibition of Cdc42, formin, and Arp2/3 activities blocked the initiation, but not the maintenance of established cell–cell adhesions. These results indicate that the same signaling molecules activated by E-cadherin rigidity sensing on PA gels contribute to actin organization and membrane dynamics during cell–cell adhesion. We hypothesize that a transition in the stiffness of E-cadherin homotypic interactions regulates actin and membrane dynamics during initial stages of cell–cell adhesion. PMID:28674019

Mitochondrial reactive oxygen species regulate the strength of inhibitory GABA-mediated synaptic transmission

NASA Astrophysics Data System (ADS)

Accardi, Michael V.; Daniels, Bryan A.; Brown, Patricia M. G. E.; Fritschy, Jean-Marc; Tyagarajan, Shiva K.; Bowie, Derek

2014-01-01

Neuronal communication imposes a heavy metabolic burden in maintaining ionic gradients essential for action potential firing and synaptic signalling. Although cellular metabolism is known to regulate excitatory neurotransmission, it is still unclear whether the brain’s energy supply affects inhibitory signalling. Here we show that mitochondrial-derived reactive oxygen species (mROS) regulate the strength of postsynaptic GABAA receptors at inhibitory synapses of cerebellar stellate cells. Inhibition is strengthened through a mechanism that selectively recruits α3-containing GABAA receptors into synapses with no discernible effect on resident α1-containing receptors. Since mROS promotes the emergence of postsynaptic events with unique kinetic properties, we conclude that newly recruited α3-containing GABAA receptors are activated by neurotransmitter released onto discrete postsynaptic sites. Although traditionally associated with oxidative stress in neurodegenerative disease, our data identify mROS as a putative homeostatic signalling molecule coupling cellular metabolism to the strength of inhibitory transmission.

Quantitative comparison of cancer and normal cell adhesion using organosilane monolayer templates: an experimental study on the anti-adhesion effect of green-tea catechins.

PubMed

Sakamoto, Rumi; Kakinuma, Eisuke; Masuda, Kentaro; Takeuchi, Yuko; Ito, Kosaku; Iketaki, Kentaro; Matsuzaki, Takahisa; Nakabayashi, Seiichiro; Yoshikawa, Hiroshi Y; Yamamoto, Hideaki; Sato, Yuko; Tanii, Takashi

2016-09-01

The main constituent of green tea, (-)-Epigallocatechin-3-O-gallate (EGCG), is known to have cancer-specific chemopreventive effects. In the present work, we investigated how EGCG suppresses cell adhesion by comparing the adhesion of human pancreatic cancer cells (AsPC-1 and BxPC-3) and their counterpart, normal human embryonic pancreas-derived cells (1C3D3), in catechin-containing media using organosilane monolayer templates (OMTs). The purpose of this work is (1) to evaluate the quantitativeness in the measurement of cell adhesion with the OMT and (2) to show how green-tea catechins suppress cell adhesion in a cancer-specific manner. For the first purpose, the adhesion of cancer and normal cells was compared using the OMT. The cell adhesion in different type of catechins such as EGCG, (-)-Epicatechin-3-O-gallate (ECG) and (-)-Epicatechin (EC) was also evaluated. The measurements revealed that the anti-adhesion effect of green-tea catechins is cancer-specific, and the order is EGCG≫ECG>EC. The results agree well with the data reported to date, showing the quantitativeness of the new method. For the second purpose, the contact area of cells on the OMT was measured by reflection interference contrast microscopy. The cell-OMT contact area of cancer cells decreases with increasing EGCG concentration, whereas that of normal cells remains constant. The results reveal a twofold action of EGCG on cancer cell adhesion-suppressing cell attachment to a candidate adhesion site and decreasing the contact area of the cells-and validates the use of OMT as a tool for screening cancer cell adhesion.

Fine structure of synapses of the central nervous system in resinless sections.

PubMed

Cohen, R S; Wolosewick, J J; Becker, R P; Pappas, G D

1983-10-01

The cytoskeleton has been implicated in neuronal function, particularly in axonal transport, excitability at axonal membranes, and movement of synaptic vesicles at preganglionic endings. The present study demonstrates the presence of a pre- and postsynaptic cytoskeleton in resinless sections of CNS tissue by use of the polyethylene glycol (PEG) technique of Wolosewick (1980) viewed by conventional transmission EM, scanning transmission EM, and surface scanning EM. The PEG technique permits visualization of the cytoskeletal network unobscured by the electron scattering properties of epoxy embedment. In the presynaptic process, synaptic vesicles appear to be suspended in a filamentous network that is contiguous with the synaptic vesicle membrane and with the presynaptic plasma membrane and its dense material. In the postsynaptic process, the postsynaptic density (PSD) is seen in intimate contact with the postsynaptic membrane. En face images of the PSD in some synapses appear as a torus. Emanating from the filamentous web of the PSD are filaments which extend to the adjacent plasma membrane. We conclude that membranous synaptic elements are contiguous with a three-dimensional lattice network that is similar to that described in whole unembedded cells (Wolosewick and Porter, 1976). Moreover, the synaptic densities represent a specialized elaboration of the cytoskeleton.

Transfer characteristics of the hair cell's afferent synapse

NASA Astrophysics Data System (ADS)

Keen, Erica C.; Hudspeth, A. J.

2006-04-01

The sense of hearing depends on fast, finely graded neurotransmission at the ribbon synapses connecting hair cells to afferent nerve fibers. The processing that occurs at this first chemical synapse in the auditory pathway determines the quality and extent of the information conveyed to the central nervous system. Knowledge of the synapse's input-output function is therefore essential for understanding how auditory stimuli are encoded. To investigate the transfer function at the hair cell's synapse, we developed a preparation of the bullfrog's amphibian papilla. In the portion of this receptor organ representing stimuli of 400-800 Hz, each afferent nerve fiber forms several synaptic terminals onto one to three hair cells. By performing simultaneous voltage-clamp recordings from presynaptic hair cells and postsynaptic afferent fibers, we established that the rate of evoked vesicle release, as determined from the average postsynaptic current, depends linearly on the amplitude of the presynaptic Ca2+ current. This result implies that, for receptor potentials in the physiological range, the hair cell's synapse transmits information with high fidelity. auditory system | exocytosis | glutamate | ribbon synapse | synaptic vesicle

Mechanisms of polarized membrane trafficking in neurons – focusing in on endosomes

PubMed Central

Lasiecka, Zofia M.; Winckler, Bettina

2011-01-01

Neurons are polarized cells that have a complex and unique morphology: long processes (axons and dendrites) extending far from the cell body. In addition, the somatodendritic and axonal domains are further divided into specific subdomains, such as synapses (pre- and postsynaptic specializations), proximal and distal dendrites, axon initial segments, nodes of Ranvier, and axon growth cones. The striking asymmetry and complexity of neuronal cells is necessary for their function in receiving, processing and transferring electrical signals, with each domain playing a precise function in these processes. In order to establish and maintain distinct neuronal domains, mechanisms must exist for protein delivery to specific neuronal compartments, such that each compartment has the correct functional molecular composition. How polarized membrane domains are established and maintained is a long-standing question. Transmembrane proteins, such as receptors and adhesion molecules, can be transported to their proper membrane domains by several pathways. The biosynthetic secretory system delivers newly synthesized transmembrane proteins from the ER-Golgi via the trans-Golgi network (TGN) to the plasma membrane. In addition, the endosomal system is critically involved in many instances in ensuring proper (re)targeting of membrane components because it can internalize and degrade mislocalized proteins, or recycle proteins from one domain to another. The endosomal system is thus crucial for establishing and maintaining neuronal polarity. In this review, we focus mainly on the intracellular compartments that serve as sorting stations for polarized transport, with particular emphasis on the emerging roles of endosomes. PMID:21762782

Removal of area CA3 from hippocampal slices induces postsynaptic plasticity at Schaffer collateral synapses that normalizes CA1 pyramidal cell discharge.

PubMed

Dumas, Theodore C; Uttaro, Michael R; Barriga, Carolina; Brinkley, Tiffany; Halavi, Maryam; Wright, Susan N; Ferrante, Michele; Evans, Rebekah C; Hawes, Sarah L; Sanders, Erin M

2018-06-21

Neural networks that undergo acute insults display remarkable reorganization. This injury related plasticity is thought to permit recovery of function in the face of damage that cannot be reversed. Previously, an increase in the transmission strength at Schaffer collateral to CA1 pyramidal cell synapses was observed after long-term activity reduction in organotypic hippocampal slices. Here we report that, following acute preparation of adult rat hippocampal slices and surgical removal of area CA3, input to area CA1 was reduced and Schaffer collateral synapses underwent functional strengthening. This increase in synaptic strength was limited to Schaffer collateral inputs (no alteration to temporoammonic synapses) and acted to normalize postsynaptic discharge, supporting a homeostatic or compensatory response. Short-term plasticity was not altered, but an increase in immunohistochemical labeling of GluA1 subunits was observed in the stratum radiatum (but not stratum moleculare), suggesting increased numbers of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors and a postsynaptic locus of expression. Combined, these data support the idea that, in response to the reduction in presynaptic activity caused by removal of area CA3, Schaffer collateral synapses undergo a relatively rapid increase in functional efficacy likely supported by insertion of more AMPARs, which maintains postsynaptic excitability in CA1 pyramidal neurons. This novel fast compensatory plasticity exhibits properties that would allow it to maintain optimal network activity levels in the hippocampus, a brain structure lauded for its ongoing experience-dependent malleability. Copyright © 2018 Elsevier B.V. All rights reserved.

An Elmo–Dock complex locally controls Rho GTPases and actin remodeling during cadherin-mediated adhesion

PubMed Central

Collins, Caitlin

2014-01-01

Cell–cell contact formation is a dynamic process requiring the coordination of cadherin-based cell–cell adhesion and integrin-based cell migration. A genome-wide RNA interference screen for proteins required specifically for cadherin-dependent cell–cell adhesion identified an Elmo–Dock complex. This was unexpected as Elmo–Dock complexes act downstream of integrin signaling as Rac guanine-nucleotide exchange factors. In this paper, we show that Elmo2 recruits Dock1 to initial cell–cell contacts in Madin–Darby canine kidney cells. At cell–cell contacts, both Elmo2 and Dock1 are essential for the rapid recruitment and spreading of E-cadherin, actin reorganization, localized Rac and Rho GTPase activities, and the development of strong cell–cell adhesion. Upon completion of cell–cell adhesion, Elmo2 and Dock1 no longer localize to cell–cell contacts and are not required subsequently for the maintenance of cell–cell adhesion. These studies show that Elmo–Dock complexes are involved in both integrin- and cadherin-based adhesions, which may help to coordinate the transition of cells from migration to strong cell–cell adhesion. PMID:25452388

GluD1 is a common altered player in neuronal differentiation from both MECP2-mutated and CDKL5-mutated iPS cells.

PubMed

Livide, Gabriella; Patriarchi, Tommaso; Amenduni, Mariangela; Amabile, Sonia; Yasui, Dag; Calcagno, Eleonora; Lo Rizzo, Caterina; De Falco, Giulia; Ulivieri, Cristina; Ariani, Francesca; Mari, Francesca; Mencarelli, Maria Antonietta; Hell, Johannes Wilhelm; Renieri, Alessandra; Meloni, Ilaria

2015-02-01

Rett syndrome is a monogenic disease due to de novo mutations in either MECP2 or CDKL5 genes. In spite of their involvement in the same disease, a functional interaction between the two genes has not been proven. MeCP2 is a transcriptional regulator; CDKL5 encodes for a kinase protein that might be involved in the regulation of gene expression. Therefore, we hypothesized that mutations affecting the two genes may lead to similar phenotypes by dysregulating the expression of common genes. To test this hypothesis we used induced pluripotent stem (iPS) cells derived from fibroblasts of one Rett patient with a MECP2 mutation (p.Arg306Cys) and two patients with mutations in CDKL5 (p.Gln347Ter and p.Thr288Ile). Expression profiling was performed in CDKL5-mutated cells and genes of interest were confirmed by real-time RT-PCR in both CDKL5- and MECP2-mutated cells. The only major change in gene expression common to MECP2- and CDKL5-mutated cells was for GRID1, encoding for glutamate D1 receptor (GluD1), a member of the δ-family of ionotropic glutamate receptors. GluD1 does not form AMPA or NMDA glutamate receptors. It acts like an adhesion molecule by linking the postsynaptic and presynaptic compartments, preferentially inducing the inhibitory presynaptic differentiation of cortical neurons. Our results demonstrate that GRID1 expression is downregulated in both MECP2- and CDKL5-mutated iPS cells and upregulated in neuronal precursors and mature neurons. These data provide novel insights into disease pathophysiology and identify possible new targets for therapeutic treatment of Rett syndrome.

GluD1 is a common altered player in neuronal differentiation from both MECP2-mutated and CDKL5-mutated iPS cells

PubMed Central

Livide, Gabriella; Patriarchi, Tommaso; Amenduni, Mariangela; Amabile, Sonia; Yasui, Dag; Calcagno, Eleonora; Lo Rizzo, Caterina; De Falco, Giulia; Ulivieri, Cristina; Ariani, Francesca; Mari, Francesca; Mencarelli, Maria Antonietta; Hell, Johannes Wilhelm; Renieri, Alessandra; Meloni, Ilaria

2015-01-01

Rett syndrome is a monogenic disease due to de novo mutations in either MECP2 or CDKL5 genes. In spite of their involvement in the same disease, a functional interaction between the two genes has not been proven. MeCP2 is a transcriptional regulator; CDKL5 encodes for a kinase protein that might be involved in the regulation of gene expression. Therefore, we hypothesized that mutations affecting the two genes may lead to similar phenotypes by dysregulating the expression of common genes. To test this hypothesis we used induced pluripotent stem (iPS) cells derived from fibroblasts of one Rett patient with a MECP2 mutation (p.Arg306Cys) and two patients with mutations in CDKL5 (p.Gln347Ter and p.Thr288Ile). Expression profiling was performed in CDKL5-mutated cells and genes of interest were confirmed by real-time RT-PCR in both CDKL5- and MECP2-mutated cells. The only major change in gene expression common to MECP2- and CDKL5-mutated cells was for GRID1, encoding for glutamate D1 receptor (GluD1), a member of the δ-family of ionotropic glutamate receptors. GluD1 does not form AMPA or NMDA glutamate receptors. It acts like an adhesion molecule by linking the postsynaptic and presynaptic compartments, preferentially inducing the inhibitory presynaptic differentiation of cortical neurons. Our results demonstrate that GRID1 expression is downregulated in both MECP2- and CDKL5-mutated iPS cells and upregulated in neuronal precursors and mature neurons. These data provide novel insights into disease pathophysiology and identify possible new targets for therapeutic treatment of Rett syndrome. PMID:24916645

Nanowell-Trapped Charged Ligand-Bearing Nanoparticle Surfaces – A Novel Method of Enhancing Flow-Resistant Cell Adhesion

PubMed Central

Tran, Phat L.; Gamboa, Jessica R.; McCracken, Katherine E.; Riley, Mark R.

2014-01-01

Assuring cell adhesion to an underlying biomaterial surface is vital in implant device design and tissue engineering, particularly under circumstances where cells are subjected to potential detachment from overriding fluid flow. Cell-substrate adhesion is a highly regulated process involving the interplay of mechanical properties, surface topographic features, electrostatic charge, and biochemical mechanisms. At the nanoscale level the physical properties of the underlying substrate are of particular importance in cell adhesion. Conventionally, natural, pro-adhesive, and often thrombogenic, protein biomaterials are frequently utilized to facilitate adhesion. In the present study nanofabrication techniques are utilized to enhance the biological functionality of a synthetic polymer surface, polymethymethacrylate, with respect to cell adhesion. Specifically we examine the effect on cell adhesion of combining: 1. optimized surface texturing, 2. electrostatic charge and 3. cell adhesive ligands, uniquely assembled on the substrata surface, as an ensemble of nanoparticles trapped in nanowells. Our results reveal that the ensemble strategy leads to enhanced, more than simply additive, endothelial cell adhesion under both static and flow conditions. This strategy may be of particular utility for enhancing flow-resistant endothelialization of blood-contacting surfaces of cardiovascular devices subjected to flow-mediated shear. PMID:23225491

USP5/Leon deubiquitinase confines postsynaptic growth by maintaining ubiquitin homeostasis through Ubiquilin.

PubMed

Wang, Chien-Hsiang; Huang, Yi-Chun; Chen, Pei-Yi; Cheng, Ying-Ju; Kao, Hsiu-Hua; Pi, Haiwei; Chien, Cheng-Ting

2017-05-10

Synapse formation and growth are tightly controlled processes. How synaptic growth is terminated after reaching proper size remains unclear. Here, we show that Leon, the Drosophila USP5 deubiquitinase, controls postsynaptic growth. In leon mutants, postsynaptic specializations of neuromuscular junctions are dramatically expanded, including the subsynaptic reticulum, the postsynaptic density, and the glutamate receptor cluster. Expansion of these postsynaptic features is caused by a disruption of ubiquitin homeostasis with accumulation of free ubiquitin chains and ubiquitinated substrates in the leon mutant. Accumulation of Ubiquilin (Ubqn), the ubiquitin receptor whose human homolog ubiquilin 2 is associated with familial amyotrophic lateral sclerosis, also contributes to defects in postsynaptic growth and ubiquitin homeostasis. Importantly, accumulations of postsynaptic proteins cause different aspects of postsynaptic overgrowth in leon mutants. Thus, the deubiquitinase Leon maintains ubiquitin homeostasis and proper Ubqn levels, preventing postsynaptic proteins from accumulation to confine postsynaptic growth.

Cholinergic interneurons in the feeding system of the pond snail Lymnaea stagnalis. II. N1 interneurons make cholinergic synapses with feeding motoneurons.

PubMed

Elliott, C J; Kemenes, G

1992-05-29

The N1 neurons are a population of interneurons active during the protraction phase of the feeding rhythm. All the N1 neurons are coupled by electrical synapses which persist in a high Mg/low Ca saline which blocks chemical synapses. Individual N1 spikes produce discrete electrotonic postsynaptic potentials (PSPS) in other N1 cells, but the coupling is not strong enough to ensure 1:1 firing. Bursts of N1 spikes generate compound PSPS in the feeding motoneurons. The sign (excitation or inhibition) of the N1 input corresponds with the synaptic barrage recorded during the protraction phase. Discrete PSPS are only resolved in a Hi-Di saline. Their variation in latency and number can be explained by variation in electrotonic propagation within the electrically coupled network of N1 cells. The excitatory postsynaptic potentials (ESPS) in the 1 cell are reduced by 0.5 mM antagonists hexamethonium (HMT), atropine (ATR), curare (d-TC) and by methylxylocholine (MeXCh), all of which block the excitatory cholinergic receptor (Elliott et al. (Phil. Trans. R. Soc. Lond. 336, 157-166 (Preceding paper.) (1992)). The 1 cell EPSPS were transiently blocked by phenyltrimethylammonium (PTMA), which is both an agonist and antagonist at the 1 cell excitatory acetylcholine (ACh) receptor (Elliott et al. 1992). The inhibitory postsynaptic potential (IPSP) in the 3 cell is blocked by bath applications of MeXCh and PTMA, which both abolish the response of the 3 cell to ACh (Elliott et. al. 1992). The effects of the cholinergic antagonists on the response of 4 cluster and 5 cells to N1 stimulation matches their response to ACh (Elliott et al. 1992). It is concluded that the population of N1 cells are multiaction, premotor cholinergic interneurons.

An Inducible Endothelial Cell Surface Glycoprotein Mediates Melanoma Adhesion

NASA Astrophysics Data System (ADS)

Rice, G. Edgar; Bevilacqua, Michael P.

1989-12-01

Hematogenous metastasis requires the arrest and extravasation of blood-borne tumor cells, possibly involving direct adhesive interactions with vascular endothelium. Cytokine activation of cultured human endothelium increases adhesion of melanoma and carcinoma cell lines. An inducible 110-kD endothelial cell surface glycoprotein, designated INCAM-110, appears to mediate adhesion of melanoma cells. In addition, an inducible endothelial receptor for neutrophils, ELAM-1, supports the adhesion of a human colon carcinoma cell line. Thus, activation of vascular endothelium in vivo that results in increased expression of INCAM-110 and ELAM-1 may promote tumor cell adhesion and affect the incidence and distribution of metastases.

9-cis-Retinoic Acid Promotes Cell Adhesion Through Integrin Dependent and Independent Mechanisms Across Immune Lineages

PubMed Central

Whelan, Jarrett T.; Chen, Jianming; Miller, Jabin; Morrow, Rebekah L.; Lingo, Joshuah D.; Merrell, Kaitlin; Shaikh, Saame Raza; Bridges, Lance C.

2012-01-01

Retinoids are essential in the proper establishment and maintenance of immunity. Although retinoids are implicated in immune related processes, their role in immune cell adhesion has not been well established. In this study, the effect of 9-cis-retinoic acid (9-cis-RA) on human hematopoietic cell adhesion was investigated. 9-cis-RA treatment specifically induced cell adhesion of the human immune cell lines HuT-78, NB4, RPMI 8866, and U937. Due to the prominent role of integrin receptors in mediating immune cell adhesion, we sought to evaluate if cell adhesion was integrin-dependent. By employing a variety of integrin antagonist including function-blocking antibodies and EDTA, we establish that 9-cis-RA prompts immune cell adhesion through established integrin receptors in addition to a novel integrin-independent process. The novel integrin-independent adhesion required the presence of retinoid and was attenuated by treatment with synthetic corticosteroids. Finally, we demonstrate that 9-cis-RA treatment of primary murine B-cells induces ex vivo adhesion that persists in the absence of integrin function. Our study is the first to demonstrate that 9-cis-retinoic acid influences immune cell adhesion through at least two functionally distinct mechanisms. PMID:22925918

Role of cellular adhesions in tissue dynamics spectroscopy

NASA Astrophysics Data System (ADS)

Merrill, Daniel A.; An, Ran; Turek, John; Nolte, David

2014-02-01

Cellular adhesions play a critical role in cell behavior, and modified expression of cellular adhesion compounds has been linked to various cancers. We tested the role of cellular adhesions in drug response by studying three cellular culture models: three-dimensional tumor spheroids with well-developed cellular adhesions and extracellular matrix (ECM), dense three-dimensional cell pellets with moderate numbers of adhesions, and dilute three-dimensional cell suspensions in agarose having few adhesions. Our technique for measuring the drug response for the spheroids and cell pellets was biodynamic imaging (BDI), and for the suspensions was quasi-elastic light scattering (QELS). We tested several cytoskeletal chemotherapeutic drugs (nocodazole, cytochalasin-D, paclitaxel, and colchicine) on three cancer cell lines chosen from human colorectal adenocarcinoma (HT-29), human pancreatic carcinoma (MIA PaCa-2), and rat osteosarcoma (UMR-106) to exhibit differences in adhesion strength. Comparing tumor spheroid behavior to that of cell suspensions showed shifts in the spectral motion of the cancer tissues that match predictions based on different degrees of cell-cell contacts. The HT-29 cell line, which has the strongest adhesions in the spheroid model, exhibits anomalous behavior in some cases. These results highlight the importance of using three-dimensional tissue models in drug screening with cellular adhesions being a contributory factor in phenotypic differences between the drug responses of tissue and cells.

Pathway analysis of genome-wide association datasets of personality traits.

PubMed

Kim, H-N; Kim, B-H; Cho, J; Ryu, S; Shin, H; Sung, J; Shin, C; Cho, N H; Sung, Y A; Choi, B-O; Kim, H-L

2015-04-01

Although several genome-wide association (GWA) studies of human personality have been recently published, genetic variants that are highly associated with certain personality traits remain unknown, due to difficulty reproducing results. To further investigate these genetic variants, we assessed biological pathways using GWA datasets. Pathway analysis using GWA data was performed on 1089 Korean women whose personality traits were measured with the Revised NEO Personality Inventory for the 5-factor model of personality. A total of 1042 pathways containing 8297 genes were included in our study. Of these, 14 pathways were highly enriched with association signals that were validated in 1490 independent samples. These pathways include association of: Neuroticism with axon guidance [L1 cell adhesion molecule (L1CAM) interactions]; Extraversion with neuronal system and voltage-gated potassium channels; Agreeableness with L1CAM interaction, neurotransmitter receptor binding and downstream transmission in postsynaptic cells; and Conscientiousness with the interferon-gamma and platelet-derived growth factor receptor beta polypeptide pathways. Several genes that contribute to top-ranked pathways in this study were previously identified in GWA studies or by pathway analysis in schizophrenia or other neuropsychiatric disorders. Here we report the first pathway analysis of all five personality traits. Importantly, our analysis identified novel pathways that contribute to understanding the etiology of personality traits. © 2015 The Authors. Genes, Brain and Behavior published by International Behavioural and Neural Genetics Society and John Wiley & Sons Ltd.

Presynaptic congenital myasthenic syndrome with a homozygous sequence variant in LAMA5 combines myopia, facial tics, and failure of neuromuscular transmission.

PubMed

Maselli, Ricardo A; Arredondo, Juan; Vázquez, Jessica; Chong, Jessica X; Bamshad, Michael J; Nickerson, Deborah A; Lara, Marian; Ng, Fiona; Lo, Victoria L; Pytel, Peter; McDonald, Craig M

2017-08-01

Defects in genes encoding the isoforms of the laminin alpha subunit have been linked to various phenotypic manifestations, including brain malformations, muscular dystrophy, ocular defects, cardiomyopathy, and skin abnormalities. We report here a severe defect of neuromuscular transmission in a consanguineous patient with a homozygous variant in the laminin alpha-5 subunit gene (LAMA5). The variant c.8046C>T (p.Arg2659Trp) is rare and has a predicted deleterious effect. The affected individual, who also carries a rare homozygous sequence variant in LAMA1, had muscle weakness, myopia, and facial tics. Magnetic resonance imaging of brain showed mild volume loss and periventricular T2 prolongation. Repetitive nerve stimulation revealed 50% decrement of compound muscle action potential amplitudes and 250% facilitation immediately after exercise, Endplate studies identified a profound reduction of the endplate potential quantal content and endplates with normal postsynaptic folding that were denuded or partially occupied by small nerve terminals. Expression studies revealed that p.Arg2659Trp caused decreased binding of laminin alpha-5 to SV2A and impaired laminin-521 cell-adhesion and cell projection support in primary neuronal cultures. In summary, this report describing severe neuromuscular transmission failure in a patient with a LAMA5 mutation expands the list of phenotypes associated with defects in genes encoding alpha-laminins. © 2017 Wiley Periodicals, Inc.

Crystal Structure of the Extracellular Cholinesterase-Like Domain from Neuroligin-2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koehnke,J.; Jin, X.; Budreck, E.

Neuroligins (NLs) are catalytically inactive members of a family of cholinesterase-like transmembrane proteins that mediate cell adhesion at neuronal synapses. Postsynaptic neuroligins engage in Ca2+-dependent transsynaptic interactions via their extracellular cholinesterase domain with presynaptic neurexins (NRXs). These interactions may be regulated by two short splice insertions (termed A and B) in the NL cholinesterase domain. Here, we present the 3.3- Angstroms crystal structure of the ectodomain from NL2 containing splice insertion A (NL2A). The overall structure of NL2A resembles that of cholinesterases, but several structural features are unique to the NL proteins. First, structural elements surrounding the esterase active-site regionmore » differ significantly between active esterases and NL2A. On the opposite surface of the NL2A molecule, the positions of the A and B splice insertions identify a candidate NRX interaction site of the NL protein. Finally, sequence comparisons of NL isoforms allow for mapping the location of residues of previously identified mutations in NL3 and NL4 found in patients with autism spectrum disorders. Overall, the NL2 structure promises to provide a valuable model for dissecting NL isoform- and synapse-specific functions.« less

Integrating evolutionary and regulatory information with a multispecies approach implicates genes and pathways in obsessive-compulsive disorder.

PubMed

Noh, Hyun Ji; Tang, Ruqi; Flannick, Jason; O'Dushlaine, Colm; Swofford, Ross; Howrigan, Daniel; Genereux, Diane P; Johnson, Jeremy; van Grootheest, Gerard; Grünblatt, Edna; Andersson, Erik; Djurfeldt, Diana R; Patel, Paresh D; Koltookian, Michele; M Hultman, Christina; Pato, Michele T; Pato, Carlos N; Rasmussen, Steven A; Jenike, Michael A; Hanna, Gregory L; Stewart, S Evelyn; Knowles, James A; Ruhrmann, Stephan; Grabe, Hans-Jörgen; Wagner, Michael; Rück, Christian; Mathews, Carol A; Walitza, Susanne; Cath, Daniëlle C; Feng, Guoping; Karlsson, Elinor K; Lindblad-Toh, Kerstin

2017-10-17

Obsessive-compulsive disorder is a severe psychiatric disorder linked to abnormalities in glutamate signaling and the cortico-striatal circuit. We sequenced coding and regulatory elements for 608 genes potentially involved in obsessive-compulsive disorder in human, dog, and mouse. Using a new method that prioritizes likely functional variants, we compared 592 cases to 560 controls and found four strongly associated genes, validated in a larger cohort. NRXN1 and HTR2A are enriched for coding variants altering postsynaptic protein-binding domains. CTTNBP2 (synapse maintenance) and REEP3 (vesicle trafficking) are enriched for regulatory variants, of which at least six (35%) alter transcription factor-DNA binding in neuroblastoma cells. NRXN1 achieves genome-wide significance (p = 6.37 × 10 -11 ) when we include 33,370 population-matched controls. Our findings suggest synaptic adhesion as a key component in compulsive behaviors, and show that targeted sequencing plus functional annotation can identify potentially causative variants, even when genomic data are limited.Obsessive-compulsive disorder (OCD) is a neuropsychiatric disorder with symptoms including intrusive thoughts and time-consuming repetitive behaviors. Here Noh and colleagues identify genes enriched for functional variants associated with increased risk of OCD.

The possible interplay of synaptic and clock genes in autism spectrum disorders.

PubMed

Bourgeron, T

2007-01-01

Autism spectrum disorders (ASD) are complex neurodevelopmental conditions characterized by deficits in social communication, absence or delay in language, and stereotyped and repetitive behaviors. Results from genetic studies reveal one pathway associated with susceptibility to ASD, which includes the synaptic cell adhesion molecules NLGN3, NLGN4, and NRXN1 and a postsynaptic scaffolding protein SHANK3. This protein complex is crucial for the maintenance of functional synapses as well as the adequate balance between neuronal excitation and inhibition. Among the factors that could modulate this pathway are the genes controlling circadian rhythms. Indeed, sleep disorders and low melatonin levels are frequently observed in ASD. In this context, an alteration of both this synaptic pathway and the setting of the clock would greatly increase the risk of ASD. In this chapter, I report genetic and neurobiological findings that highlight the major role of synaptic and clock genes in the susceptibility to ASD. On the basis of these lines of evidence, I propose that future studies of ASD should investigate the circadian modulation of synaptic function as a focus for functional analyses and the development of new therapeutic strategies.

Crystal structure of the extracellular cholinesterase-like domain from neuroligin-2

PubMed Central

Koehnke, Jesko; Jin, Xiangshu; Budreck, Elaine C.; Posy, Shoshana; Scheiffele, Peter; Honig, Barry; Shapiro, Lawrence

2008-01-01

Neuroligins (NLs) are catalytically inactive members of a family of cholinesterase-like transmembrane proteins that mediate cell adhesion at neuronal synapses. Postsynaptic neuroligins engage in Ca2+-dependent transsynaptic interactions via their extracellular cholinesterase domain with presynaptic neurexins (NRXs). These interactions may be regulated by two short splice insertions (termed A and B) in the NL cholinesterase domain. Here, we present the 3.3-Å crystal structure of the ectodomain from NL2 containing splice insertion A (NL2A). The overall structure of NL2A resembles that of cholinesterases, but several structural features are unique to the NL proteins. First, structural elements surrounding the esterase active-site region differ significantly between active esterases and NL2A. On the opposite surface of the NL2A molecule, the positions of the A and B splice insertions identify a candidate NRX interaction site of the NL protein. Finally, sequence comparisons of NL isoforms allow for mapping the location of residues of previously identified mutations in NL3 and NL4 found in patients with autism spectrum disorders. Overall, the NL2 structure promises to provide a valuable model for dissecting NL isoform- and synapse-specific functions. PMID:18250328

Biological adhesion of the flatworm Macrostomum lignano relies on a duo-gland system and is mediated by a cell type-specific intermediate filament protein.

PubMed

Lengerer, Birgit; Pjeta, Robert; Wunderer, Julia; Rodrigues, Marcelo; Arbore, Roberto; Schärer, Lukas; Berezikov, Eugene; Hess, Michael W; Pfaller, Kristian; Egger, Bernhard; Obwegeser, Sabrina; Salvenmoser, Willi; Ladurner, Peter

2014-02-12

Free-living flatworms, in both marine and freshwater environments, are able to adhere to and release from a substrate several times within a second. This reversible adhesion relies on adhesive organs comprised of three cell types: an adhesive gland cell, a releasing gland cell, and an anchor cell, which is a modified epidermal cell responsible for structural support. However, nothing is currently known about the molecules that are involved in this adhesion process. In this study we present the detailed morphology of the adhesive organs of the free-living marine flatworm Macrostomum lignano. About 130 adhesive organs are located in a horse-shoe-shaped arc along the ventral side of the tail plate. Each organ consists of exactly three cells, an adhesive gland cell, a releasing gland cell, and an anchor cell. The necks of the two gland cells penetrate the anchor cell through a common pore. Modified microvilli of the anchor cell form a collar surrounding the necks of the adhesive- and releasing glands, jointly forming the papilla, the outer visible part of the adhesive organs. Next, we identified an intermediate filament (IF) gene, macif1, which is expressed in the anchor cells. RNA interference mediated knock-down resulted in the first experimentally induced non-adhesion phenotype in any marine animal. Specifically, the absence of intermediate filaments in the anchor cells led to papillae with open tips, a reduction of the cytoskeleton network, a decline in hemidesmosomal connections, and to shortened microvilli containing less actin. Our findings reveal an elaborate biological adhesion system in a free-living flatworm, which permits impressively rapid temporary adhesion-release performance in the marine environment. We demonstrate that the structural integrity of the supportive cell, the anchor cell, is essential for this adhesion process: the knock-down of the anchor cell-specific intermediate filament gene resulted in the inability of the animals to adhere. The RNAi mediated changes of the anchor cell morphology are comparable to situations observed in human gut epithelia. Therefore, our current findings and future investigations using this powerful flatworm model system might contribute to a better understanding of the function of intermediate filaments and their associated human diseases.

[Expression of cell adhesion molecules in acute leukemia cell].

PubMed

Ju, Xiaoping; Peng, Min; Xu, Xiaoping; Lu, Shuqing; Li, Yao; Ying, Kang; Xie, Yi; Mao, Yumin; Xia, Fang

2002-11-01

To investigate the role of cell adhesion molecule in the development and extramedullary infiltration (EI) of acute leukemia. The expressions of neural cell adhesion molecule (NCAM) gene, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule (VCAM-1) genes in 25 acute leukemia patients bone marrow cells were detected by microarray and reverse transcriptase-polymerase chain reaction (RT-PCR). The expressions of NCAM, ICAM-1 and VCAM-1 gene were significantly higher in acute leukemia cells and leukemia cells with EI than in normal tissues and leukemia cells without EI, respectively, both by cDNA microarray and by RT-PCR. The cDNA microarray is a powerful technique in analysis of acute leukemia cells associated genes. High expressions of cell adhesion molecule genes might be correlated with leukemia pathogenesis and infiltration of acute leukemia cell.

Preliminary evidence for a postsynaptic action of beta-bungarotoxin in mammalian skeletal muscle

NASA Technical Reports Server (NTRS)

Storella, R. J.; Schouchoff, A. L.; Fujii, M.; Hill, J.; Fletcher, J. E.; Jiang, M. S.; Smith, L. A.

1992-01-01

Two hours after treatment with beta-bungarotoxin (0.34-0.4 microM), when there was complete neuromuscular block, the peak contracture response to 50 microM succinylcholine was significantly reduced by about 35% in the mouse phrenic nerve-diaphragm preparation. Additionally, significant phospholipase A2 activity was detected on primary cell cultures from skeletal muscle which were incubated for 2 hr with concentrations of beta-bungarotoxin greater than or equal to 0.1 microM. Thus, beta-bungarotoxin appears to have pharmacologically and biochemically detectable postsynaptic actions in mammalian muscle systems.

Decrement of GABAA receptor-mediated inhibitory postsynaptic currents in dentate granule cells in epileptic hippocampus.

PubMed

Isokawa, M

1996-05-01

1. Inhibitory postsynaptic currents (IPSCs) were studied in hippocampal dentate granule cells (DGCs) in the pilocarpine model and human temporal lobe epilepsy, with the use of the whole cell patch-clamp recording technique in slice preparations. 2. In the pilocarpine model, hippocampal slices were prepared from rats that were allowed to experience spontaneous seizures for 2 mo. Human hippocampal specimens were obtained from epileptic patients who underwent surgical treatment for medically intractable seizures. 3. IPSCs were generated by single perforant path stimulation and recorded at a membrane potential (Vm) of 0 mV near the reversal potential of glutamate excitatory postsynaptic currents in the voltage-clamp recording. IPSCs were pharmacologically identified as gamma-aminobutyric acid-A (GABAA) IPSCs by 10 microM bicuculline methiodide. 4. During low-frequency stimulation, IPSCs were not different in amplitude among non-seizure-experienced rat hippocampi, human nonsclerotic hippocampi, seizure-experienced rat hippocampi, and human sclerotic hippocampi. In the last two groups of DGCs, current-clamp recordings indicated the presence of prolonged excitatory postsynaptic potentials (EPSPs) mediated by the N-methyl-D-aspartate (NMDA) receptor. 5. High-frequency stimulation, administered at Vm = -30 mV to activate NMDA currents, reduced GABAA IPSC amplitude specifically in seizure-experienced rat hippocampi (t = 2.5, P < 0.03) and human sclerotic hippocampi (t = 7.7, P < 0.01). This reduction was blocked by an NMDA receptor antagonist, 2-amino-5-phosphonovaleric acid (APV) (50 microM). The time for GABAA IPSCs to recover to their original amplitude was also shortened by the application of APV. 6. I conclude that, when intensively activated, NMDA receptor-mediated excitatory transmission may interact with GABAergic synaptic inhibition in DGCs in seizure-experienced hippocampus to transiently reduce GABA(A) receptor-channel function. Such interactions may contribute to give rise to epileptic excitation in chronically seizure-prone hippocampus.

Single-Molecule Discrimination within Dendritic Spines of Discrete Perisynaptic Sites of Actin Filament Assembly Driving Postsynaptic Reorganization

NASA Astrophysics Data System (ADS)

Blanpied, Thomas A.

2013-03-01

In the brain, the strength of synaptic transmission between neurons is principally set by the organization of proteins within the receptive, postsynaptic cell. Synaptic strength at an individual site of contact can remain remarkably stable for months or years. However, it also can undergo diverse forms of plasticity which change the strength at that contact independent of changes to neighboring synapses. Such activity-triggered neural plasticity underlies memory storage and cognitive development, and is disrupted in pathological physiology such as addiction and schizophrenia. Much of the short-term regulation of synaptic plasticity occurs within the postsynaptic cell, in small subcompartments surrounding the synaptic contact. Biochemical subcompartmentalization necessary for synapse-specific plasticity is achieved in part by segregation of synapses to micron-sized protrusions from the cell called dendritic spines. Dendritic spines are heavily enriched in the actin cytoskeleton, and regulation of actin polymerization within dendritic spines controls both basal synaptic strength and many forms of synaptic plasticity. However, understanding the mechanism of this control has been difficult because the submicron dimensions of spines limit examination of actin dynamics in the spine interior by conventional confocal microscopy. To overcome this, we developed single-molecule tracking photoactivated localization microscopy (smtPALM) to measure the movement of individual actin molecules within living spines. This revealed inward actin flow from broad areas of the spine plasma membrane, as well as a dense central core of heterogeneous filament orientation. The velocity of single actin molecules along filaments was elevated in discrete regions within the spine, notably near the postsynaptic density but surprisingly not at the endocytic zone which is involved in some forms of plasticity. We conclude that actin polymerization is initiated at many well-separated foci within spines, an organization that may be necessary for the finely tuned adjustment of synaptic molecular content that underlies functional plasticity. Indeed, further single-molecule mapping studies confirm that actin polymerization drives reorganization of molecular organization at the synapse itself.

The cancer cell adhesion resistome: mechanisms, targeting and translational approaches.

PubMed

Dickreuter, Ellen; Cordes, Nils

2017-06-27

Cell adhesion-mediated resistance limits the success of cancer therapies and is a great obstacle to overcome in the clinic. Since the 1990s, where it became clear that adhesion of tumor cells to the extracellular matrix is an important mediator of therapy resistance, a lot of work has been conducted to understand the fundamental underlying mechanisms and two paradigms were deduced: cell adhesion-mediated radioresistance (CAM-RR) and cell adhesion-mediated drug resistance (CAM-DR). Preclinical work has evidently demonstrated that targeting of integrins, adapter proteins and associated kinases comprising the cell adhesion resistome is a promising strategy to sensitize cancer cells to both radiotherapy and chemotherapy. Moreover, the cell adhesion resistome fundamentally contributes to adaptation mechanisms induced by radiochemotherapy as well as molecular drugs to secure a balanced homeostasis of cancer cells for survival and growth. Intriguingly, this phenomenon provides a basis for synthetic lethal targeted therapies simultaneously administered to standard radiochemotherapy. In this review, we summarize current knowledge about the cell adhesion resistome and highlight targeting strategies to override CAM-RR and CAM-DR.

Single-cell force spectroscopy as a technique to quantify human red blood cell adhesion to subendothelial laminin.

PubMed

Maciaszek, Jamie L; Partola, Kostyantyn; Zhang, Jing; Andemariam, Biree; Lykotrafitis, George

2014-12-18

Single-cell force spectroscopy (SCFS), an atomic force microscopy (AFM)-based assay, enables quantitative study of cell adhesion while maintaining the native state of surface receptors in physiological conditions. Human healthy and pathological red blood cells (RBCs) express a large number of surface proteins which mediate cell-cell interactions, or cell adhesion to the extracellular matrix. In particular, RBCs adhere with high affinity to subendothelial matrix laminin via the basal cell adhesion molecule and Lutheran protein (BCAM/Lu). Here, we established SCFS as an in vitro technique to study human RBC adhesion at baseline and following biochemical treatment. Using blood obtained from healthy human subjects, we recorded adhesion forces from single RBCs attached to AFM cantilevers as the cell was pulled-off of substrates coated with laminin protein. We found that an increase in the overall cell adhesion measured via SCFS is correlated with an increase in the resultant total force measured on 1 µm(2) areas of the RBC membrane. Further, we showed that SCFS can detect significant changes in the adhesive response of RBCs to modulation of the cyclic adenosine monophosphate (cAMP) and protein kinase A (PKA) pathway. Lastly, we identified variability in the RBC adhesion force to laminin amongst the human subjects, suggesting that RBCs maintain diverse levels of active BCAM/Lu adhesion receptors. By using single-cell measurements, we established a powerful new method for the quantitative measurement of single RBC adhesion with specific receptor-mediated binding. Copyright © 2014 Elsevier Ltd. All rights reserved.

Regulation of epithelial and lymphocyte cell adhesion by adenosine deaminase-CD26 interaction.

PubMed Central

Ginés, Silvia; Mariño, Marta; Mallol, Josefa; Canela, Enric I; Morimoto, Chikao; Callebaut, Christian; Hovanessian, Ara; Casadó, Vicent; Lluis, Carmen; Franco, Rafael

2002-01-01

The extra-enzymic function of cell-surface adenosine deaminase (ADA), an enzyme mainly localized in the cytosol but also found on the cell surface of monocytes, B cells and T cells, has lately been the subject of numerous studies. Cell-surface ADA is able to transduce co-stimulatory signals in T cells via its interaction with CD26, an integral membrane protein that acts as ADA-binding protein. The aim of the present study was to explore whether ADA-CD26 interaction plays a role in the adhesion of lymphocyte cells to human epithelial cells. To meet this aim, different lymphocyte cell lines (Jurkat and CEM T) expressing endogenous, or overexpressing human, CD26 protein were tested in adhesion assays to monolayers of colon adenocarcinoma human epithelial cells, Caco-2, which express high levels of cell-surface ADA. Interestingly, the adhesion of Jurkat and CEM T cells to a monolayer of Caco-2 cells was greatly dependent on CD26. An increase by 50% in the cell-to-cell adhesion was found in cells containing higher levels of CD26. Incubation with an anti-CD26 antibody raised against the ADA-binding site or with exogenous ADA resulted in a significant reduction (50-70%) of T-cell adhesion to monolayers of epithelial cells. The role of ADA-CD26 interaction in the lymphocyte-epithelial cell adhesion appears to be mediated by CD26 molecules that are not interacting with endogenous ADA (ADA-free CD26), since SKW6.4 (B cells) that express more cell-surface ADA showed lower adhesion than T cells. Adhesion stimulated by CD26 and ADA is mediated by T cell lymphocyte function-associated antigen. A role for ADA-CD26 interaction in cell-to-cell adhesion was confirmed further in integrin activation assays. FACS analysis revealed a higher expression of activated integrins on T cell lines in the presence of increasing amounts of exogenous ADA. Taken together, these results suggest that the ADA-CD26 interaction on the cell surface has a role in lymphocyte-epithelial cell adhesion. PMID:11772392

Protopine inhibits heterotypic cell adhesion in MDA-MB-231 cells through down-regulation of multi-adhesive factors.

PubMed

He, Kai; Gao, Jian-Li

2014-01-01

A Chinese herb Corydalis yanhusuo W.T. Wang that showed anticancer and anti-angiogenesis effects in our previous studies was presented for further studies. In the present study, we studied the anticancer proliferation and adhesion effects of five alkaloids which were isolated from Corydalis yanhusuo. MTT dose response curves, cell migration assay, cell invasion assay, as well as three types of cell adhesive assay were performed on MDA-MB-231 human breast cancer cells. The mechanism of the compounds on inhibiting heterotypic cell adhesion were further explored by determining the expression of epidermal growth factor receptor (EGFR), Intercellular adhesion molecule 1 (ICAM-1), αv-integrin, β1-integrin and β5-integrin by western blotting assay. In five tested alkaloids, only protopine exhibited anti-adhesive and anti-invasion effects in MDA-MB-231 cells, which contributed to the anti-metastasis effect of Corydalis yanhusuo. The results showed that after treatment with protopine for 90 min, the expression of EGFR, ICAM-1, αv-integrin, β1-integrin and β5-integrin were remarkably reduced. The present results suggest that protopine seems to inhibit the heterotypic cell adhesion between MDA-MB-231 cells, and human umbilical vein endothelial cells by changing the expression of adhesive factors.

Group III metabotropic glutamate receptors and exocytosed protons inhibit L-type calcium currents in cones but not in rods.

PubMed

Hosoi, Nobutake; Arai, Itaru; Tachibana, Masao

2005-04-20

Light responses of photoreceptors (rods and cones) are transmitted to the second-order neurons (bipolar cells and horizontal cells) via glutamatergic synapses located in the outer plexiform layer of the retina. Although it has been well established that postsynaptic group III metabotropic glutamate receptors (mGluRs) of ON bipolar cells contribute to generating the ON signal, presynaptic roles of group III mGluRs remain to be elucidated at this synaptic connection. We addressed this issue by applying the slice patch-clamp technique to the newt retina. OFF bipolar cells and horizontal cells generate a steady inward current in the dark and a transient inward current at light offset, both of which are mediated via postsynaptic non-NMDA receptors. A group III mGluR-specific agonist, L-2-amino-4-phosphonobutyric acid (L-AP-4), inhibited both the steady and off-transient inward currents but did not affect the glutamate-induced current in these postsynaptic neurons. L-AP-4 inhibited the presynaptic L-type calcium current (ICa) in cones by shifting the voltage dependence of activation to more positive membrane potentials. The inhibition of ICa was most prominent around the physiological range of cone membrane potentials. In contrast, L-AP-4 did not affect L-type ICa in rods. Paired recordings from photoreceptors and the synaptically connected second-order neurons confirmed that L-AP-4 inhibited both ICa and glutamate release in cones but not in rods. Furthermore, we found that exocytosed protons also inhibited ICa in cones but not in rods. Selective modulation of ICa in cones may help broaden the dynamic range of synaptic transfer by controlling the amount of transmitter release from cones.

Impaired development of cortico-striatal synaptic connectivity in a cell culture model of Huntington's disease.

PubMed

Buren, Caodu; Parsons, Matthew P; Smith-Dijak, Amy; Raymond, Lynn A

2016-03-01

Huntington's disease (HD) is a genetically inherited neurodegenerative disease caused by a mutation in the gene encoding the huntingtin protein. This mutation results in progressive cell death that is particularly striking in the striatum. Recent evidence indicates that early HD is initially a disease of the synapse, in which subtle alterations in synaptic neurotransmission, particularly at the cortico-striatal (C-S) synapse, can be detected well in advance of cell death. Here, we used a cell culture model in which striatal neurons are co-cultured with cortical neurons, and monitored the development of C-S connectivity up to 21days in vitro (DIV) in cells cultured from either the YAC128 mouse model of HD or the background strain, FVB/N (wild-type; WT) mice. Our data demonstrate that while C-S connectivity in WT co-cultures develops rapidly and continuously from DIV 7 to 21, YAC128 C-S connectivity shows no significant growth from DIV 14 onward. Morphological and electrophysiological data suggest that a combination of pre- and postsynaptic mechanisms contribute to this effect, including a reduction in both the postsynaptic dendritic arborization and the size and replenishment rate of the presynaptic readily releasable pool of excitatory vesicles. Moreover, a chimeric culture strategy confirmed that the most robust impairment in C-S connectivity was only observed when mutant huntingtin was expressed both pre- and postsynaptically. In all, our data demonstrate a progressive HD synaptic phenotype in this co-culture system that may be exploited as a platform for identifying promising therapeutic strategies to prevent early HD-associated synaptopathy. Copyright © 2015 Elsevier Inc. All rights reserved.

Synaptic calcium regulation in hair cells of the chicken basilar papilla.

PubMed

Im, Gi Jung; Moskowitz, Howard S; Lehar, Mohammed; Hiel, Hakim; Fuchs, Paul Albert

2014-12-10

Cholinergic inhibition of hair cells occurs by activation of calcium-dependent potassium channels. A near-membrane postsynaptic cistern has been proposed to serve as a store from which calcium is released to supplement influx through the ionotropic ACh receptor. However, the time and voltage dependence of acetylcholine (ACh)-evoked potassium currents reveal a more complex relationship between calcium entry and release from stores. The present work uses voltage steps to regulate calcium influx during the application of ACh to hair cells in the chicken basilar papilla. When calcium influx was terminated at positive membrane potential, the ACh-evoked potassium current decayed exponentially over ∼100 ms. However, at negative membrane potentials, this current exhibited a secondary rise in amplitude that could be eliminated by dihydropyridine block of the voltage-gated calcium channels of the hair cell. Calcium entering through voltage-gated channels may transit through the postsynaptic cistern, since ryanodine and sarcoendoplasmic reticulum calcium-ATPase blockers altered the time course and magnitude of this secondary, voltage-dependent contribution to ACh-evoked potassium current. Serial section electron microscopy showed that efferent and afferent synaptic structures are juxtaposed, supporting the possibility that voltage-gated influx at afferent ribbon synapses influences calcium homeostasis during long-lasting cholinergic inhibition. In contrast, spontaneous postsynaptic currents ("minis") resulting from stochastic efferent release of ACh were made briefer by ryanodine, supporting the hypothesis that the synaptic cistern serves primarily as a calcium barrier and sink during low-level synaptic activity. Hypolemmal cisterns such as that at the efferent synapse of the hair cell can play a dynamic role in segregating near-membrane calcium for short-term and long-term signaling. Copyright © 2014 the authors 0270-6474/14/3416688-10$15.00/0.

Synaptic Calcium Regulation in Hair Cells of the Chicken Basilar Papilla

PubMed Central

Im, Gi Jung; Moskowitz, Howard S.; Lehar, Mohammed; Hiel, Hakim

2014-01-01

Cholinergic inhibition of hair cells occurs by activation of calcium-dependent potassium channels. A near-membrane postsynaptic cistern has been proposed to serve as a store from which calcium is released to supplement influx through the ionotropic ACh receptor. However, the time and voltage dependence of acetylcholine (ACh)-evoked potassium currents reveal a more complex relationship between calcium entry and release from stores. The present work uses voltage steps to regulate calcium influx during the application of ACh to hair cells in the chicken basilar papilla. When calcium influx was terminated at positive membrane potential, the ACh-evoked potassium current decayed exponentially over ∼100 ms. However, at negative membrane potentials, this current exhibited a secondary rise in amplitude that could be eliminated by dihydropyridine block of the voltage-gated calcium channels of the hair cell. Calcium entering through voltage-gated channels may transit through the postsynaptic cistern, since ryanodine and sarcoendoplasmic reticulum calcium-ATPase blockers altered the time course and magnitude of this secondary, voltage-dependent contribution to ACh-evoked potassium current. Serial section electron microscopy showed that efferent and afferent synaptic structures are juxtaposed, supporting the possibility that voltage-gated influx at afferent ribbon synapses influences calcium homeostasis during long-lasting cholinergic inhibition. In contrast, spontaneous postsynaptic currents (“minis”) resulting from stochastic efferent release of ACh were made briefer by ryanodine, supporting the hypothesis that the synaptic cistern serves primarily as a calcium barrier and sink during low-level synaptic activity. Hypolemmal cisterns such as that at the efferent synapse of the hair cell can play a dynamic role in segregating near-membrane calcium for short-term and long-term signaling. PMID:25505321

Low-expression of E-cadherin in leukaemia cells causes loss of homophilic adhesion and promotes cell growth.

PubMed

Rao, Qing; Wang, Ji-Ying; Meng, Jihong; Tang, Kejing; Wang, Yanzhong; Wang, Min; Xing, Haiyan; Tian, Zheng; Wang, Jianxiang

2011-09-01

E-cadherin (epithelial cadherin) belongs to the calcium-dependent adhesion molecule superfamily and is implicated in the interactions of haematopoietic progenitors and bone marrow stromal cells. Adhesion capacity to bone marrow stroma was impaired for leukaemia cells, suggesting that a breakdown of adhesive mechanisms governed by an adhesion molecule may exist in leukaemic microenvironment. We previously found that E-cadherin was low expressed in primary acute leukaemia cells compared with normal bone marrow mononuclear cells. In this study, we investigate the functional importance of low E-cadherin expression in leukaemia cell behaviours and investigate its effects in the abnormal interaction of leukaemic cells with stromal cells. After expression of E-cadherin was restored by a demethylating agent in leukaemia cells, E-cadherin-specific adhesion was enhanced. Additionally, siRNA (small interfering RNA)-mediated silencing of E-cadherin in Raji cells resulted in a reduction of cell homophilic adhesion and enhancement of cell proliferation and colony formation. These results suggest that low expression of E-cadherin contributes to the vigorous growth and transforming ability of leukaemic cells.

Cellular level robotic surgery: Nanodissection of intermediate filaments in live keratinocytes.

PubMed

Yang, Ruiguo; Song, Bo; Sun, Zhiyong; Lai, King Wai Chiu; Fung, Carmen Kar Man; Patterson, Kevin C; Seiffert-Sinha, Kristina; Sinha, Animesh A; Xi, Ning

2015-01-01

We present the nanosurgery on the cytoskeleton of live cells using AFM based nanorobotics to achieve adhesiolysis and mimic the effect of pathophysiological modulation of intercellular adhesion. Nanosurgery successfully severs the intermediate filament bundles and disrupts cell-cell adhesion similar to the desmosomal protein disassembly in autoimmune disease, or the cationic modulation of desmosome formation. Our nanomechanical analysis revealed that adhesion loss results in a decrease in cellular stiffness in both cases of biochemical modulation of the desmosome junctions and mechanical disruption of intercellular adhesion, supporting the notion that intercellular adhesion through intermediate filaments anchors the cell structure as focal adhesion does and that intermediate filaments are integral components in cell mechanical integrity. The surgical process could potentially help reveal the mechanism of autoimmune pathology-induced cell-cell adhesion loss as well as its related pathways that lead to cell apoptosis. Copyright © 2015 Elsevier Inc. All rights reserved.

Endothelial NOS is required for SDF-1alpha/CXCR4-mediated peripheral endothelial adhesion of c-kit+ bone marrow stem cells.

PubMed

Kaminski, Alexander; Ma, Nan; Donndorf, Peter; Lindenblatt, Nicole; Feldmeier, Gregor; Ong, Lee-Lee; Furlani, Dario; Skrabal, Christian A; Liebold, Andreas; Vollmar, Brigitte; Steinhoff, Gustav

2008-01-01

In the era of intravascular approaches for regenerative cell therapy, the underlying mechanisms of stem cell migration to non-marrow tissue have not been clarified. We hypothesized that next to a local inflammatory response implying adhesion molecule expression, endothelial nitric oxide synthase (eNOS)-dependent signaling is required for stromal- cell-derived factor-1 alpha (SDF-1alpha)-induced adhesion of c-kit+ cells to the vascular endothelium. SDF-1alpha/tumor necrosis factor-alpha (TNF-alpha)-induced c-kit+-cell shape change and migration capacity was studied in vitro using immunohistochemistry and Boyden chamber assays. In vivo interaction of c-kit+ cells from bone marrow with the endothelium in response to SDF-1alpha/TNF-alpha stimulation was visualized in the cremaster muscle microcirculation of wild-type (WT) and eNOS (-/-) mice using intravital fluorescence microscopy. In addition, NOS activity was inhibited with N-nitro-L-arginine-methylester-hydrochloride in WT mice. To reveal c-kit+-specific adhesion behavior, endogenous leukocytes (EL) and c-kit+ cells from peripheral blood served as control. Moreover, intercellular adhesion molecule-1 (ICAM-1) and CXCR4 were blocked systemically to determine their role in inflammation-related c-kit+-cell adhesion. In vitro, SDF-1alpha enhanced c-kit+-cell migration. In vivo, SDF-1alpha alone triggered endothelial rolling-not firm adherence-of c-kit+ cells in WT mice. While TNF-alpha alone had little effect on adhesion of c-kit+ cells, it induced maximum endothelial EL adherence. However, after combined treatment with SDF-1alpha+TNF-alpha, endothelial adhesion of c-kit+ cells increased independent of their origin, while EL adhesion was not further incremented. Systemic treatment with anti-ICAM-1 and anti-CXCR4-monoclonal antibody completely abolished endothelial c-kit+-cell adhesion. In N-nitro-L-arginine-methylester-hydrochloride-treated WT mice as well as in eNOS (-/-) mice, firm endothelial adhesion of c-kit+ cells was entirely abrogated, while EL adhesion was significantly increased. The chemokine SDF-1alpha mediates firm adhesion c-kit+ cells only in the presence of TNF-alpha stimulation via an ICAM-1- and CXCR4-dependent mechanism. The presence of eNOS appears to be a crucial and specific factor for firm c-kit+-cell adhesion to the vascular endothelium.

Development of Ca2+ hotspots between Lymnaea neurons during synaptogenesis

PubMed Central

Feng, Zhong-Ping; Grigoriev, Nikita; Munno, David; Lukowiak, Ken; MacVicar, Brian A; Goldberg, Jeffrey I; Syed, Naweed I

2002-01-01

Calcium (Ca2+) channel clustering at specific presynaptic sites is a hallmark of mature synapses. However, the spatial distribution patterns of Ca2+ channels at newly formed synapses have not yet been demonstrated. Similarly, it is unclear whether Ca2+ ‘hotspots’ often observed at the presynaptic sites are indeed target cell contact specific and represent a specialized mechanism by which Ca2+ channels are targeted to select synaptic sites. Utilizing both soma–soma paired (synapsed) and single neurons from the mollusk Lymnaea, we have tested the hypothesis that differential gradients of voltage-dependent Ca2+ signals develop in presynaptic neuron at its contact point with the postsynaptic neuron; and that these Ca2+ hotspots are target cell contact specific. Fura-2 imaging, or two-photon laser scanning microscopy of Calcium Green, was coupled with electrophysiological techniques to demonstrate that voltage-induced Ca2+ gradients (hotspots) develop in the presynaptic cell at its contact point with the postsynaptic neuron, but not in unpaired single cells. The incidence of Ca2+ hotspots coincided with the appearance of synaptic transmission between the paired cells, and these gradients were target cell contact specific. In contrast, the voltage-induced Ca2+ signal in unpaired neurons was uniformly distributed throughout the somata; a similar pattern of Ca2+ gradient was observed in the presynaptic neuron when it was soma–soma paired with a non-synaptic partner cell. Moreover, voltage clamp recording techniques, in conjunction with a fast, optical differential perfusion system, were used to demonstrate that the total whole-cell Ca2+ (or Ba2+) current density in single and paired cells was not significantly different. However, the amplitude of Ba2+ current was significantly higher in the presynaptic cell at its contact side with the postsynaptic neurons, compared with non-contacted regions. In summary, this study demonstrates that voltage-induced Ca2+ hotspots develop in the presynaptic cell, concomitant with the appearance of synaptic transmission between the soma–soma paired cells. The appearance of Ca2+ gradients in presynaptic neurons is target cell contact specific and is probably due to a spatial redistribution of existing channels during synaptogenesis. PMID:11850501

The Molecular Architecture of Cell Adhesion: Dynamic Remodeling Revealed by Videonanoscopy.

PubMed

Sergé, Arnauld

2016-01-01

The plasma membrane delimits the cell, which is the basic unit of living organisms, and is also a privileged site for cell communication with the environment. Cell adhesion can occur through cell-cell and cell-matrix contacts. Adhesion proteins such as integrins and cadherins also constitute receptors for inside-out and outside-in signaling within proteolipidic platforms. Adhesion molecule targeting and stabilization relies on specific features such as preferential segregation by the sub-membrane cytoskeleton meshwork and within membrane proteolipidic microdomains. This review presents an overview of the recent insights brought by the latest developments in microscopy, to unravel the molecular remodeling occurring at cell contacts. The dynamic aspect of cell adhesion was recently highlighted by super-resolution videomicroscopy, also named videonanoscopy. By circumventing the diffraction limit of light, nanoscopy has allowed the monitoring of molecular localization and behavior at the single-molecule level, on fixed and living cells. Accessing molecular-resolution details such as quantitatively monitoring components entering and leaving cell contacts by lateral diffusion and reversible association has revealed an unexpected plasticity. Adhesion structures can be highly specialized, such as focal adhesion in motile cells, as well as immune and neuronal synapses. Spatiotemporal reorganization of adhesion molecules, receptors, and adaptors directly relates to structure/function modulation. Assembly of these supramolecular complexes is continuously balanced by dynamic events, remodeling adhesions on various timescales, notably by molecular conformation switches, lateral diffusion within the membrane and endo/exocytosis. Pathological alterations in cell adhesion are involved in cancer evolution, through cancer stem cell interaction with stromal niches, growth, extravasation, and metastasis.

GLUTAMATE NEUROTOXICITY IN THE DEVELOPING RAT COCHLEA IS ANTAGONIZED BY KUNURENIC ACID AND MK-801

EPA Science Inventory

Glutamate (GLU) is neurotoxic in the neonatal rat cochlea, producing hearing impairment which is largely due to the death of spiral ganglion cells, whereas the receptor hair cells are spared. endritic fibers of the spiral ganglion are post-synaptic to the primary afferent synapse...

Adhesion mechanisms in embryogenesis and in cancer invasion and metastasis.

PubMed

Thiery, J P; Boyer, B; Tucker, G; Gavrilovic, J; Valles, A M

1988-01-01

Cell-substratum and cell-cell adhesion mechanisms contribute to the development of animal form. The adhesive status of embryonic cells has been analysed during epithelial-mesenchymal cell interconversion and in cell migrations. Clear-cut examples of the modulation of cell adhesion molecules (CAMs) have been described at critical periods of morphogenesis. In chick embryos the three primary CAMs (N-CAM. L-CAM and N-cadherin) present early in embryogenesis are expressed later in a defined pattern during morphogenesis and histogenesis. The axial mesoderm derived from gastrulating cells expresses increasing amounts of N-cadherin and N-CAM. During metamerization these two adhesion molecules become abundant at somitic cell surfaces. Both CAMs are functional in an in vitro aggregation assay; however, the calcium-dependent adhesion molecule N-cadherin is more sensitive to perturbation by specific antibodies. Neural crest cells which separate from the neural epithelium lose their primary CAMs in a defined time-sequence. Adhesion to fibronectins via specific surface receptors becomes a predominant interaction during the migratory process, while some primary and secondary CAMs are expressed de novo during the ontogeny of the peripheral nervous system. In vitro, different fibronectin functional domains have been identified in the attachment, spreading and migration of neural crest cells. The fibronectin receptors which transduce the adhesive signals play a key role in the control of cell movement. All these results have prompted us to examine whether similar mechanisms operate in carcinoma cell invasion and metastasis. In vitro, rat bladder transitional carcinoma cells convert reversibly into invasive mesenchymal cells. A rapid modulation of adhesive properties is found during the epithelial-mesenchymal carcinoma cell interconversion. The different model systems analysed demonstrate that a limited repertoire of adhesion molecules, expressed in a well-defined spatiotemporal pattern, is involved in tissue formation and in key processes of tumour spread.

In vitro adhesion of fibroblastic cells to titanium alloy discs treated with sodium hydroxide.

PubMed

Al Mustafa, Maisa; Agis, Hermann; Müller, Heinz-Dieter; Watzek, Georg; Gruber, Reinhard

2015-01-01

Adhesion of osteogenic cells on titanium surfaces is a prerequisite for osseointegration. Alkali treatment can increase the hydrophilicity of titanium implant surfaces, thereby supporting the adhesion of blood components. However, it is unclear if alkali treatment also supports the adhesion of cells with a fibroblastic morphology to titanium. Here, we have used a titanium alloy (Ti-6AL-4V) processed by alkali treatment to demonstrate the impact of hydrophilicity on the adhesion of primary human gingival fibroblast and bone cells. Also included were the osteosarcoma and fibroblastoma cell lines, MG63 and L929, respectively. Cell adhesion was determined by scanning electron microscopy. We also measured viability, proliferation, and protein synthesis of the adherent cells. Alkali treatment increased the adhesion of gingival fibroblasts, bone cells, and the two cell lines when seeded onto the titanium alloy surface for 1 h. At 3 h, no significant changes in cell adhesion were observed. Cells grown for 1 day on the titanium alloy surfaces processed by alkali treatment behave similarly to untreated controls with regard to viability, proliferation, and protein synthesis. Based on these preliminary In vitro findings, we conclude that alkali treatment can support the early adhesion of cells with fibroblastic characteristics to a titanium alloy surface. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Cell Adhesion-dependent Serine 85 Phosphorylation of Paxillin Modulates Focal Adhesion Formation and Haptotactic Migration via Association with the C-terminal Tail Domain of Talin*

PubMed Central

Kwak, Tae Kyoung; Lee, Mi-Sook; Ryu, Jihye; Choi, Yoon-Ju; Kang, Minkyung; Jeong, Doyoung; Lee, Jung Weon

2012-01-01

Integrin-mediated adhesion to extracellular matrix proteins is dynamically regulated during morphological changes and cell migration. Upon cell adhesion, protein-protein interactions among molecules at focal adhesions (FAs) play major roles in the regulation of cell morphogenesis and migration. Although tyrosine phosphorylation of paxillin is critically involved in adhesion-mediated signaling, the significance of paxillin phosphorylation at Ser-85 and the mechanism by which it regulates cell migration remain unclear. In this study, we examined how Ser-85 phosphorylation of paxillin affects FA formation and cell migration. We found that paxillin phosphorylation at Ser-85 occurred during HeLa cell adhesion to collagen I and was concomitant with tyrosine phosphorylation of both focal adhesion kinase and talin. However, the non-phosphorylatable S85A mutant of paxillin impaired cell spreading, FA turnover, and migration toward collagen I but not toward serum. Furthermore, whereas the (presumably indirect) interaction between paxillin and the C-terminal tail of talin led to dynamic FAs at the cell boundary, S85A paxillin did not bind talin and caused stabilized FAs in the central region of cells. Together, these observations suggest that cell adhesion-dependent Ser-85 phosphorylation of paxillin is important for its interaction with talin and regulation of dynamic FAs and cell migration. PMID:22761432

A mucus adhesion promoting protein, MapA, mediates the adhesion of Lactobacillus reuteri to Caco-2 human intestinal epithelial cells.

PubMed

Miyoshi, Yukihiro; Okada, Sanae; Uchimura, Tai; Satoh, Eiichi

2006-07-01

Lactobacillus reuteri is one of the dominant lactobacilli found in the gastrointestinal tract of various animals. A surface protein of L. reuteri 104R, mucus adhesion promoting protein (MapA), is considered to be an adhesion factor of this strain. We investigated the relation between MapA and adhesion of L. reuteri to human intestinal (Caco-2) cells. Quantitative analysis of the adhesion of L. reuteri strains to Caco-2 cells showed that various L. reuteri strains bind not only to mucus but also to intestinal epithelial cells. In addition, purified MapA bound to Caco-2 cells, and this binding inhibited the adhesion of L. reuteri in a concentration-dependent manner. Based on these observations, the adhesion of L. reuteri appears due to the binding of MapA to receptor-like molecules on Caco-2 cells. Further, far-western analysis indicated the existence of multiple receptor-like molecules in Caco-2 cells.

Synergistic effect of two cell recognition systems: glycosphingolipid-glycosphingolipid interaction and integrin receptor interaction with pericellular matrix protein.

PubMed

Kojima, N; Hakomori, S

1991-12-01

GM3-expressing cells adhere, spread and migrate on plastic plates coated with Gg3, LacCer and Gb4, but not with other glycosphingolipids (GSLs). Thus, cell adhesion, spreading and migration through GSL-GSL interaction occur in an analogous fashion to the interaction of cells with adhesive matrix proteins [AP, e.g. fibronectin (FN), laminin (LN)] through their integrin receptors. In this study, the adhesion of two GM3-expressing cell lines (B16 melanoma and HEL299 fibroblast) on plastic plates co-coated with GSL plus AP is compared with adhesion on plates coated with GSL (Gg3 or LacCer) alone, or coated with AP alone. Results show that: (i) cell adhesion on GSL-coated plates takes place earlier in the incubation period than that on AP-coated plates; (ii) cell adhesion, as well as spreading, was greatly enhanced (in terms of strength and rapidity) on plates co-coated with GSL plus AP; (iii) repulsion (negative adhesion) of cells was observed on plates co-coated with AP plus N-acetyl-GM3 (NAcGM3) and was presumably based on repulsive NAcGM3-NAcGM3 interaction; (iv) GM3-dependent cell adhesion on GSL-coated plates, as well as synergistic promotion of cell adhesion (based on the GSL-GSL and AP-integrin systems), was suppressed by incubation of cells with anti-GM3 monoclonal antibody DH2 or sialidase. Synergistic adhesion of cells on GSL/AP co-coated plates was less inhibited by incubation with peptide sequences RGDS or YIGSR than was adhesion on plates coated with AP alone.(ABSTRACT TRUNCATED AT 250 WORDS)

In vivo studies of sickle red blood cells.

PubMed

Kaul, Dhananjay K; Fabry, Mary E

2004-03-01

The defining clinical feature of sickle cell anemia is periodic occurrence of painful vasoocclusive crisis. Factors that promote trapping and sickling of red cells in the microcirculation are likely to trigger vasoocclusion. The marked red cell heterogeneity in sickle blood and abnormal adhesion of sickle red cells to vascular endothelium would be major disruptive influences. Using ex vivo and in vivo models, the authors show how to dissect the relative contribution of heterogeneous sickle red cell classes to adhesive and obstructive events. These studies revealed that (1) both rheological abnormalities and adhesion of sickle red cells contribute to their abnormal hemodynamic behavior, (2) venules are the sites of sickle cell adhesion, and (3) sickle red cell deformability plays an important role in adhesive and obstructive events. Preferential adhesion of deformable sickle red cells in postcapillary venules followed by selective trapping of dense sickle red cells could result in vasoocclusion. An updated version of this 2-step model is presented. The multifactorial nature of sickle red cell adhesion needs to be considered in designing antiadhesive therapy in vivo.

Impaired Integrin-mediated Adhesion and Signaling in Fibroblasts Expressing a Dominant-negative Mutant PTP1B

PubMed Central

Arregui, Carlos O.; Balsamo, Janne; Lilien, Jack

1998-01-01

To investigate the role of nonreceptor protein tyrosine phosphatase 1B (PTP1B) in β1-integrin– mediated adhesion and signaling, we transfected mouse L cells with normal and catalytically inactive forms of the phosphatase. Parental cells and cells expressing the wild-type or mutant PTP1B were assayed for (a) adhesion, (b) spreading, (c) presence of focal adhesions and stress fibers, and (d) tyrosine phosphorylation. Parental cells and cells expressing wild-type PTP1B show similar morphology, are able to attach and spread on fibronectin, and form focal adhesions and stress fibers. In contrast, cells expressing the inactive PTP1B have a spindle-shaped morphology, reduced adhesion and spreading on fibronectin, and almost a complete absence of focal adhesions and stress fibers. Attachment to fibronectin induces tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin in parental cells and cells transfected with the wild-type PTP1B, while in cells transfected with the mutant PTP1B, such induction is not observed. Additionally, in cells expressing the mutant PTP1B, tyrosine phosphorylation of Src is enhanced and activity is reduced. Lysophosphatidic acid temporarily reverses the effects of the mutant PTP1B, suggesting the existence of a signaling pathway triggering focal adhesion assembly that bypasses the need for active PTP1B. PTP1B coimmunoprecipitates with β1-integrin from nonionic detergent extracts and colocalizes with vinculin and the ends of actin stress fibers in focal adhesions. Our data suggest that PTP1B is a critical regulatory component of integrin signaling pathways, which is essential for adhesion, spreading, and formation of focal adhesions. PMID:9813103

Flagellin based biomimetic coatings: From cell-repellent surfaces to highly adhesive coatings.

PubMed

Kovacs, Boglarka; Patko, Daniel; Szekacs, Inna; Orgovan, Norbert; Kurunczi, Sandor; Sulyok, Attila; Khanh, Nguyen Quoc; Toth, Balazs; Vonderviszt, Ferenc; Horvath, Robert

2016-09-15

Biomimetic coatings with cell-adhesion-regulating functionalities are intensively researched today. For example, cell-based biosensing for drug development, biomedical implants, and tissue engineering require that the surface adhesion of living cells is well controlled. Recently, we have shown that the bacterial flagellar protein, flagellin, adsorbs through its terminal segments to hydrophobic surfaces, forming an oriented monolayer and exposing its variable D3 domain to the solution. Here, we hypothesized that this nanostructured layer is highly cell-repellent since it mimics the surface of the flagellar filaments. Moreover, we proposed flagellin as a carrier molecule to display the cell-adhesive RGD (Arg-Gly-Asp) peptide sequence and induce cell adhesion on the coated surface. The D3 domain of flagellin was replaced with one or more RGD motifs linked by various oligopeptides modulating flexibility and accessibility of the inserted segment. The obtained flagellin variants were applied to create surface coatings inducing cell adhesion and spreading to different levels, while wild-type flagellin was shown to form a surface layer with strong anti-adhesive properties. As reference surfaces synthetic polymers were applied which have anti-adhesive (PLL-g-PEG poly(l-lysine)-graft-poly(ethylene glycol)) or adhesion inducing properties (RGD-functionalized PLL-g-PEG). Quantitative adhesion data was obtained by employing optical biochips and microscopy. Cell-adhesion-regulating coatings can be simply formed on hydrophobic surfaces by using the developed flagellin-based constructs. The developed novel RGD-displaying flagellin variants can be easily obtained by bacterial production and can serve as alternatives to create cell-adhesion-regulating biomimetic coatings. In the present work, we show for the first time that. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Regulation of T-lymphocyte motility, adhesion and de-adhesion by a cell surface mechanism directed by low density lipoprotein receptor-related protein 1 and endogenous thrombospondin-1.

PubMed

Talme, Toomas; Bergdahl, Eva; Sundqvist, Karl-Gösta

2014-06-01

T lymphocytes are highly motile and constantly reposition themselves between a free-floating vascular state, transient adhesion and migration in tissues. The regulation behind this unique dynamic behaviour remains unclear. Here we show that T cells have a cell surface mechanism for integrated regulation of motility and adhesion and that integrin ligands and CXCL12/SDF-1 influence motility and adhesion through this mechanism. Targeting cell surface-expressed low-density lipoprotein receptor-related protein 1 (LRP1) with an antibody, or blocking transport of LRP1 to the cell surface, perturbed the cell surface distribution of endogenous thrombospondin-1 (TSP-1) while inhibiting motility and potentiating cytoplasmic spreading on intercellular adhesion molecule 1 (ICAM-1) and fibronectin. Integrin ligands and CXCL12 stimulated motility and enhanced cell surface expression of LRP1, intact TSP-1 and a 130,000 MW TSP-1 fragment while preventing formation of a de-adhesion-coupled 110 000 MW TSP-1 fragment. The appearance of the 130 000 MW TSP-1 fragment was inhibited by the antibody that targeted LRP1 expression, inhibited motility and enhanced spreading. The TSP-1 binding site in the LRP1-associated protein, calreticulin, stimulated adhesion to ICAM-1 through intact TSP-1 and CD47. Shear flow enhanced cell surface expression of intact TSP-1. Hence, chemokines and integrin ligands up-regulate a dominant motogenic pathway through LRP1 and TSP-1 cleavage and activate an associated adhesion pathway through the LRP1-calreticulin complex, intact TSP-1 and CD47. This regulation of T-cell motility and adhesion makes pro-adhesive stimuli favour motile responses, which may explain why T cells prioritize movement before permanent adhesion.

Redistribution of Adhesive Forces through Src/FAK Drives Contact Inhibition of Locomotion in Neural Crest.

PubMed

Roycroft, Alice; Szabó, András; Bahm, Isabel; Daly, Liam; Charras, Guillaume; Parsons, Maddy; Mayor, Roberto

2018-06-04

Contact inhibition of locomotion is defined as the behavior of cells to cease migrating in their former direction after colliding with another cell. It has been implicated in multiple developmental processes and its absence has been linked to cancer invasion. Cellular forces are thought to govern this process; however, the exact role of traction through cell-matrix adhesions and tension through cell-cell adhesions during contact inhibition of locomotion remains unknown. Here we use neural crest cells to address this and show that cell-matrix adhesions are rapidly disassembled at the contact between two cells upon collision. This disassembly is dependent upon the formation of N-cadherin-based cell-cell adhesions and driven by Src and FAK activity. We demonstrate that the loss of cell-matrix adhesions near the contact leads to a buildup of tension across the cell-cell contact, a step that is essential to drive cell-cell separation after collision. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Tuning cell adhesive properties via layer-by-layer assembly of chitosan and alginate

PubMed Central

Silva, Joana M.; García, José R.; Reis, Rui L.; García, Andrés J.; Mano, João F.

2017-01-01

Understanding the mechanisms controlling cell-multilayer film interactions is crucial to the successful engineering of these coatings for biotechnological and biomedical applications. Herein, we present a strategy to tune the cell adhesive properties of multilayers based on marine polysaccharides with and without cross-linking and/or coating with extracellular matrix proteins. Chemical cross-linking of multilayers improved mechanical properties of the coatings but also elicited changes in surface chemistry that alter the adhesion of human umbilical vein endothelial cells. We evaluated a strategy to decouple the mechanical and chemical properties of these films, enabling the transition from cell-adhesive to cell-resistant multilayers. Addition of chitosan/alginate multilayers on top of cross-linked films decreased endothelial cell adhesion, spreading, and proliferation to similar levels as uncross-linked films. Our findings highlight the key role of surface chemistry in cell-multilayer film interactions, and these engineered nanocoatings represent a tunable model of cell adhesive and non-adhesive multilayered films. PMID:28126597

Molecular analysis of antigen-independent adhesion forces between T and B lymphocytes.

PubMed Central

Amblard, F; Auffray, C; Sekaly, R; Fischer, A

1994-01-01

The low-affinity interactions underlying antigen recognition by T-cell receptors (TCRs) are thought to involve antigen-independent adhesion mechanisms. Using a hydrodynamic approach, we found that antigen-independent adhesion occurred between human B cells and resting T cells in a transient and temperature-dependent fashion. The mean cell-cell adhesion force was 0.32 x 10(-9) N and was generated by similar contributions (0.16 x 10(-9) N) of the LFA-1- and CD2-dependent adhesion pathways. After T-cell stimulation with a phorbol ester, the force contributed by LFA-1 was drastically increased, while that of CD2 was unaffected. We propose that weak receptor-mediated adhesion initiates antigen-independent intercellular contacts required for antigen recognition by the TCR and is upregulated following TCR engagement. The method used permits adhesion forces between living cells to be resolved at the molecular level and should prove valuable for the rapid assessment of interaction forces between various types of cells and cell-sized particles. Images PMID:7909604

Cell-cell adhesion in the cnidaria: insights into the evolution of tissue morphogenesis.

PubMed

Magie, Craig R; Martindale, Mark Q

2008-06-01

Cell adhesion is a major aspect of cell biology and one of the fundamental processes involved in the development of a multicellular animal. Adhesive mechanisms, both cell-cell and between cell and extracellular matrix, are intimately involved in assembling cells into the three-dimensional structures of tissues and organs. The modulation of adhesive complexes could therefore be seen as a central component in the molecular control of morphogenesis, translating information encoded within the genome into organismal form. The availability of whole genomes from early-branching metazoa such as cnidarians is providing important insights into the evolution of adhesive processes by allowing for the easy identification of the genes involved in adhesion in these organisms. Discovery of the molecular nature of cell adhesion in the early-branching groups, coupled with comparisons across the metazoa, is revealing the ways evolution has tinkered with this vital cellular process in the generation of the myriad forms seen across the animal kingdom.

Identification of a Monocyte Receptor on Herpesvirus-Infected Endothelial Cells

NASA Astrophysics Data System (ADS)

Etingin, Orli R.; Silverstein, Roy L.; Hajjar, David P.

1991-08-01

The adhesion of circulating blood cells to vascular endothelium may be an initial step in atherosclerosis, inflammation, and wound healing. One mechanism for promoting cell-cell adhesion involves the expression of adhesion molecules on the surface of the target cell. Herpes simplex virus infection of endothelium induces arterial injury and has been implicated in the development of human atherosclerosis. We now demonstrate that HSV-infected endothelial cells express the adhesion molecule GMP140 and that this requires cell surface expression of HSV glycoprotein C and local thrombin generation. Monocyte adhesion to HSV-infected endothelial cells was completely inhibited by anti-GMP140 antibodies but not by antibodies to other adhesion molecules such as VCAM and ELAM-1. The induction of GMP140 expression on HSV-infected endothelium may be an important pathophysiological mechanism in virus-induced cell injury and inflammation.

Remodeling of the postsynaptic plasma membrane during neural development.

PubMed

Tulodziecka, Karolina; Diaz-Rohrer, Barbara B; Farley, Madeline M; Chan, Robin B; Di Paolo, Gilbert; Levental, Kandice R; Waxham, M Neal; Levental, Ilya

2016-11-07

Neuronal synapses are the fundamental units of neural signal transduction and must maintain exquisite signal fidelity while also accommodating the plasticity that underlies learning and development. To achieve these goals, the molecular composition and spatial organization of synaptic terminals must be tightly regulated; however, little is known about the regulation of lipid composition and organization in synaptic membranes. Here we quantify the comprehensive lipidome of rat synaptic membranes during postnatal development and observe dramatic developmental lipidomic remodeling during the first 60 postnatal days, including progressive accumulation of cholesterol, plasmalogens, and sphingolipids. Further analysis of membranes associated with isolated postsynaptic densities (PSDs) suggests the PSD-associated postsynaptic plasma membrane (PSD-PM) as one specific location of synaptic remodeling. We analyze the biophysical consequences of developmental remodeling in reconstituted synaptic membranes and observe remarkably stable microdomains, with the stability of domains increasing with developmental age. We rationalize the developmental accumulation of microdomain-forming lipids in synapses by proposing a mechanism by which palmitoylation of the immobilized scaffold protein PSD-95 nucleates domains at the postsynaptic plasma membrane. These results reveal developmental changes in lipid composition and palmitoylation that facilitate the formation of postsynaptic membrane microdomains, which may serve key roles in the function of the neuronal synapse. © 2016 Tulodziecka et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Human immunodeficiency virus-1 protein Tat induces excitotoxic loss of presynaptic terminals in hippocampal cultures.

PubMed

Shin, Angela H; Thayer, Stanley A

2013-05-01

Human immunodeficiency virus (HIV) infection of the CNS produces dendritic damage that correlates with cognitive decline in patients with HIV-associated neurocognitive disorders (HAND). HIV-induced neurotoxicity results in part from viral proteins shed from infected cells, including the HIV transactivator of transcription (Tat). We previously showed that Tat binds to the low density lipoprotein receptor-related protein (LRP), resulting in overactivation of NMDA receptors, activation of the ubiquitin-proteasome pathway, and subsequent loss of postsynaptic densities. Here, we show that Tat also induces a loss of presynaptic terminals. The number of presynaptic terminals was quantified using confocal imaging of synaptophysin fused to green fluorescent protein (Syn-GFP). Tat-induced loss of presynaptic terminals was secondary to excitatory postsynaptic mechanisms because treatment with an LRP antagonist or an NMDA receptor antagonist inhibited this loss. Treatment with nutlin-3, an E3 ligase inhibitor, prevented Tat-induced loss of presynaptic terminals. These data suggest that Tat-induced loss of presynaptic terminals is a consequence of excitotoxic postsynaptic activity. We previously found that ifenprodil, an NR2B subunit-selective NMDA receptor antagonist, induced recovery of postsynaptic densities. Here we show that Tat-induced loss of presynaptic terminals was reversed by ifenprodil treatment. Thus, Tat-induced loss of presynaptic terminals is reversible, and this recovery can be initiated by inhibiting a subset of postsynaptic NMDA receptors. Understanding the dynamics of synaptic changes in response to HIV infection of the CNS may lead to the design of improved pharmacotherapies for HAND patients. Copyright © 2012 Elsevier Inc. All rights reserved.

Cadherin-23 Mediates Heterotypic Cell-Cell Adhesion between Breast Cancer Epithelial Cells and Fibroblasts

PubMed Central

Apostolopoulou, Maria; Ligon, Lee

2012-01-01

In the early stages of breast cancer metastasis, epithelial cells penetrate the basement membrane and invade the surrounding stroma, where they encounter fibroblasts. Paracrine signaling between fibroblasts and epithelial tumor cells contributes to the metastatic cascade, but little is known about the role of adhesive contacts between these two cell types in metastasis. Here we show that MCF-7 breast cancer epithelial cells and normal breast fibroblasts form heterotypic adhesions when grown together in co-culture, as evidenced by adhesion assays. PCR and immunoblotting show that both cell types express multiple members of the cadherin superfamily, including the atypical cadherin, cadherin-23, when grown in isolation and in co-culture. Immunocytochemistry experiments show that cadherin-23 localizes to homotypic adhesions between MCF-7 cells and also to heterotypic adhesions between the epithelial cells and fibroblasts, and antibody inhibition and RNAi experiments show that cadherin-23 plays a role in mediating these adhesive interactions. Finally, we show that cadherin-23 is upregulated in breast cancer tissue samples, and we hypothesize that heterotypic adhesions mediated by this atypical cadherin may play a role in the early stages of metastasis. PMID:22413011

Strong adhesion by regulatory T cells induces dendritic cell cytoskeletal polarization and contact-dependent lethargy.

PubMed

Chen, Jiahuan; Ganguly, Anutosh; Mucsi, Ashley D; Meng, Junchen; Yan, Jiacong; Detampel, Pascal; Munro, Fay; Zhang, Zongde; Wu, Mei; Hari, Aswin; Stenner, Melanie D; Zheng, Wencheng; Kubes, Paul; Xia, Tie; Amrein, Matthias W; Qi, Hai; Shi, Yan

2017-02-01

Dendritic cells are targeted by regulatory T (T reg) cells, in a manner that operates as an indirect mode of T cell suppression. In this study, using a combination of single-cell force spectroscopy and structured illumination microscopy, we analyze individual T reg cell-DC interaction events and show that T reg cells exhibit strong intrinsic adhesiveness to DCs. This increased DC adhesion reduces the ability of contacted DCs to engage other antigen-specific cells. We show that this unusually strong LFA-1-dependent adhesiveness of T reg cells is caused in part by their low calpain activities, which normally release integrin-cytoskeleton linkage, and thereby reduce adhesion. Super resolution imaging reveals that such T reg cell adhesion causes sequestration of Fascin-1, an actin-bundling protein essential for immunological synapse formation, and skews Fascin-1-dependent actin polarization in DCs toward the T reg cell adhesion zone. Although it is reversible upon T reg cell disengagement, this sequestration of essential cytoskeletal components causes a lethargic state of DCs, leading to reduced T cell priming. Our results reveal a dynamic cytoskeletal component underlying T reg cell-mediated DC suppression in a contact-dependent manner. © 2017 Chen et al.

Bidirectional control of spike timing by GABA(A) receptor-mediated inhibition during theta oscillation in CA1 pyramidal neurons.

PubMed

Kwag, Jeehyun; Paulsen, Ole

2009-08-26

Precisely controlled spike times relative to theta-frequency network oscillations play an important role in hippocampal memory processing. Here we study how inhibitory synaptic input during theta oscillation contributes to the control of spike timing. Using whole-cell patch-clamp recordings from CA1 pyramidal cells in vitro with dynamic clamp to simulate theta-frequency oscillation (5 Hz), we show that gamma-aminobutyric acid-A (GABA(A)) receptor-mediated inhibitory postsynaptic potentials (IPSPs) can not only delay but also advance the postsynaptic spike depending on the timing of the inhibition relative to the oscillation. Spike time advancement with IPSP was abolished by the h-channel blocker ZD7288 (10 microM), suggesting that IPSPs can interact with intrinsic membrane conductances to yield bidirectional control of spike timing.

Paired-Pulse Depression at Photoreceptor Synapses

PubMed Central

Rabl, Katalin; Cadetti, Lucia; Thoreson, Wallace B.

2011-01-01

Synaptic depression produced by repetitive stimulation is likely to be particularly important in shaping responses of second-order retinal neurons at the tonically active photoreceptor synapse. We analyzed the time course and mechanisms of synaptic depression at rod and cone synapses using paired-pulse protocols involving two complementary measurements of exocytosis: (1) paired whole-cell recordings of the postsynaptic current (PSC) in second-order retinal neurons and (2) capacitance measurements of vesicular membrane fusion in rods and cones. PSCs in ON bipolar, OFF bipolar, and horizontal cells evoked by stimulation of either rods or cones recovered from paired-pulse depression (PPD) at rates similar to the recovery of exocytotic capacitance changes in rods and cones. Correlation between presynaptic and postsynaptic measures of recovery from PPD suggests that 80 –90% of the depression at these synapses is presynaptic in origin. Consistent with a predominantly presynaptic mechanism, inhibiting desensitization of postsynaptic glutamate receptors had little effect on PPD. The depression of exocytotic capacitance changes exceeded depression of the presynaptic calcium current, suggesting that it is primarily caused by a depletion of synaptic vesicles. In support of this idea, limiting Ca2+ influx by using weaker depolarizing stimuli promoted faster recovery from PPD. Although cones exhibit much faster exocytotic kinetics than rods, exocytotic capacitance changes recovered from PPD at similar rates in both cell types. Thus, depression of release is not likely to contribute to differences in the kinetics of transmission from rods and cones. PMID:16510733

CD147-targeting siRNA inhibits cell-matrix adhesion of human malignant melanoma cells by phosphorylating focal adhesion kinase.

PubMed

Nishibaba, Rie; Higashi, Yuko; Su, Juan; Furukawa, Tatsuhiko; Kawai, Kazuhiro; Kanekura, Takuro

2012-01-01

CD147/basigin, highly expressed on the surface of malignant tumor cells including malignant melanoma (MM) cells, plays a critical role in the invasiveness and metastasis of MM. Metastasis is an orchestrated process comprised of multiple steps including adhesion and invasion. Integrin, a major adhesion molecule, co-localizes with CD147/basigin on the cell surface. Using the human MM cell line A375 that highly expresses CD147/basigin, we investigated whether CD147/basigin is involved in adhesion in association with integrin. CD147/basigin was knocked-down using siRNA targeting CD147 to elucidate the role of CD147/basigin. Cell adhesion was evaluated by adhesion assay on matrix-coated plates. The localization of integrin was inspected under a confocal microscope and the expression and phosphorylation of focal adhesion kinase (FAK), a downstream kinase of integrin, were examined by western blot analysis. Silencing of CD147/basigin in A375 cells by siRNA induced the phosphorylation of FAK at Y397. Integrin identified on the surface of parental cells was distributed in a speckled fashion in the cytoplasm of CD147 knockdown cells, resulting in morphological changes from a round to a polygonal shape with pseudopodial protrusions. Silencing of CD147/basigin in A375 cells clearly weakened their adhesiveness to collagen I and IV. Our results suggest that CD147/basigin regulates the adhesion of MM cells to extracellular matrices and of integrin β1 signaling via the phosphorylation of FAK. © 2011 Japanese Dermatological Association.

Paired-pulse facilitation and depression at unitary synapses in rat hippocampus: quantal fluctuation affects subsequent release.

PubMed Central

Debanne, D; Guérineau, N C; Gähwiler, B H; Thompson, S M

1996-01-01

1. Excitatory synaptic transmission between pairs of monosynaptically coupled pyramidal cells was examined in rat hippocampal slice cultures. Action potentials were elicited in single CA3 pyramidal cells impaled with microelectrodes and unitary excitatory postsynaptic currents (EPSCs) were recorded in whole-cell voltage-clamped CA1 or CA3 cells. 2. The amplitude of successive unitary EPSCs in response to single action potentials varied. The amplitude of EPSCs was altered by adenosine or changes in the [Mg2+]/[CA2+] ratio. We conclude that single action potentials triggered the release of multiple quanta of glutamate. 3. When two action potentials were elicited in the presynaptic cell, the amplitude of the second EPSC was inversely related to the amplitude of the first. Paired-pulse facilitation (PPF) was observed when the first EPSC was small, i.e. the second EPSC was larger than the first, whereas paired-pulse depression (PPD) was observed when the first EPSC was large. 4. The number of trials displaying PPD was greater when release probability was increased, and smaller when release probability was decreased. 5. PPD was not postsynaptically mediated because it was unaffected by decreasing ionic flux with 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or receptor desensitization with aniracetam. 6. PPF was maximal at an interstimulus interval of 70 ms and recovered within 500 ms. Recovery from PPD occurred within 5 s. 7. We propose that multiple release sites are formed by the axon of a CA3 pyramidal cell and a single postsynaptic CA1 or CA3 cell. PPF is observed if the first action potential fails to release transmitter at most release sites. PPD is observed if the first action potential successfully triggers release at most release sites. 8. Our observations of PPF are consistent with the residual calcium hypothesis. We conclude that PPD results from a decrease in quantal content, perhaps due to short-term depletion of readily releasable vesicles. PMID:9011608

Cell adhesion molecules, the extracellular matrix and oral squamous carcinoma.

PubMed

Lyons, A J; Jones, J

2007-08-01

Carcinomas are characterized by invasion of malignant cells into the underlying connective tissue and migration of malignant cells to form metastases at distant sites. These processes require alterations in cell-cell and cell-extracellular matrix interactions. As cell adhesion molecules play a role in cell-cell and cell-extracellular matrix adhesion and interactions they are involved in the process of tumour invasion and metastases. In epithelial tissues, receptors of the integrin family mediate adhesion to the adjacent matrix whereas cadherins largely mediate intercellular adhesion. These and other cell adhesion molecules such as intercellular adhesion molecule-1, CD44, dystroglycans and selectins, are involved and undergo changes in carcinomas, which provide possible targets for anti-cancer drug treatments. In the extracellular matrix that is associated with tumours, laminin 5, oncofetal fibronectin and tenascin C appear. The degree of expression of some of these moieties indicates prognosis in oral cancer and offer targets for antibody-directed radiotherapy. Metalloproteases which degrade the extracellular matrix are increased in carcinomas, and their activity is necessary for tumour angiogenesis and consequent invasion and metastases. Metalloprotease inhibitors have begun to produce decreases in mortality in clinical trials. This report provides a brief overview of our current understanding of cell adhesion molecules, the extracellular matrix, tumour invasion and metastasis.

Application of Organosilane Monolayer Template to Quantitative Evaluation of Cancer Cell Adhesive Ability

NASA Astrophysics Data System (ADS)

Tanii, Takashi; Sasaki, Kosuke; Ichisawa, Kota; Demura, Takanori; Beppu, Yuichi; Vu, Hoan Anh; Thanh Chi, Hoan; Yamamoto, Hideaki; Sato, Yuko

2011-06-01

The adhesive ability of two human pancreatic cancer cell lines was evaluated using organosilane monolayer templates (OMTs). Using the OMT, the spreading area of adhered cells can be limited, and this enables us to focus on the initial attachment process of adhesion. Moreover, it becomes possible to arrange the cells in an array and to quantitatively evaluate the number of attached cells. The adhesive ability of the cancer cells cultured on the OMT was controlled by adding (-)-epigallocatechin-3-gallate (EGCG), which blocks a receptor that mediates cell adhesion and is overexpressed in cancer cells. Measurement of the relative ability of the cancer cells to attach to the OMT revealed that the ability for attachment decreased with increasing EGCG concentration. The results agreed well with the western blot analysis, indicating that the OMT can potentially be employed to evaluate the adhesive ability of various cancer cells.

Modeling keratinocyte wound healing dynamics: Cell-cell adhesion promotes sustained collective migration.

PubMed

Nardini, John T; Chapnick, Douglas A; Liu, Xuedong; Bortz, David M

2016-07-07

The in vitro migration of keratinocyte cell sheets displays behavioral and biochemical similarities to the in vivo wound healing response of keratinocytes in animal model systems. In both cases, ligand-dependent Epidermal Growth Factor Receptor (EGFR) activation is sufficient to elicit collective cell migration into the wound. Previous mathematical modeling studies of in vitro wound healing assays assume that physical connections between cells have a hindering effect on cell migration, but biological literature suggests a more complicated story. By combining mathematical modeling and experimental observations of collectively migrating sheets of keratinocytes, we investigate the role of cell-cell adhesion during in vitro keratinocyte wound healing assays. We develop and compare two nonlinear diffusion models of the wound healing process in which cell-cell adhesion either hinders or promotes migration. Both models can accurately fit the leading edge propagation of cell sheets during wound healing when using a time-dependent rate of cell-cell adhesion strength. The model that assumes a positive role of cell-cell adhesion on migration, however, is robust to changes in the leading edge definition and yields a qualitatively accurate density profile. Using RNAi for the critical adherens junction protein, α-catenin, we demonstrate that cell sheets with wild type cell-cell adhesion expression maintain migration into the wound longer than cell sheets with decreased cell-cell adhesion expression, which fails to exhibit collective migration. Our modeling and experimental data thus suggest that cell-cell adhesion promotes sustained migration as cells pull neighboring cells into the wound during wound healing. Copyright © 2016 Elsevier Ltd. All rights reserved.

Adrenoceptor-Mediated Post- and Pre-Synaptic Regulations of the Reticulospinal Neurons in Rat Caudal Pontine Reticular Nucleus.

PubMed

Yang, Nian; Qiao, Qi-Cheng; Liu, Yu-Hui; Zhang, Ji-Qiang; Hu, Zhi-An; Zhang, Jun

2016-12-01

The central noradrenergic system participates in diverse nervous functions. Nevertheless, our knowledge of the action of adrenoceptors in motor regulation is still lacking. Intriguingly, reticulospinal neurons in the caudal pontine reticular nucleus (PnC) receive fairly dense noradrenergic innervation and play an important role in motor control. Here, after demonstrating the expression of α1- and α2-adrenoceptors in the PnC, we found that noradrenaline elicited a post-synaptic effect (inward or outward whole-cell current at -70 mV holding) on PnC reticulospinal neurons. The α1- and α2-adrenoceptors were co-expressed in individual PnC reticulospinal neurons to mediate an inward and an outward current component at -70 mV holding, respectively, which, when superposed, produced the overall post-synaptic effects of noradrenaline (NA). More importantly, the activation of post-synaptic α1- or α2-adrenoceptors indeed exerted opposing modulations (excitation vs. inhibition) on the firing activities of individual PnC reticulospinal neurons. Furthermore, the activation and inhibition of the Na + -permeable non-selective cationic conductance (NSCC) were demonstrated to be coupled to α1- and α2-adrenoceptors, respectively. Additionally, the activation of α2-adrenoceptors activated K + conductance. Pre-synaptically, the α2-adrenoceptors were expressed to attenuate the miniature excitatory postsynaptic current (mEPSC) in PnC reticulospinal neurons, but not to affect the miniature inhibitory postsynaptic current (mIPSC). Consistently, the evoked EPSC in PnC reticulospinal neurons was suppressed after the activation of pre-synaptic α2-adrenoceptors. Thus, the excitatory input and post-synaptic dynamics of PnC reticulospinal neurons are indeed intricately modulated by the activation of α1- and α2-adrenoceptors, through which motor control may be regulated in an adaptive manner by the central noradrenergic system.

Ketamine attenuates the glutamatergic neurotransmission in the ventral posteromedial nucleus slices of rats.

PubMed

Fu, Bao; Liu, Chengxi; Zhang, Yajun; Fu, Xiaoyun; Zhang, Lin; Yu, Tian

2017-08-23

Ketamine is a frequently used intravenous anesthetic, which can reversibly induce loss of consciousness (LOC). Previous studies have demonstrated that thalamocortical system is critical for information transmission and integration in the brain. The ventral posteromedial nucleus (VPM) is a critical component of thalamocortical system. Glutamate is an important excitatory neurotransmitter in the brain and may be involved in ketamine-induced LOC. The study used whole-cell patch-clamp to observe the effect of ketamine (30 μM-1000 μM) on glutamatergic neurotransmission in VPM slices. Ketamine significantly decreased the amplitude of glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs), but only higher concentration of ketamine (300 μM and 1000 μM) suppressed the frequency of sEPSCs. Ketamine (100 μM-1000 μM) also decreased the amplitude of glutamatergic miniature excitatory postsynaptic currents (mEPSCs), without altering the frequency. In VPM neurons, ketamine attenuates the glutamatergic neurotransmission mainly through postsynaptic mechanism and action potential may be involved in the process.

END-PLATE ACETYLCHOLINE RECEPTOR: STRUCTURE, MECHANISM, PHARMACOLOGY, AND DISEASE

PubMed Central

Sine, Steven M.

2012-01-01

The synapse is a localized neurohumoral contact between a neuron and an effector cell and may be considered the quantum of fast intercellular communication. Analogously, the postsynaptic neurotransmitter receptor may be considered the quantum of fast chemical to electrical transduction. Our understanding of postsynaptic receptors began to develop about a hundred years ago with the demonstration that electrical stimulation of the vagus nerve released acetylcholine and slowed the heart beat. During the past 50 years, advances in understanding postsynaptic receptors increased at a rapid pace, owing largely to studies of the acetylcholine receptor (AChR) at the motor endplate. The endplate AChR belongs to a large superfamily of neurotransmitter receptors, called Cys-loop receptors, and has served as an exemplar receptor for probing fundamental structures and mechanisms that underlie fast synaptic transmission in the central and peripheral nervous systems. Recent studies provide an increasingly detailed picture of the structure of the AChR and the symphony of molecular motions that underpin its remarkably fast and efficient chemoelectrical transduction. PMID:22811427

Subunit-dependent postsynaptic expression of kainate receptors on hippocampal interneurons in area CA1

PubMed Central

Wondolowski, Joyce; Frerking, Matthew

2009-01-01

Kainate receptors (KARs) contribute to postsynaptic excitation in only a select subset of neurons. To define the parameters that specify the postsynaptic expression of KARs, we examined the contribution of KARs to EPSCs on hippocampal interneurons in area CA1. Interneurons in stratum radiatum/lacunosum-moleculare (SR/SLM) express KARs both with and without the GluR5 subunit, but KAR-mediated EPSCs are generated mainly, if not entirely, by GluR5-containing KARs. Extrasynaptic glutamate spillover profoundly recruits AMPARs with little effect on KARs, indicating that KARs are targeted at the synapse more precisely than AMPARs. However, spontaneous EPSCs with a conventional AMPAR component did not have a resolvable contribution of KARs, suggesting that the KARs that contribute to the evoked EPSCs are at a distinct set of synapses. GluR5-containing KARs on interneurons in stratum oriens do not contribute substantially to the EPSC. We conclude that KARs are localized to synapses by cell type-, synapse-, and subunit-selective mechanisms. PMID:19144856

Dendritic position is a major determinant of presynaptic strength

PubMed Central

de Jong, Arthur P.H.; Schmitz, Sabine K.; Toonen, Ruud F.G.

2012-01-01

Different regulatory principles influence synaptic coupling between neurons, including positional principles. In dendrites of pyramidal neurons, postsynaptic sensitivity depends on synapse location, with distal synapses having the highest gain. In this paper, we investigate whether similar rules exist for presynaptic terminals in mixed networks of pyramidal and dentate gyrus (DG) neurons. Unexpectedly, distal synapses had the lowest staining intensities for vesicular proteins vGlut, vGAT, Synaptotagmin, and VAMP and for many nonvesicular proteins, including Bassoon, Munc18, and Syntaxin. Concomitantly, distal synapses displayed less vesicle release upon stimulation. This dependence of presynaptic strength on dendritic position persisted after chronically blocking action potential firing and postsynaptic receptors but was markedly reduced on DG dendrites compared with pyramidal dendrites. These data reveal a novel rule, independent of neuronal activity, which regulates presynaptic strength according to dendritic position, with the strongest terminals closest to the soma. This gradient is opposite to postsynaptic gradients observed in pyramidal dendrites, and different cell types apply this rule to a different extent. PMID:22492722

Retinoids induce integrin-independent lymphocyte adhesion through RAR-α nuclear receptor activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Whelan, Jarrett T.; Wang, Lei; Chen, Jianming

2014-11-28

Highlights: • Transcription and translation are required for retinoid-induced lymphocyte adhesion. • RAR activation is sufficient to induced lymphocyte cell adhesion. • Vitamin D derivatives inhibit RAR-prompted lymphocyte adhesion. • Adhesion occurs through a novel binding site within ADAM disintegrin domains. • RARα is a key nuclear receptor for retinoid-dependent lymphocyte cell adhesion. - Abstract: Oxidative metabolites of vitamin A, in particular all-trans-retinoic acid (atRA), have emerged as key factors in immunity by specifying the localization of immune cells to the gut. Although it is appreciated that isomers of retinoic acid activate the retinoic acid receptor (RAR) and retinoid Xmore » receptor (RXR) family of nuclear receptors to elicit cellular changes, the molecular details of retinoic acid action remain poorly defined in immune processes. Here we employ a battery of agonists and antagonists to delineate the specific nuclear receptors utilized by retinoids to evoke lymphocyte cell adhesion to ADAM (adisintegrin and metalloprotease) protein family members. We report that RAR agonism is sufficient to promote immune cell adhesion in both immortal and primary immune cells. Interestingly, adhesion occurs independent of integrin function, and mutant studies demonstrate that atRA-induced adhesion to ADAM members required a distinct binding interface(s) as compared to integrin recognition. Anti-inflammatory corticosteroids as well as 1,25-(OH){sub 2}D{sub 3}, a vitamin D metabolite that prompts immune cell trafficking to the skin, potently inhibited the observed adhesion. Finally, our data establish that induced adhesion was specifically attributable to the RAR-α receptor isotype. The current study provides novel molecular resolution as to which nuclear receptors transduce retinoid exposure into immune cell adhesion.« less

Molecules mediating adhesion of T and B cells, monocytes and granulocytes to vascular endothelial cells.

PubMed Central

Prieto, J; Beatty, P G; Clark, E A; Patarroyo, M

1988-01-01

Leucocytes interact with vascular endothelial cells (EC), and adhesion between these two cell types in vitro is modulated by phorbol ester. Monocytes were found to display the highest basal adhesion to EC, followed by Epstein-Barr virus-immortalized normal B cells (EBV-B), T cells and granulocytes. Phorbol ester treatment increased the adhesion of all types of leucocytes, except monocytes. In the presence of this compound, monoclonal antibody 60.3 to GP90 (CD18, a leucocyte-adhesion protein which is non-covalently associated to either GP160, GP155, or GP130) was found to inhibit the adhesion of the four types of leucocytes to a considerable extent, while anti-lymphocyte function-associated antigen-1 (LFA-1) antibody to GP160 (CD11a) inhibited the adhesion of T and B cells only. Antibody 60.1 to GP155 (CD11b) had a major inhibitory activity exclusively on granulocytes, while antibody LB-2, which recognizes a distinct adhesion molecule (GP84) and, in contrast to the previous antibodies, reacts with EC, mainly inhibited adhesion of EBV-B and did not increase the inhibition obtained with antibody 60.3 alone. Fab fragments of antibody 60.3 inhibited leucocyte adhesion more efficiently, in either the absence or presence of phorbol ester, than the intact antibody molecule. It is concluded the GP90, either alone or associated to the larger glycoproteins, mediates the adhesion in all types of leucocytes, while GP84 mediates the adhesion of the activated B cells. Images Figure 2 PMID:3259203

Circulating vascular cell adhesion molecule-1 in pre-eclampsia, gestational hypertension, and normal pregnancy: evidence of selective dysregulation of vascular cell adhesion molecule-1 homeostasis in pre-eclampsia.

PubMed

Higgins, J R; Papayianni, A; Brady, H R; Darling, M R; Walshe, J J

1998-08-01

Our purpose was to investigate circulating levels of vascular cell adhesion molecule-1 in the peripheral and uteroplacental circulations during normotensive and hypertensive pregnancies. This prospective observational study involved 2 patient groups. Group 1 consisted of 22 women with pre-eclampsia and 30 normotensive women followed up longitudinally through pregnancy and post partum. There were an additional 13 women with established gestational hypertension. Group 2 consisted of 20 women with established pre-eclampsia and 19 normotensive control subjects undergoing cesarean delivery. Plasma levels of vascular cell adhesion molecule-1 were measured in blood drawn from the antecubital vein (group 1) and from both the antecubital and uterine veins (group 2). Data were analyzed by analysis of variance. In group 1 vascular cell adhesion molecule-1 levels did not change significantly throughout normal pregnancy and post partum. Women with established pre-eclampsia had increased vascular cell adhesion molecule-1 levels compared with the normotensive pregnancy group (P = .01). Vascular cell adhesion molecule-1 levels were not elevated in women with established gestational hypertension. In group 2 significantly higher levels of vascular cell adhesion molecule-1 were detected in the uteroplacental (P < .0001) and peripheral (P < .0001) circulations of pre-eclamptic women by comparison with normotensive women. In the pre-eclamptic group there was a tendency toward higher vascular cell adhesion molecule-1 levels in the peripheral circulation than in the uteroplacental circulation (P = .06). In contrast to vascular cell adhesion molecule-1, circulating levels of E-selectin and intercellular adhesion molecule-1, other major leukocyte adhesion molecules expressed by the endothelium, were not different in pre-eclamptic and normotensive pregnancies. Established pre-eclampsia is characterized by selective dysregulation of vascular cell adhesion molecule-1 homeostasis. This event is not an early preclinical feature of pre-eclampsia, does not persist post partum, is not a feature of nonproteinuric gestational hypertension, and is not observed with other major leukocyte adhesion molecules. Induction of vascular cell adhesion molecule-1 expression in pre-eclampsia may contribute to leukocyte-mediated tissue injury in this condition or may reflect perturbation of other, previously unrecognized, functions of this molecule in pregnancy.

The CD44-initiated pathway of T-cell extravasation uses VLA-4 but not LFA-1 for firm adhesion

PubMed Central

Siegelman, Mark H.; Stanescu, Diana; Estess, Pila

2000-01-01

Leukocytes extravasate from the blood in response to physiologic or pathologic demands by means of complementary ligand interactions between leukocytes and endothelial cells. The multistep model of leukocyte extravasation involves an initial transient interaction (“rolling” adhesion), followed by secondary (firm) adhesion. We recently showed that binding of CD44 on activated T lymphocytes to endothelial hyaluronan (HA) mediates a primary adhesive interaction under shear stress, permitting extravasation at sites of inflammation. The mechanism for subsequent firm adhesion has not been elucidated. Here we demonstrate that the integrin VLA-4 is used in secondary adhesion after CD44-mediated primary adhesion of human and mouse T cells in vitro, and by mouse T cells in an in vivo model. We show that clonal cell lines and polyclonally activated normal T cells roll under physiologic shear forces on hyaluronate and require VCAM-1, but not ICAM-1, as ligand for subsequent firm adhesion. This firm adhesion is also VLA-4 dependent, as shown by antibody inhibition. Moreover, in vivo short-term homing experiments in a model dependent on CD44 and HA demonstrate that superantigen-activated T cells require VLA-4, but not LFA-1, for entry into an inflamed peritoneal site. Thus, extravasation of activated T cells initiated by CD44 binding to HA depends upon VLA-4–mediated firm adhesion, which may explain the frequent association of these adhesion receptors with diverse chronic inflammatory processes. PMID:10712440

Sickle red cell-endothelium interactions.

PubMed

Kaul, Dhananjay K; Finnegan, Eileen; Barabino, Gilda A

2009-01-01

Periodic recurrence of painful vaso-occlusive crisis is the defining feature of sickle cell disease. Among multiple pathologies associated with this disease, sickle red cell-endothelium interaction has been implicated as a potential initiating mechanism in vaso-occlusive events. This review focuses on various interrelated mechanisms involved in human sickle red cell adhesion. We discuss in vitro and microcirculatory findings on sickle red cell adhesion, its potential role in vaso-occlusion, and the current understanding of receptor-ligand interactions involved in this pathological phenomenon. In addition, we discuss the contribution of other cellular interactions (leukocytes recruitment and leukocyte-red cell interaction) to vaso-occlusion, as observed in transgenic sickle mouse models. Emphasis is given to recently discovered adhesion molecules that play a predominant role in mediating human sickle red cell adhesion. Finally, we analyze various therapeutic approaches for inhibiting sickle red cell adhesion by targeting adhesion molecules and also consider therapeutic strategies that target stimuli involved in endothelial activation and initiation of adhesion.

Angiogenesis mediated by soluble forms of E-selectin and vascular cell adhesion molecule-1

NASA Astrophysics Data System (ADS)

Koch, Alisa E.; Halloran, Margaret M.; Haskell, Catherine J.; Shah, Manisha R.; Polverini, Peter J.

1995-08-01

ENDOTHELIAL adhesion molecules facilitate the entry of leukocytes into inflamed tissues. This in turn promotes neovascularization, a process central to the progression of rheumatoid arthritis, tumour growth and wound repair1. Here we test the hypothesis that soluble endothelial adhesion molecules promote angiogenesis2á¤-4. Human recombinant soluble E-selectin and soluble vascular cell adhesion molecule-1 induced chemotaxis of human endothelial cells in vitro and were angiogenic in rat cornea. Soluble E-selectin acted on endothelial cells in part through a sialyl Lewis-X-dependent mechanism, while soluble vascular cell adhesion molecule-1 acted on endothelial cells in part through a very late antigen (VLA)-4 dependent mechanism. The chemotactic activity of rheumatoid synovial fluid for endothelial cells, and also its angiogenic activity, were blocked by antibodies to either soluble E-selectin or soluble vascular cell adhesion molecule-1. These results suggest a novel function for soluble endothelial adhesion molecules as mediators of angiogenesis.

Strong adhesion by regulatory T cells induces dendritic cell cytoskeletal polarization and contact-dependent lethargy

PubMed Central

Mucsi, Ashley D.; Meng, Junchen; Yan, Jiacong; Zhang, Zongde; Wu, Mei; Hari, Aswin; Stenner, Melanie D.; Zheng, Wencheng; Kubes, Paul; Xia, Tie; Amrein, Matthias W.

2017-01-01

Dendritic cells are targeted by regulatory T (T reg) cells, in a manner that operates as an indirect mode of T cell suppression. In this study, using a combination of single-cell force spectroscopy and structured illumination microscopy, we analyze individual T reg cell–DC interaction events and show that T reg cells exhibit strong intrinsic adhesiveness to DCs. This increased DC adhesion reduces the ability of contacted DCs to engage other antigen-specific cells. We show that this unusually strong LFA-1–dependent adhesiveness of T reg cells is caused in part by their low calpain activities, which normally release integrin–cytoskeleton linkage, and thereby reduce adhesion. Super resolution imaging reveals that such T reg cell adhesion causes sequestration of Fascin-1, an actin-bundling protein essential for immunological synapse formation, and skews Fascin-1–dependent actin polarization in DCs toward the T reg cell adhesion zone. Although it is reversible upon T reg cell disengagement, this sequestration of essential cytoskeletal components causes a lethargic state of DCs, leading to reduced T cell priming. Our results reveal a dynamic cytoskeletal component underlying T reg cell–mediated DC suppression in a contact-dependent manner. PMID:28082358

Centrifugation assay for measuring adhesion of serially passaged bovine chondrocytes to polystyrene surfaces.

PubMed

Kaplan, David S; Hitchins, Victoria M; Vegella, Thomas J; Malinauskas, Richard A; Ferlin, Kimberly M; Fisher, John P; Frondoza, Carmelita G

2012-07-01

A major obstacle in chondrocyte-based therapy for cartilage repair is the limited availability of cells that maintain their original phenotype. Propagation of chondrocytes as monolayer cultures on polystyrene surfaces is used extensively for amplifying cell numbers. However, chondrocytes undergo a phenotypic shift when propagated in this manner and display characteristics of more adherent fibroblastic cells. Little information is available about the effect of this phenotypic shift on cellular adhesion properties. We evaluated changes in adhesion property as bovine chondrocytes were serially propagated up to five passages in monolayer culture using a centrifugation cell adhesion assay, which was based on counting of cells before and after being exposed to centrifugal dislodgement forces of 120 and 350 g. Chondrocytes proliferated well in a monolayer culture with doubling times of 2-3 days, but they appeared more fibroblastic and exhibited elongated cell morphology with continued passage. The centrifugation cell adhesion assay showed that chondrocytes became more adhesive with passage as the percentage of adherent cells after centrifugation increased and was not statistically different from the adhesion of the fibroblast cell line, L929, starting at passage 3. This increased adhesiveness correlated with a shift to a fibroblastic morphology and increased collagen I mRNA expression starting at passage 2. Our findings indicate that the centrifugation cell adhesion assay may serve as a reproducible tool to track alterations in chondrocyte phenotype during their extended propagation in culture.

Effect of Rebamipide, a Novel Antiulcer Agent, on Helicobacter pylori Adhesion to Gastric Epithelial Cells

PubMed Central

Hayashi, Shunji; Sugiyama, Toshiro; Amano, Ken-Ichi; Isogai, Hiroshi; Isogai, Emiko; Aihara, Miki; Kikuchi, Mikio; Asaka, Masahiro; Yokota, Kenji; Oguma, Keiji; Fujii, Nobuhiro; Hirai, Yoshikazu

1998-01-01

Helicobacter pylori is a major etiological agent in gastroduodenal disorders. The adhesion of H. pylori to human gastric epithelial cells is the initial step of H. pylori infection. Inhibition of H. pylori adhesion is thus a therapeutic target in the prevention of H. pylori infection. Experiments were performed to evaluate the effect of rebamipide, a novel antiulcer agent, on H. pylori adhesion to gastric epithelial cells. MKN-28 and MKN-45 cells, derived from human gastric carcinomas, were used as target cells. Ten H. pylori strains isolated from patients with chronic gastritis and gastric ulcer were used in the study. We evaluated the effect of rebamipide on H. pylori adhesion to MKN-28 and MKN-45 cells quantitatively using our previously established enzyme-linked immunosorbent assay. The adhesion of H. pylori to MKN-28 and MKN-45 cells was significantly inhibited by pretreatment of these cells with 100 μg of rebamipide per ml. However, the adhesion was not affected by the pretreatment of H. pylori with rebamipide. On the other hand, the viabilities of H. pylori, MKN-28 cells, and MKN-45 cells were not affected by rebamipide. Our studies suggest that rebamipide inhibits the adhesion of H. pylori to gastric epithelial cells. PMID:9687380

The effect of the physical properties of the substrate on the kinetics of cell adhesion and crawling studied by an axisymmetric diffusion-energy balance coupled model.

PubMed

Samadi-Dooki, Aref; Shodja, Hossein M; Malekmotiei, Leila

2015-05-14

In this paper an analytical approach to study the effect of the substrate physical properties on the kinetics of adhesion and motility behavior of cells is presented. Cell adhesion is mediated by the binding of cell wall receptors and substrate's complementary ligands, and tight adhesion is accomplished by the recruitment of the cell wall binders to the adhesion zone. The binders' movement is modeled as their axisymmetric diffusion in the fluid-like cell membrane. In order to preserve the thermodynamic consistency, the energy balance for the cell-substrate interaction is imposed on the diffusion equation. Solving the axisymmetric diffusion-energy balance coupled equations, it turns out that the physical properties of the substrate (substrate's ligand spacing and stiffness) have considerable effects on the cell adhesion and motility kinetics. For a rigid substrate with uniform distribution of immobile ligands, the maximum ligand spacing which does not interrupt adhesion growth is found to be about 57 nm. It is also found that as a consequence of the reduction in the energy dissipation in the isolated adhesion system, cell adhesion is facilitated by increasing substrate's stiffness. Moreover, the directional movement of cells on a substrate with gradients in mechanical compliance is explored with an extension of the adhesion formulation. It is shown that cells tend to move from soft to stiff regions of the substrate, but their movement is decelerated as the stiffness of the substrate increases. These findings based on the proposed theoretical model are in excellent agreement with the previous experimental observations.

Focal Adhesion-Independent Cell Migration.

PubMed

Paluch, Ewa K; Aspalter, Irene M; Sixt, Michael

2016-10-06

Cell migration is central to a multitude of physiological processes, including embryonic development, immune surveillance, and wound healing, and deregulated migration is key to cancer dissemination. Decades of investigations have uncovered many of the molecular and physical mechanisms underlying cell migration. Together with protrusion extension and cell body retraction, adhesion to the substrate via specific focal adhesion points has long been considered an essential step in cell migration. Although this is true for cells moving on two-dimensional substrates, recent studies have demonstrated that focal adhesions are not required for cells moving in three dimensions, in which confinement is sufficient to maintain a cell in contact with its substrate. Here, we review the investigations that have led to challenging the requirement of specific adhesions for migration, discuss the physical mechanisms proposed for cell body translocation during focal adhesion-independent migration, and highlight the remaining open questions for the future.

Rapid surface accumulation of NMDA receptors increases glutamatergic excitation during status epilepticus.

PubMed

Naylor, David E; Liu, Hantao; Niquet, Jerome; Wasterlain, Claude G

2013-06-01

After 1h of lithium-pilocarpine status epilepticus (SE), immunocytochemical labeling of NMDA receptor NR1 subunits reveals relocation of subunits from the interior to the cell surface of dentate gyrus granule cells and CA3 pyramidal cells. Simultaneously, an increase in NMDA-miniature excitatory postsynaptic currents (mEPSC) as well as an increase in NMDA receptor-mediated tonic currents is observed in hippocampal slices after SE. Mean-variance analysis of NMDA-mEPSCs estimates that the number of functional postsynaptic NMDA receptors per synapse increases 38% during SE, and antagonism by ifenprodil suggests that an increase in the surface representation of NR2B-containing NMDA receptors is responsible for the augmentation of both the phasic and tonic excitatory currents with SE. These results provide a potential mechanism for an enhancement of glutamatergic excitation that maintains SE and may contribute to excitotoxic injury during SE. Therapies that directly antagonize NMDA receptors may be a useful therapeutic strategy during refractory SE. Copyright © 2013 Elsevier Inc. All rights reserved.

Rapid surface accumulation of NMDA receptors increases glutamatergic excitation during status epilepticus

PubMed Central

Naylor, David E.; Liu, Hantao; Niquet, Jerome; Wasterlain, Claude G.

2017-01-01

After 1 h of lithium-pilocarpine status epilepticus (SE), immunocytochemical labeling of NMDA receptor NR1 subunits reveals relocation of subunits from the interior to the cell surface of dentate gyrus granule cells and CA3 pyramidal cells. Simultaneously, an increase in NMDA-miniature excitatory postsynaptic currents (mEPSC) as well as an increase in NMDA receptor-mediated tonic currents is observed in hippocampal slices after SE. Mean-variance analysis of NMDA-mEPSCs estimates that the number of functional postsynaptic NMDA receptors per synapse increases 38% during SE, and antagonism by ifenprodil suggests that an increase in the surface representation of NR2B-containing NMDA receptors is responsible for the augmentation of both the phasic and tonic excitatory currents with SE. These results provide a potential mechanism for an enhancement of glutamatergic excitation that maintains SE and may contribute to excitotoxic injury during SE. Therapies that directly antagonize NMDA receptors may be a useful therapeutic strategy during refractory SE. PMID:23313318

Reversing the Outcome of Synapse Elimination at Developing Neuromuscular Junctions In Vivo: Evidence for Synaptic Competition and Its Mechanism

PubMed Central

Turney, Stephen G.; Lichtman, Jeff W.

2012-01-01

During mammalian development, neuromuscular junctions and some other postsynaptic cells transition from multiple- to single-innervation as synaptic sites are exchanged between different axons. It is unclear whether one axon invades synaptic sites to drive off other inputs or alternatively axons expand their territory in response to sites vacated by other axons. Here we show that soon-to-be-eliminated axons rapidly reverse fate and grow to occupy vacant sites at a neuromuscular junction after laser removal of a stronger input. This reversal supports the idea that axons take over sites that were previously vacated. Indeed, during normal development we observed withdrawal followed by takeover. The stimulus for axon growth is not postsynaptic cell inactivity because axons grow into unoccupied sites even when target cells are functionally innervated. These results demonstrate competition at the synaptic level and enable us to provide a conceptual framework for understanding this form of synaptic plasticity. PMID:22745601

Impedimetric Analysis of the Effect of Decellularized Porcine Heart Scaffold on Human Fibrosarcoma, Endothelial, and Cardiomyocyte Cell Lines

PubMed Central

Bäcker, Henrik; Polgár, Livia; Soós, Pal; Lajkó, Eszter; Láng, Orsolya; Merkely, Bela; Szabó, Gabor; Dohmen, Pascal M.; Weymann, Alexander; Kőhidai, Laszlo

2017-01-01

Background Experiments on porcine heart scaffold represent significant assays in development of immunoneutral materials for cardiac surgery. Characterization of cell-cell and cell-scaffold interactions is essential to understand the homing process of cardiac cells into the scaffolds. Material/Methods In the present study, the highly sensitive and real-time impedimetric technique of xCELLigence SP was used to monitor cell adhesion, which is the key process of recellularization in heart scaffolds. Our objectives were: (i) to characterize the effect of decellularized porcine heart scaffold on cell adhesion of human cardiovascular cells potentially used in the recellularization process; and (ii) to investigate cell-extracellular matrix element interactions for building artificial multi-layer systems, applied as cellular models of recellularization experiments. Human fibrosarcoma, endothelial, and cardiomyocyte cells were investigated and the effect of decellularized porcine heart scaffold (HS) and fibronectin on cell adhesion was examined. Adhesion was quantified as slope of curves. Results Heart scaffold had neutral effect on cardiomyocytes as well as on endothelial cells. Adhesion of cardiomyocytes was increased by fibronectin (1.480±0.021) compared to control (0.745±0.029). The combination of fibronectin and HS induced stronger adhesion of cardiomyocytes (2.407±0.634) than fibronectin alone. Endothelial and fibrosarcoma cells showed similarly strong adhesion profiles with marked enhancer effect by fibronectin. Conclusions Decellularized porcine HS does not inhibit adhesion of human cardiovascular cells at the cell biological level, while fibronectin has strong cell adhesion-inducer effect, as well as an enhancer effect on activity of HS. Consequently, decellularized porcine hearts could be used as scaffolds for recellularization with cardiomyocytes and endothelial cells with fibronectin acting as a regulator, leading to construction of working bioartificial hearts. PMID:28493851

Interaction of tumor and host cells with adhesion and extracellular matrix molecules in the development of multiple myeloma.

PubMed

Teoh, G; Anderson, K C

1997-02-01

Adhesion molecules play an important role in the growth regulation and migration of multiple myeloma (MM) cells. They mediate homing of MM cells to the bone marrow and MM cell to bone marrow stromal cell adhesion, with resultant interleukin-6 related autocrine and paracine growth and antiapoptotic affects. Their pattern of expression on tumor cells correlates with the development of plasma cell leukemia or extramedullary disease. Clinically, expression of adhesion molecules on tumor cells or in the serum has already shown prognostic utility. Finally, since adhesion molecules are involved at multiple steps in the pathogenesis of MM, therapeutic studies may target these molecules.

Fermented soya bean (tempe) extracts reduce adhesion of enterotoxigenic Escherichia coli to intestinal epithelial cells.

PubMed

Roubos-van den Hil, P J; Nout, M J R; Beumer, R R; van der Meulen, J; Zwietering, M H

2009-03-01

This study aimed to investigate the effect of processed soya bean, during the successive stages of tempe fermentation and different fermentation times, on adhesion of enterotoxigenic Escherichia coli (ETEC) K88 to intestinal brush border cells as well as Caco-2 intestinal epithelial cells; and to clarify the mechanism of action. Tempe was prepared at controlled laboratory scale using Rhizopus microsporus var. microsporus as the inoculum. Extracts of raw, soaked and cooked soya beans reduced ETEC adhesion to brush border cells by 40%. Tempe extracts reduced adhesion by 80% or more. ETEC adhesion to Caco-2 cells reduced by 50% in the presence of tempe extracts. ETEC K88 bacteria were found to interact with soya bean extracts, and this may contribute to the observed decrease of ETEC adhesion to intestinal epithelial cells. Fermented soya beans (tempe) reduce the adhesion of ETEC to intestinal epithelial cells of pig and human origin. This reduced adhesion is caused by an interaction between ETEC K88 bacteria and soya bean compounds. The results strengthen previous observations on the anti-diarrhoeal effect of tempe. This effect indicates that soya-derived compounds may reduce adhesion of ETEC to intestinal cells in pigs as well as in humans and prevent against diarrhoeal diseases.

Formation and stability of synaptic receptor domains.

PubMed

Haselwandter, Christoph A; Calamai, Martino; Kardar, Mehran; Triller, Antoine; da Silveira, Rava Azeredo

2011-06-10

Neurotransmitter receptor molecules, concentrated in postsynaptic domains along with scaffold and a number of other molecules, are key regulators of signal transmission across synapses. Combining experiment and theory, we develop a quantitative description of synaptic receptor domains in terms of a reaction-diffusion model. We show that interactions between only receptors and scaffolds, together with the rapid diffusion of receptors on the cell membrane, are sufficient for the formation and stable characteristic size of synaptic receptor domains. Our work reconciles long-term stability of synaptic receptor domains with rapid turnover and diffusion of individual receptors, and suggests novel mechanisms for a form of short-term, postsynaptic plasticity.

Functional Evaluation of Biological Neurotoxins in Networked Cultures of Stem Cell-derived Central Nervous System Neurons

DTIC Science & Technology

2015-02-05

botulism or tetanus , whole-cell patch clamp electrophysiology was used to quantify spontaneous miniature excitory post-synaptic currents (mEPSCs) in...ESNs exposed to tetanus neurotoxin (TeNT) or botulinum neurotoxin (BoNT) serotypes / A-/G. In all cases, ESNs exhibited near-complete loss of synaptic

Activity-induced Ca2+ signaling in perisynaptic Schwann cells of the early postnatal mouse is mediated by P2Y1 receptors and regulates muscle fatigue

PubMed Central

Heredia, Dante J; Feng, Cheng-Yuan; Hennig, Grant W; Renden, Robert B

2018-01-01

Perisynaptic glial cells respond to neural activity by increasing cytosolic calcium, but the significance of this pathway is unclear. Terminal/perisynaptic Schwann cells (TPSCs) are a perisynaptic glial cell at the neuromuscular junction that respond to nerve-derived substances such as acetylcholine and purines. Here, we provide genetic evidence that activity-induced calcium accumulation in neonatal TPSCs is mediated exclusively by one subtype of metabotropic purinergic receptor. In P2ry1 mutant mice lacking these responses, postsynaptic, rather than presynaptic, function was altered in response to nerve stimulation. This impairment was correlated with a greater susceptibility to activity-induced muscle fatigue. Interestingly, fatigue in P2ry1 mutants was more greatly exacerbated by exposure to high potassium than in control mice. High potassium itself increased cytosolic levels of calcium in TPSCs, a response which was also reduced P2ry1 mutants. These results suggest that activity-induced calcium responses in TPSCs regulate postsynaptic function and muscle fatigue by regulating perisynaptic potassium. PMID:29384476

The molecular mechanism of mediation of adsorbed serum proteins to endothelial cells adhesion and growth on biomaterials.

PubMed

Yang, Dayun; Lü, Xiaoying; Hong, Ying; Xi, Tingfei; Zhang, Deyuan

2013-07-01

To explore molecular mechanism of mediation of adsorbed proteins to cell adhesion and growth on biomaterials, this study examined endothelial cell adhesion, morphology and viability on bare and titanium nitride (TiN) coated nickel titanium (NiTi) alloys and chitosan film firstly, and then identified the type and amount of serum proteins adsorbed on the three surfaces by proteomic technology. Subsequently, the mediation role of the identified proteins to cell adhesion and growth was investigated with bioinformatics analyses, and further confirmed by a series of cellular and molecular biological experiments. Results showed that the type and amount of adsorbed serum proteins associated with cell adhesion and growth was obviously higher on the alloys than on the chitosan film, and these proteins mediated endothelial cell adhesion and growth on the alloys via four ways. First, proteins such as adiponectin in the adsorbed protein layer bound with cell surface receptors to generate signal transduction, which activated cell surface integrins through increasing intracellular calcium level. Another way, thrombospondin 1 in the adsorbed protein layer promoted TGF-β signaling pathway activation and enhanced integrins expression. The third, RGD sequence containing proteins such as fibronectin 1, vitronectin and thrombospondin 1 in the adsorbed protein layer bound with activated integrins to activate focal adhesion pathway, increased focal adhesion formation and actin cytoskeleton organization and mediated cell adhesion and spreading. In addition, the activated focal adhesion pathway promoted the expression of cell growth related genes and resulted in cell proliferation. The fourth route, coagulation factor II (F2) and fibronectin 1 in the adsorbed protein layer bound with cell surface F2 receptor and integrin, activated regulation of actin cytoskeleton pathway and regulated actin cytoskeleton organization. Copyright © 2013 Elsevier Ltd. All rights reserved.

Design rules for biomolecular adhesion: lessons from force measurements.

PubMed

Leckband, Deborah

2010-01-01

Cell adhesion to matrix, other cells, or pathogens plays a pivotal role in many processes in biomolecular engineering. Early macroscopic methods of quantifying adhesion led to the development of quantitative models of cell adhesion and migration. The more recent use of sensitive probes to quantify the forces that alter or manipulate adhesion proteins has revealed much greater functional diversity than was apparent from population average measurements of cell adhesion. This review highlights theoretical and experimental methods that identified force-dependent molecular properties that are central to the biological activity of adhesion proteins. Experimental and theoretical methods emphasized in this review include the surface force apparatus, atomic force microscopy, and vesicle-based probes. Specific examples given illustrate how these tools have revealed unique properties of adhesion proteins and their structural origins.

Modeling cell adhesion and proliferation: a cellular-automata based approach.

PubMed

Vivas, J; Garzón-Alvarado, D; Cerrolaza, M

Cell adhesion is a process that involves the interaction between the cell membrane and another surface, either a cell or a substrate. Unlike experimental tests, computer models can simulate processes and study the result of experiments in a shorter time and lower costs. One of the tools used to simulate biological processes is the cellular automata, which is a dynamic system that is discrete both in space and time. This work describes a computer model based on cellular automata for the adhesion process and cell proliferation to predict the behavior of a cell population in suspension and adhered to a substrate. The values of the simulated system were obtained through experimental tests on fibroblast monolayer cultures. The results allow us to estimate the cells settling time in culture as well as the adhesion and proliferation time. The change in the cells morphology as the adhesion over the contact surface progress was also observed. The formation of the initial link between cell and the substrate of the adhesion was observed after 100 min where the cell on the substrate retains its spherical morphology during the simulation. The cellular automata model developed is, however, a simplified representation of the steps in the adhesion process and the subsequent proliferation. A combined framework of experimental and computational simulation based on cellular automata was proposed to represent the fibroblast adhesion on substrates and changes in a macro-scale observed in the cell during the adhesion process. The approach showed to be simple and efficient.

Atomic Force Microscopy Mechanical Mapping of Micropatterned Cells Shows Adhesion Geometry-Dependent Mechanical Response on Local and Global Scales

PubMed Central

Rigato, Annafrancesca; Rico, Felix; Eghiaian, Frédéric; Piel, Mathieu; Scheuring, Simon

2015-01-01

In multicellular organisms cell shape and organization are dictated by cell-cell or cell-extracellular matrix adhesion interactions. Adhesion complexes crosstalk with the cytoskeleton enabling cells to sense their mechanical environment. Unfortunately, most of cell biology studies, and cell mechanics studies in particular, are conducted on cultured cells adhering to a hard, homogeneous and unconstrained substrate with non-specific adhesion sites – thus far from physiological and reproducible conditions. Here, we grew cells on three different fibronectin patterns with identical overall dimensions but different geometries (▽, T and Y), and investigated their topography and mechanics by atomic force microscopy (AFM). The obtained mechanical maps were reproducible for cells grown on patterns of the same geometry, revealing pattern-specific subcellular differences. We found that local Young’s moduli variations are related to the cell adhesion geometry. Additionally, we detected local changes of cell mechanical properties induced by cytoskeletal drugs. We thus provide a method to quantitatively and systematically investigate cell mechanics and their variations, and present further evidence for a tight relation between cell adhesion and mechanics. PMID:26013956

Atomic Force Microscopy Mechanical Mapping of Micropatterned Cells Shows Adhesion Geometry-Dependent Mechanical Response on Local and Global Scales.

PubMed

Rigato, Annafrancesca; Rico, Felix; Eghiaian, Frédéric; Piel, Mathieu; Scheuring, Simon

2015-06-23

In multicellular organisms, cell shape and organization are dictated by cell-cell or cell-extracellular matrix adhesion interactions. Adhesion complexes crosstalk with the cytoskeleton enabling cells to sense their mechanical environment. Unfortunately, most of cell biology studies, and cell mechanics studies in particular, are conducted on cultured cells adhering to a hard, homogeneous, and unconstrained substrate with nonspecific adhesion sites, thus far from physiological and reproducible conditions. Here, we grew cells on three different fibronectin patterns with identical overall dimensions but different geometries (▽, T, and Y), and investigated their topography and mechanics by atomic force microscopy (AFM). The obtained mechanical maps were reproducible for cells grown on patterns of the same geometry, revealing pattern-specific subcellular differences. We found that local Young's moduli variations are related to the cell adhesion geometry. Additionally, we detected local changes of cell mechanical properties induced by cytoskeletal drugs. We thus provide a method to quantitatively and systematically investigate cell mechanics and their variations, and present further evidence for a tight relation between cell adhesion and mechanics.

Emergence of collective propulsion through cell-cell adhesion.

PubMed

Matsushita, Katsuyoshi

2018-04-01

The mechanisms driving the collective movement of cells remain poorly understood. To contribute toward resolving this mystery, a model was formulated to theoretically explore the possible functions of polarized cell-cell adhesion in collective cell migration. The model consists of an amoeba cell with polarized cell-cell adhesion, which is controlled by positive feedback with cell motion. This model cell has no persistent propulsion and therefore exhibits a simple random walk when in isolation. However, at high density, these cells acquire collective propulsion and form ordered movement. This result suggests that cell-cell adhesion has a potential function, which induces collective propulsion with persistence.

Emergence of collective propulsion through cell-cell adhesion

NASA Astrophysics Data System (ADS)

Matsushita, Katsuyoshi

2018-04-01

The mechanisms driving the collective movement of cells remain poorly understood. To contribute toward resolving this mystery, a model was formulated to theoretically explore the possible functions of polarized cell-cell adhesion in collective cell migration. The model consists of an amoeba cell with polarized cell-cell adhesion, which is controlled by positive feedback with cell motion. This model cell has no persistent propulsion and therefore exhibits a simple random walk when in isolation. However, at high density, these cells acquire collective propulsion and form ordered movement. This result suggests that cell-cell adhesion has a potential function, which induces collective propulsion with persistence.

Surfactant Functionalization Induces Robust, Differential Adhesion of Tumor Cells and Blood Cells to Charged Nanotube-Coated Biomaterials Under Flow

PubMed Central

Mitchell, Michael J.; Castellanos, Carlos A.; King, Michael R.

2015-01-01

The metastatic spread of cancer cells from the primary tumor to distant sites leads to a poor prognosis in cancers originating from multiple organs. Increasing evidence has linked selectin-based adhesion between circulating tumor cells (CTCs) and endothelial cells of the microvasculature to metastatic dissemination, in a manner similar to leukocyte adhesion during inflammation. Functionalized biomaterial surfaces hold promise as a diagnostic tool to separate CTCs and potentially treat metastasis, utilizing antibody and selectin-mediated interactions for cell capture under flow. However, capture at high purity levels is challenged by the fact that CTCs and leukocytes both possess selectin ligands. Here, a straightforward technique to functionalize and alter the charge of naturally occurring halloysite nanotubes using surfactants is reported to induce robust, differential adhesion of tumor cells and blood cells to nanotube-coated surfaces under flow. Negatively charged sodium dodecanoate-functionalized nanotubes simultaneously enhanced tumor cell capture while negating leukocyte adhesion, both in the presence and absence of adhesion proteins, and can be utilized to isolate circulating tumor cells regardless of biomarker expression. Conversely, diminishing nanotube charge via functionalization with decyltrimethylammonium bromide both abolished tumor cell capture while promoting leukocyte adhesion. PMID:25934290

Reinjury risk of nano-calcium oxalate monohydrate and calcium oxalate dihydrate crystals on injured renal epithelial cells: aggravation of crystal adhesion and aggregation

PubMed Central

Gan, Qiong-Zhi; Sun, Xin-Yuan; Bhadja, Poonam; Yao, Xiu-Qiong; Ouyang, Jian-Ming

2016-01-01

Background Renal epithelial cell injury facilitates crystal adhesion to cell surface and serves as a key step in renal stone formation. However, the effects of cell injury on the adhesion of nano-calcium oxalate crystals and the nano-crystal-induced reinjury risk of injured cells remain unclear. Methods African green monkey renal epithelial (Vero) cells were injured with H2O2 to establish a cell injury model. Cell viability, superoxide dismutase (SOD) activity, malonaldehyde (MDA) content, propidium iodide staining, hematoxylin–eosin staining, reactive oxygen species production, and mitochondrial membrane potential (Δψm) were determined to examine cell injury during adhesion. Changes in the surface structure of H2O2-injured cells were assessed through atomic force microscopy. The altered expression of hyaluronan during adhesion was examined through laser scanning confocal microscopy. The adhesion of nano-calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) crystals to Vero cells was observed through scanning electron microscopy. Nano-COM and COD binding was quantitatively determined through inductively coupled plasma emission spectrometry. Results The expression of hyaluronan on the cell surface was increased during wound healing because of Vero cell injury. The structure and function of the cell membrane were also altered by cell injury; thus, nano-crystal adhesion occurred. The ability of nano-COM to adhere to the injured Vero cells was higher than that of nano-COD crystals. The cell viability, SOD activity, and Δψm decreased when nano-crystals attached to the cell surface. By contrast, the MDA content, reactive oxygen species production, and cell death rate increased. Conclusion Cell injury contributes to crystal adhesion to Vero cell surface. The attached nano-COM and COD crystals can aggravate Vero cell injury. As a consequence, crystal adhesion and aggregation are enhanced. These findings provide further insights into kidney stone formation. PMID:27382277

Quantitative characterization of mesenchymal stem cell adhesion to the articular cartilage surface.

PubMed

Hung, Ben P; Babalola, Omotunde M; Bonassar, Lawrence J

2013-12-01

There has been great interest in use of mesenchymal stem cell (MSC)-based therapies for cartilage repair. Most recently, treatments involving intra-articular injection of MSCs have shown great promise for cartilage repair and arthritis therapy, which rely on MSC adhesion to cartilage. While there is some information on chondrocyte adhesion to cartilage, there is relatively little known about the kinetics and strength of MSC adhesion to cartilage. The goals of this study were as follows: (1) to quantify the kinetics and strength of adhesion of marrow-derived MSCs to articular cartilage using standard laboratory hardware; (2) to compare this adhesion behavior to that of articular chondrocytes; and (3) to assess the effect of serial monolayer culture on MSC adhesion. First through fourth passage MSCs and primary articular chondrocytes were allowed to adhere to the articular surface of cartilage disks for up to 30 h and the number of adhered cells was recorded to quantify adhesion kinetics. After 30 h, adherent cells were subjected to centrifugal shear to determine adhesion strength, quantified as the shear necessary to detach half the adhered cells (σ50 ). The number of adhered MSCs and adhesion strength increased with passage number and MSCs adhered more strongly than did primary articular chondrocytes. As such, the kinetics and strength of MSC adhesion to cartilage is not dramatically lower than that for articular chondrocytes. This protocol for assessing cell adhesion to cartilage is simple to implement and may represent an important screening tool for assessing the efficacy of cell-based therapies for cartilage repair. Copyright © 2013 Wiley Periodicals, Inc., a Wiley Company.

Surgery-derived reactive oxygen species produced by polymorphonuclear leukocytes promote tumor recurrence: studies in an in vitro model.

PubMed

van Grevenstein, Wilhelmina M U; Aalbers, Arend G J; Ten Raa, Sander; Sluiter, Wim; Hofland, Leo J; Jeekel, Hans; van Eijck, Casper H J

2007-06-01

Tissue injury induces the acute phase response, aimed at minimizing damage and starting the healing process. Polymorphonuclear leukocytes (PMNs) respond to the presence of specific chemoattractants and begin to appear in large numbers. The aim of this study was to investigate the influence of reactive oxygen species (ROS) produced by PMNs on the interaction between colon carcinoma cells and mesothelial cells. An experimental human in vitro model was designed using Caco-2 colon carcinoma cells and primary cultures of mesothelial cells. Tumor cell adhesion to a mesothelial monolayer was assessed after preincubation of the mesothelium with stimulated PMNs and unstimulated PMNs. Mesothelial cells were also incubated with xanthine/xanthine oxidase (X/XO) complex producing ROS after which adhesion of Caco-2 cells was investigated and the expression of adhesion molecules (ICAM-1, VCAM-1, and CD44) by means of enzyme immunoassay. In the control situation the average adhesion of Caco-2 cells to the mesothelial monolayers was 23%. Mesothelial monolayers incubated with unstimulated PMNs showed a 25% increase of tumor cell adhesion (P < 0.05). The adhesion of tumor to the monolayers incubated with the N-formyl-methionyl-leucyl-phenylalanine-stimulated PMNs increased with 40% (P < 0.01). Incubation of the mesothelium with X/XO resulted in an enhancement of adhesion of Caco-2 cells of 70% and an up-regulation of expression of ICAM-1, VCAM-1, and CD44. This study reveals an increase of tumor cell adhesion to the mesothelium induced by incubating the mesothelial monolayers with PMNs. PMNs are producing a number of products, like proteolytic enzymes, cytokines, and ROS. These factors up-regulate the expression of adhesion molecules and in that way stimulate the adhesion of tumor to the mesothelium.

The evaluation of p,p'-DDT exposure on cell adhesion of hepatocellular carcinoma.

PubMed

Jin, Xiaoting; Chen, Meilan; Song, Li; Li, Hanqing; Li, Zhuoyu

2014-08-01

Many studies have found a positive association between the progression of hepatocellular carcinoma and DDT exposure. These studies mainly focus on the effect of DDT exposure on cell proliferation and epithelial to mesenchymal transition (EMT) promotion. However, the influence of DDT on cell adhesion of hepatocellular carcinoma remains to be unclear. The aim of our study was to determine the effect of p,p'-DDT on cell adhesion of hepatocellular carcinoma in vitro and in vivo. The data showed that p,p'-DDT, exposing HepG2 cells for 6 days, decreased cell-cell adhesion and elevated cell-matrix adhesion. Strikingly, p,p'-DDT increased reactive oxygen species (ROS) content, and this was accompanied by the activation of JAK/STAT3 pathway. Moreover, ROS inhibitor supplement reversed these effects significantly. However, the addition of ER inhibitor, ICI, had no effect on the p,p'-DDT-induced effects. p,p'-DDT altered the mRNA levels of related adhesion molecules, including inhibition of E-cadherin and promotion of N-cadherin along with CD29. Interestingly, the p,p'-DDT-altered adhesion molecules could be reversed with JAK inhibitor or STAT3 inhibitor. Likewise, p,p'-DDT stimulated the JAK/STAT3 pathway in nude mice, as well as altered the mRNA levels of E-cadherin, N-cadherin, and CD29. Taken together, these results indicate that p,p'-DDT profoundly promotes the adhesion process by decreasing cell-cell adhesion and inducing cell-matrix adhesion via the ROS-mediated JAK/STAT3 pathway. All these events account for the carcinogenic potential of p,p'-DDT in liver. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Becker, Ines; Schillig, Cora

A double-sided adhesive metal-based tape for use as contacting aid for SOFC fuel cells is provided. The double-sided metal-based adhesive tape is suitable for simplifying the construction of cell bundles. The double-sided metal-based adhesive tape is used for electrical contacting of the cell connector with the anode and for electrical contacting of the interconnector of the fuel cells with the cell connector. A method for producing the double-sided adhesive metal-base tape is also provided.

Targeted cell adhesion on selectively micropatterned polymer arrays on a poly(dimethylsiloxane) surface.

PubMed

Tang, Linzhi; Min, Junhong; Lee, Eun-Cheol; Kim, Jong Sung; Lee, Nae Yoon

2010-02-01

Herein, we introduce the fabrication of polymer micropattern arrays on a chemically inert poly(dimethylsiloxane) (PDMS) surface and employ them for the selective adhesion of cells. To fabricate the micropattern arrays, a mercapto-ester-based photocurable adhesive was coated onto a mercaptosilane-coated PDMS surface and photopolymerized using a photomask to obtain patterned arrays at the microscale level. Robust polymer patterns, 380 microm in diameter, were successfully fabricated onto a PDMS surface, and cells were selectively targeted toward the patterned regions. Next, the performance of the cell adhesion was observed by anchoring cell adhesive linker, an RGD oligopeptide, on the surface of the mercapto-ester-based adhesive-cured layer. The successful anchoring of the RGD linker was confirmed through various surface characterizations such as water contact angle measurement, XPS analysis, FT-IR analysis, and AFM measurement. The micropatterning of a photocurable adhesive onto a PDMS surface can provide high structural rigidity, a highly-adhesive surface, and a physical pathway for selective cell adhesion, while the incorporated polymer micropattern arrays inside a PDMS microfluidic device can serve as a microfluidic platform for disease diagnoses and high-throughput drug screening.

Perfluorocarbon induces alveolar epithelial cell response through structural and mechanical remodeling.

PubMed

André Dias, Sofia; Planus, Emmanuelle; Angely, Christelle; Lotteau, Luc; Tissier, Renaud; Filoche, Marcel; Louis, Bruno; Pelle, Gabriel; Isabey, Daniel

2018-02-15

During total liquid ventilation, lung cells are exposed to perfluorocarbon (PFC) whose chemophysical properties highly differ from standard aqueous cell feeding medium (DMEM). We herein perform a systematic study of structural and mechanical properties of A549 alveolar epithelial cells in order to characterize their response to PFC exposure, using DMEM as control condition. Changes in F-actin structure, focal adhesion density and glycocalyx distribution are evaluated by confocal fluorescent microscopy. Changes in cell mechanics and adhesion are measured by multiscale magnetic twisting cytometry (MTC). Two different microrheological models (single Voigt and power law) are used to analyze the cell mechanics characterized by cytoskeleton (CSK) stiffness and characteristic relaxation times. Cell-matrix adhesion is analyzed using a stochastic multibond deadhesion model taking into account the non-reversible character of the cell response, allowing us to quantify the adhesion weakness and the number of associated bonds. The roles of F-actin structure and glycocalyx layer are evaluated by depolymerizing F-actin and degrading glycocalyx, respectively. Results show that PFC exposure consistently induces F-actin remodeling, CSK softening and adhesion weakening. These results demonstrate that PFC triggers an alveolar epithelial cell response herein evidenced by a decay in intracellular CSK tension, an adhesion weakening and a glycocalyx layer redistribution. These PFC-induced cell adjustments are consistent with the hypothesis that cells respond to a decrease in adhesion energy at cell surface. This adhesion energy can be even further reduced in the presence of surfactant adsorbed at the cell surface.

Dissecting the roles of ROCK isoforms in stress-induced cell detachment.

PubMed

Shi, Jianjian; Surma, Michelle; Zhang, Lumin; Wei, Lei

2013-05-15

The homologous Rho kinases, ROCK1 and ROCK2, are involved in stress fiber assembly and cell adhesion and are assumed to be functionally redundant. Using mouse embryonic fibroblasts (MEFs) derived from ROCK1(-/-) and ROCK2(-/-) mice, we have recently reported that they play different roles in regulating doxorubicin-induced stress fiber disassembly and cell detachment: ROCK1 is involved in destabilizing the actin cytoskeleton and cell detachment, whereas ROCK2 is required for stabilizing the actin cytoskeleton and cell adhesion. Here, we present additional insights into the roles of ROCK1 and ROCK2 in regulating stress-induced impairment of cell-matrix and cell-cell adhesion. In response to doxorubicin, ROCK1(-/-) MEFs showed significant preservation of both focal adhesions and adherens junctions, while ROCK2(-/-) MEFs exhibited impaired focal adhesions but preserved adherens junctions compared with the wild-type MEFs. Additionally, inhibition of focal adhesion or adherens junction formations by chemical inhibitors abolished the anti-detachment effects of ROCK1 deletion. Finally, ROCK1(-/-) MEFs, but not ROCK2(-/-) MEFs, also exhibited preserved central stress fibers and reduced cell detachment in response to serum starvation. These results add new insights into a novel mechanism underlying the anti-detachment effects of ROCK1 deletion mediated by reduced peripheral actomyosin contraction and increased actin stabilization to promote cell-cell and cell-matrix adhesion. Our studies further support the differential roles of ROCK isoforms in regulating stress-induced loss of central stress fibers and focal adhesions as well as cell detachment.

Galaptin Mediates the Effect of Hypergravity on Vascular Smooth Muscle cell (SMC) Adhesion to Laminin Containing Matrices

NASA Technical Reports Server (NTRS)

Enahora, Fatisha T.; Bosah, Francis N.; Harris-Hooker, Sandra; Sanford, Gary L.

1997-01-01

Galaptin, an endogenous beta-galactoside specific lectin, has been reported to bind to laminin and subsequently decrease the binding of SMC. Cellular function depend on cell:matrix interactions. Hypergravity (HGrav) affect a number of cellular functions, yet little is known about its affect on cell adhesion. We examined the possibility that galaptin mediates the effects of hypergravity on SMC adherence. Confluent primate aorta SMC cultures were subjected to Hgrav (centrifuged at 6G) for 24 and 48 hr. Cells were non-enzymatically dispersed, pretreated with antisense (AS-oligo) or control sense (SS-oligo) oligonucleotides to galaptin mRNA (0.01 micro g/ml), then seeded in uncoated or ECL-matrix coated plates. Adhesion of cells were monitored after 6 hr. HGrav increased adhesion by 100-300% compared to controls. AS-oligo decreased adhesion for both HGrav and control cells. SS-oligo did not affect adhesion for either HGrav or control cells. These studies show that HGrav affects cell adhesion and that galaptin expression is required for this effect.

The presynaptic ribbon maintains vesicle populations at the hair cell afferent fiber synapse

PubMed Central

Becker, Lars; Schnee, Michael E; Niwa, Mamiko; Sun, Willy; Maxeiner, Stephan; Talaei, Sara; Kachar, Bechara; Rutherford, Mark A

2018-01-01

The ribbon is the structural hallmark of cochlear inner hair cell (IHC) afferent synapses, yet its role in information transfer to spiral ganglion neurons (SGNs) remains unclear. We investigated the ribbon’s contribution to IHC synapse formation and function using KO mice lacking RIBEYE. Despite loss of the entire ribbon structure, synapses retained their spatiotemporal development and KO mice had a mild hearing deficit. IHCs of KO had fewer synaptic vesicles and reduced exocytosis in response to brief depolarization; a high stimulus level rescued exocytosis in KO. SGNs exhibited a lack of sustained excitatory postsynaptic currents (EPSCs). We observed larger postsynaptic glutamate receptor plaques, potentially compensating for the reduced EPSC rate in KO. Surprisingly, large-amplitude EPSCs were maintained in KO, while a small population of low-amplitude slower EPSCs was increased in number. The ribbon facilitates signal transduction at physiological stimulus levels by retaining a larger residency pool of synaptic vesicles. PMID:29328021

E-selectin: sialyl Lewis, a dependent adhesion of colon cancer cells, is inhibited differently by antibodies against E-selectin ligands.

PubMed

Srinivas, U; Påhlsson, P; Lundblad, A

1996-09-01

Recent studies have demonstrated that selectins, a new family of cell-adhesion molecules with similar domain structures, mediate the adhesion of peripheral blood cells to interleukin-1 (IL-1)-activated endothelium. In the present study the authors evaluated the role of E-selectin-Sialyl Lewis x (SLe(x))/ Sialyl Lewis a (SLe(a)) interaction in mediating in vitro adhesion of two colon cancer cell lines, HT-29 and COLO 201, to human umbilical cord endothelial cells (HUVEC). Colon cancer cell lines had a strong expression of blood group-related carbohydrate epitopes as evaluated by fluorescence-activated cell sorter (FACS) analysis. It was established that adhesion of HT-29 and COLO 201 cells to IL-1 stimulated HUVEC was calcium dependent and could be inhibited by a monoclonal antibody directed against E-selectin. Prior incubation of cells with two different antibodies directed against SLe(x) and antibodies directed against related Lewis epitopes, Le(x) and Le(a), had no significant effect on adhesion. Three antibodies directed against SLe(a) differed in their capacity to inhibit the adhesion of HT-29 and COLO 201 cells to HUVEC. Only one antibody directed against the SLe(a) structure was effective in inhibiting adhesion of both COLO 201 and HT-29 cells. The difference could not be attributed to titre, the type or number of glycoproteins, or to a difference in the amount of SLe(a) present on individual proteins, suggesting that presence and right presentation of SLe(a) epitope might be important for adhesion of colon cancer cells. Finally, in the in vitro system used, adhesion of HT-29 and COLO 201 cells to activated HUVEC is mediated predominantly by E-selectin/SLe(a) interaction. SLe(x) and related epitopes, Le(x) and Le(a), seem to have limited relevance for colon cancer cell recognition of E-selectin.

Neuron-Glia Adhesion is Inhibited by Antibodies to Neural Determinants

NASA Astrophysics Data System (ADS)

Grumet, M.; Rutishauser, U.; Edelman, G. M.

1983-10-01

Suspensions of embryonic chick neuronal cells adhered to monolayers of glial cells, but few neurons bound to control monolayers of fibroblastic cells from meninges or skin. Neuronal cell-glial cell adhesion was inhibited by prior incubation of the neurons with Fab' fragments of antibodies to neuronal membranes. In contrast, antibodies to the neural cell adhesion molecule (N-CAM) did not inhibit the binding. These results suggest that a specific adhesive mechanism between neurons and glial cells exists and that it is mediated by CAM's that differ from those so far identified.

Withaferin A inhibits tumor necrosis factor-alpha-induced expression of cell adhesion molecules by inactivation of Akt and NF-kappaB in human pulmonary epithelial cells.

PubMed

Oh, Jung Hwa; Kwon, Taeg Kyu

2009-05-01

We here investigated the functional effect of withaferin A on airway inflammation and its action mechanism. Withaferin A inhibited the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in human lung epithelial A549 cells stimulated with tumor necrosis factor-alpha (TNF-alpha), resulting in the suppression of leukocyte adhesion to lung epithelial A549 cells. In addition, withaferin A inhibited TNF-alpha-induced expression of adhesion molecules (ICAM-1 and VCAM-1) protein and mRNA in a dose-dependent manner. Withaferin A prevented DNA binding activity of nuclear factor-kappaB (NF-kappaB) and nuclear translocation of NF-kappaB. It also inhibited phosphorylation of Akt and extracellular signal-regulated kinase (ERK), which are upstream in the regulation of adhesion molecules by TNF-alpha. Furthermore, withaferin A inhibited U937 monocyte adhesion to A549 cells stimulated by TNF-alpha, suggesting that it may inhibit the binding of these cells by regulating the expression of critical adhesion molecules by TNF-alpha. Taken together, these results suggest that withaferin A inhibits cell adhesion through inhibition of ICAM-1 and VCAM-1 expression, at least in part, by blocking Akt and down-regulating NF-kappaB activity.

Mesenchymal Stem/Multipotent Stromal Cells from Human Decidua Basalis Reduce Endothelial Cell Activation.

PubMed

Alshabibi, Manal A; Al Huqail, Al Joharah; Khatlani, Tanvir; Abomaray, Fawaz M; Alaskar, Ahmed S; Alawad, Abdullah O; Kalionis, Bill; Abumaree, Mohamed Hassan

2017-09-15

Recently, we reported the isolation and characterization of mesenchymal stem cells from the decidua basalis of human placenta (DBMSCs). These cells express a unique combination of molecules involved in many important cellular functions, which make them good candidates for cell-based therapies. The endothelium is a highly specialized, metabolically active interface between blood and the underlying tissues. Inflammatory factors stimulate the endothelium to undergo a change to a proinflammatory and procoagulant state (ie, endothelial cell activation). An initial response to endothelial cell activation is monocyte adhesion. Activation typically involves increased proliferation and enhanced expression of adhesion and inflammatory markers by endothelial cells. Sustained endothelial cell activation leads to a type of damage to the body associated with inflammatory diseases, such as atherosclerosis. In this study, we examined the ability of DBMSCs to protect endothelial cells from activation through monocyte adhesion, by modulating endothelial proliferation, migration, adhesion, and inflammatory marker expression. Endothelial cells were cocultured with DBMSCs, monocytes, monocyte-pretreated with DBMSCs and DBMSC-pretreated with monocytes were also evaluated. Monocyte adhesion to endothelial cells was examined following treatment with DBMSCs. Expression of endothelial cell adhesion and inflammatory markers was also analyzed. The interaction between DBMSCs and monocytes reduced endothelial cell proliferation and monocyte adhesion to endothelial cells. In contrast, endothelial cell migration increased in response to DBMSCs and monocytes. Endothelial cell expression of adhesion and inflammatory molecules was reduced by DBMSCs and DBMSC-pretreated with monocytes. The mechanism of reduced endothelial proliferation involved enhanced phosphorylation of the tumor suppressor protein p53. Our study shows for the first time that DBMSCs protect endothelial cells from activation by inflammation triggered by monocyte adhesion and increased endothelial cell proliferation. These events are manifest in inflammatory diseases, such as atherosclerosis. Therefore, our results suggest that DBMSCs could be usefully employed as a therapeutic strategy for atherosclerosis.

Clustering of adhesion receptors following exposure of insect blood cells to foreign surfaces.

PubMed

Nardi, James B; Zhuang, Shufei; Pilas, Barbara; Bee, Charles Mark; Kanost, Michael R

2005-05-01

Cell-mediated immune responses of insects involve interactions of two main classes of blood cells (hemocytes) known as granular cells and plasmatocytes. In response to a foreign surface, these hemocytes suddenly transform from circulating, non-adherent cells to cells that interact and adhere to each other and the foreign surface. This report presents evidence that during this adhesive transformation the extracellular matrix (ECM) proteins lacunin and a ligand for peanut agglutinin (PNA) lectin are released by granular cells and bind to surfaces of both granular cells and plasmatocytes. ECM protein co-localizes on cell surfaces with the adhesive receptors integrin and neuroglian, a member of the immunoglobulin superfamily. The ECM protein(s) secreted by granular cells are hypothesized to interact with adhesion receptors such as neuroglian and integrin by cross linking and clustering them on hemocyte surfaces. This clustering of receptors is known to enhance the adhesiveness (avidity) of interacting mammalian immune cells. The formation of ring-shaped clusters of these adhesion receptors on surfaces of insect immune cells represents an evolutionary antecedent of the mammalian immunological synapse.

GABAergic miniature postsynaptic currents in septal neurons show differential allosteric sensitivity after binge-like ethanol exposure.

PubMed

DuBois, Dustin W; Trzeciakowski, Jerome P; Parrish, Alan R; Frye, Gerald D

2006-05-17

Binge-like ethanol treatment of septal neurons blunts GABAAR-mediated miniature postsynaptic currents (mPSCs), suggesting it arrests synaptic development. Ethanol may disrupt postsynaptic maturation by blunting feedback signaling through immature GABAARs. Here, the impact of ethanol on the sensitivity of mPSCs to zolpidem, zinc and 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha-OH-DHP) was tested. The decay phase of mPSCs showed concentration-dependent potentiation by zolpidem (0.03-100 microM), which was substantially blunted after ethanol exposure. Since zolpidem potentiation exhibited a substantial age-dependent increase in untreated neurons, this finding supported the idea that ethanol arrests synaptic development. GABAAR alpha1 subunit protein also increased with age in untreated neurons, paralleling enhanced sensitivity to zolpidem. Surprisingly, alpha1 levels were not reduced by binge ethanol even though mPSCs were relatively zolpidem-insensitive. Zinc (3-30 microM) decreased mPSC parameters in a concentration- and age-related manner with older untreated cells showing less inhibition. However, there was no increase in mPSC zinc sensitivity after binge ethanol as would be expected if a general arrest of synaptic maturation had occurred. 3alpha-OH-DHP (3-1000 nM) induced concentration-dependent potentiation of mPSC decay. Although potentiation was age-independent, binge ethanol treatment exaggerated sensitivity to this neurosteroid. Finally, chronic picrotoxin pretreatment (100 microM) intended to mimic GABAAR inhibition from ethanol pretreatment did not significantly change mPSC modulation by zolpidem, zinc or 3alpha-OH-DHP. These results suggest that binge ethanol treatment selectively arrests a subset of processes important for maturation of postsynaptic GABAA Rs. However, it is unlikely that ethanol causes a broad arrest of postsynaptic development through a direct inhibition of GABAAR signaling.

Feedforward and feedback inhibition in neostriatal GABAergic spiny neurons.

PubMed

Tepper, James M; Wilson, Charles J; Koós, Tibor

2008-08-01

There are two distinct inhibitory GABAergic circuits in the neostriatum. The feedforward circuit consists of a relatively small population of GABAergic interneurons that receives excitatory input from the neocortex and exerts monosynaptic inhibition onto striatal spiny projection neurons. The feedback circuit comprises the numerous spiny projection neurons and their interconnections via local axon collaterals. This network has long been assumed to provide the majority of striatal GABAergic inhibition and to sharpen and shape striatal output through lateral inhibition, producing increased activity in the most strongly excited spiny cells at the expense of their less strongly excited neighbors. Recent results, mostly from recording experiments of synaptically connected pairs of neurons, have revealed that the two GABAergic circuits differ markedly in terms of the total number of synapses made by each, the strength of the postsynaptic response detected at the soma, the extent of presynaptic convergence and divergence and the net effect of the activation of each circuit on the postsynaptic activity of the spiny neuron. These data have revealed that the feedforward inhibition is powerful and widespread, with spiking in a single interneuron being capable of significantly delaying or even blocking the generation of spikes in a large number of postsynaptic spiny neurons. In contrast, the postsynaptic effects of spiking in a single presynaptic spiny neuron on postsynaptic spiny neurons are weak when measured at the soma, and unable to significantly affect spike timing or generation. Further, reciprocity of synaptic connections between spiny neurons is only rarely observed. These results suggest that the bulk of the fast inhibition that has the strongest effects on spiny neuron spike timing comes from the feedforward interneuronal system whereas the axon collateral feedback system acts principally at the dendrites to control local excitability as well as the overall level of activity of the spiny neuron.

The coffee diterpene kahweol inhibits tumor necrosis factor-{alpha}-induced expression of cell adhesion molecules in human endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Hyung Gyun; Kim, Ji Young; Hwang, Yong Pil

2006-12-15

Endothelial cells produce adhesion molecules after being stimulated with various inflammatory cytokines. These adhesion molecules play an important role in the development of atherogenesis. Recent studies have highlighted the chemoprotective and anti-inflammatory effects of kahweol, a coffee-specific diterpene. This study examined the effects of kahweol on the cytokine-induced monocyte/human endothelial cell interaction, which is a crucial early event in atherogenesis. Kahweol inhibited the adhesion of TNF{alpha}-induced monocytes to endothelial cells and suppressed the TNF{alpha}-induced protein and mRNA expression of the cell adhesion molecules, VCAM-1 and ICAM-1. Furthermore, kahweol inhibited the TNF{alpha}-induced JAK2-PI3K/Akt-NF-{kappa}B activation pathway in these cells. Overall, kahweol hasmore » anti-inflammatory and anti-atherosclerotic activities, which occurs partly by down-regulating the pathway that affects the expression and interaction of the cell adhesion molecules on endothelial cells.« less

High-content adhesion assay to address limited cell samples†

PubMed Central

Warrick, Jay W.; Young, Edmond W. K.; Schmuck, Eric G.; Saupe, Kurt W.

2013-01-01

Cell adhesion is a broad topic in cell biology that involves physical interactions between cells and other cells or the surrounding extracellular matrix, and is implicated in major research areas including cancer, development, tissue engineering, and regenerative medicine. While current methods have contributed significantly to our understanding of cell adhesion, these methods are unsuitable for tackling many biological questions requiring intermediate numbers of cells (102–105), including small animal biopsies, clinical samples, and rare cell isolates. To overcome this fundamental limitation, we developed a new assay to quantify the adhesion of ~102–103 cells at a time on engineered substrates, and examined the adhesion strength and population heterogeneity via distribution-based modeling. We validated the platform by testing adhesion strength of cancer cells from three different cancer types (breast, prostate, and multiple myeloma) on both IL-1β activated and non-activated endothelial monolayers, and observed significantly increased adhesion for each cancer cell type upon endothelial activation, while identifying and quantifying distinct subpopulations of cell-substrate interactions. We then applied the assay to characterize adhesion of primary bone marrow stromal cells to different cardiac fibroblast-derived matrix substrates to demonstrate the ability to study limited cell populations in the context of cardiac cell-based therapies. Overall, these results demonstrate the sensitivity and robustness of the assay as well as its ability to enable extraction of high content, functional data from limited and potentially rare primary samples. We anticipate this method will enable a new class of biological studies with potential impact in basic and translational research. PMID:23426645

Dissecting the Impact of Matrix Anchorage and Elasticity in Cell Adhesion

PubMed Central

Pompe, Tilo; Glorius, Stefan; Bischoff, Thomas; Uhlmann, Ina; Kaufmann, Martin; Brenner, Sebastian; Werner, Carsten

2009-01-01

Abstract Extracellular matrices determine cellular fate decisions through the regulation of intracellular force and stress. Previous studies suggest that matrix stiffness and ligand anchorage cause distinct signaling effects. We show herein how defined noncovalent anchorage of adhesion ligands to elastic substrates allows for dissection of intracellular adhesion signaling pathways related to matrix stiffness and receptor forces. Quantitative analysis of the mechanical balance in cell adhesion using traction force microscopy revealed distinct scalings of the strain energy imparted by the cells on the substrates dependent either on matrix stiffness or on receptor force. Those scalings suggested the applicability of a linear elastic theoretical framework for the description of cell adhesion in a certain parameter range, which is cell-type-dependent. Besides the deconvolution of biophysical adhesion signaling, site-specific phosphorylation of focal adhesion kinase, dependent either on matrix stiffness or on receptor force, also demonstrated the dissection of biochemical signaling events in our approach. Moreover, the net contractile moment of the adherent cells and their strain energy exerted on the elastic substrate was found to be a robust measure of cell adhesion with a unifying power-law scaling exponent of 1.5 independent of matrix stiffness. PMID:19843448

Unified pre- and postsynaptic long-term plasticity enables reliable and flexible learning.

PubMed

Costa, Rui Ponte; Froemke, Robert C; Sjöström, P Jesper; van Rossum, Mark Cw

2015-08-26

Although it is well known that long-term synaptic plasticity can be expressed both pre- and postsynaptically, the functional consequences of this arrangement have remained elusive. We show that spike-timing-dependent plasticity with both pre- and postsynaptic expression develops receptive fields with reduced variability and improved discriminability compared to postsynaptic plasticity alone. These long-term modifications in receptive field statistics match recent sensory perception experiments. Moreover, learning with this form of plasticity leaves a hidden postsynaptic memory trace that enables fast relearning of previously stored information, providing a cellular substrate for memory savings. Our results reveal essential roles for presynaptic plasticity that are missed when only postsynaptic expression of long-term plasticity is considered, and suggest an experience-dependent distribution of pre- and postsynaptic strength changes.

Glutamatergic postsynaptic block by Pamphobeteus spider venoms in crayfish.

PubMed

Araque, A; Ferreira, W; Lucas, S; Buño, W

1992-01-31

The effects of toxins from venom glands of two south american spiders (Pamphobeteus platyomma and P. soracabae) on glutamatergic excitatory synaptic transmission were studied in the neuromuscular junction of the opener muscle of crayfish. The toxins selectively and reversibly blocked both excitatory postsynaptic currents and potentials in a dose-dependent manner. They also reversibly abolished glutamate-induced postsynaptic membrane depolarization. They had no effect on resting postsynaptic membrane conductance nor on postsynaptic voltage-gated currents. The synaptic facilitation and the frequency of miniature postsynaptic potentials were unaffected by the toxins, indicating that presynaptic events were not modified. Picrotoxin, a selective antagonist of the gamma-aminobutyric acid (GABA)A receptor, did not modify toxin effects. We conclude that both toxins specifically block the postsynaptic glutamate receptor-channel complex.

Occludin confers adhesiveness when expressed in fibroblasts.

PubMed

Van Itallie, C M; Anderson, J M

1997-05-01

Occludin is an integral membrane protein specifically associated with tight junctions. Previous studies suggest it is likely to function in forming the intercellular seal. In the present study, we expressed occludin under an inducible promotor in occludin-null fibroblasts to determine whether this protein confers intercellular adhesion. When human occludin is stably expressed in NRK and Rat-1 fibroblasts, which lack endogenous occludin and tight junctions but do have well developed ZO-1-containing adherens-like junctions, occludin colocalizes with ZO-1 to points of cell-cell contact. In contrast, L-cell fibroblasts which lack cadherin-based adherens junctions, target neither ZO-1 nor occludin to sites of cell contact. Occludin-induced adhesion was next quantified using a suspended cell assay. In NRK and Rat-1 cells, occludin expression induces adhesion in the absence of calcium, thus independent of cadherin-cadherin contacts. In contrast, L-cells are nonadhesive in this assay and show no increase in adhesion after induction of occludin expression. Binding of an antibody to the first of the putative extracellular loops of occludin confirmed that this sequence was exposed on the cell surface, and synthetic peptides containing the amino acid sequence of this loop inhibit adhesion induced by occludin expression. These results suggest that the extracellular surface of occludin is directly involved in cell-cell adhesion and the ability to confer adhesiveness correlates with the ability to colocalize with its cytoplasmic binding protein, ZO-1.

Inhibition of cell adhesion by anti–P-selectin aptamer: a new potential therapeutic agent for sickle cell disease

PubMed Central

Gutsaeva, Diana R.; Parkerson, James B.; Yerigenahally, Shobha D.; Kurz, Jeffrey C.; Schaub, Robert G.; Ikuta, Tohru

2011-01-01

Adhesive interactions between circulating sickle red blood cells (RBCs), leukocytes, and endothelial cells are major pathophysiologic events in sickle cell disease (SCD). To develop new therapeutics that efficiently inhibit adhesive interactions, we generated an anti–P-selectin aptamer and examined its effects on cell adhesion using knockout-transgenic SCD model mice. Aptamers, single-stranded oligonucleotides that bind molecular targets with high affinity and specificity, are emerging as new therapeutics for cardiovascular and hematologic disorders. In vitro studies found that the anti–P-selectin aptamer exhibits high specificity to mouse P-selectin but not other selectins. SCD mice were injected with the anti–P-selectin aptamer, and cell adhesion was observed under hypoxia. The anti–P-selectin aptamer inhibited the adhesion of sickle RBCs and leukocytes to endothelial cells by 90% and 80%, respectively. The anti–P-selectin aptamer also increased microvascular flow velocities and reduced the leukocyte rolling flux. SCD mice treated with the anti–P-selectin aptamer demonstrated a reduced mortality rate associated with the experimental procedures compared with control mice. These results demonstrate that anti–P-selectin aptamer efficiently inhibits the adhesion of both sickle RBCs and leukocytes to endothelial cells in SCD model mice, suggesting a critical role for P-selectin in cell adhesion. Anti–P-selectin aptamer may be useful as a novel therapeutic agent for SCD. PMID:20926770

Platelet-independent adhesion of calcium-loaded erythrocytes to von Willebrand factor

PubMed Central

Bierings, Ruben; Meems, Henriet; Mul, Frederik P. J.; Geerts, Dirk; Vlaar, Alexander P. J.; Voorberg, Jan; Hordijk, Peter L.

2017-01-01

Adhesion of erythrocytes to endothelial cells lining the vascular wall can cause vaso-occlusive events that impair blood flow which in turn may result in ischemia and tissue damage. Adhesion of erythrocytes to vascular endothelial cells has been described in multiple hemolytic disorders, especially in sickle cell disease, but the adhesion of normal erythrocytes to endothelial cells has hardly been described. It was shown that calcium-loaded erythrocytes can adhere to endothelial cells. Because sickle erythrocyte adhesion to ECs can be enhanced by ultra-large von Willebrand factor multimers, we investigated whether calcium loading of erythrocytes could promote binding to endothelial cells via ultra-large von Willebrand factor multimers. We used (immunofluorescent) live-cell imaging of washed erythrocytes perfused over primary endothelial cells at venular flow rate. Using this approach, we show that calcium-loaded erythrocytes strongly adhere to histamine-stimulated primary human endothelial cells. This adhesion is mediated by ultra-large von Willebrand factor multimers. Von Willebrand factor knockdown or ADAMTS13 cleavage abolished the binding of erythrocytes to activated endothelial cells under flow. Platelet depletion did not interfere with erythrocyte binding to von Willebrand factor. Our results reveal platelet-independent adhesion of calcium-loaded erythrocytes to endothelium-derived von Willebrand factor. Erythrocyte adhesion to von Willebrand factor may be particularly relevant for venous thrombosis, which is characterized by the formation of erythrocyte-rich thrombi. PMID:28249049

Revealing Early Steps of α2β1 Integrin-mediated Adhesion to Collagen Type I by Using Single-Cell Force Spectroscopy

PubMed Central

Taubenberger, Anna; Cisneros, David A.; Friedrichs, Jens; Puech, Pierre-Henri; Muller, Daniel J.

2007-01-01

We have characterized early steps of α2β1 integrin-mediated cell adhesion to a collagen type I matrix by using single-cell force spectroscopy. In agreement with the role of α2β1 as a collagen type I receptor, α2β1-expressing Chinese hamster ovary (CHO)-A2 cells spread rapidly on the matrix, whereas α2β1-negative CHO wild-type cells adhered poorly. Probing CHO-A2 cell detachment forces over a contact time range of 600 s revealed a nonlinear adhesion response. During the first 60 s, cell adhesion increased slowly, and forces associated with the smallest rupture events were consistent with the breakage of individual integrin–collagen bonds. Above 60 s, a fraction of cells rapidly switched into an activated adhesion state marked by up to 10-fold increased detachment forces. Elevated overall cell adhesion coincided with a rise of the smallest rupture forces above the value required to break a single-integrin–collagen bond, suggesting a change from single to cooperative receptor binding. Transition into the activated adhesion mode and the increase of the smallest rupture forces were both blocked by inhibitors of actomyosin contractility. We therefore propose a two-step mechanism for the establishment of α2β1-mediated adhesion as weak initial, single-integrin–mediated binding events are superseded by strong adhesive interactions involving receptor cooperativity and actomyosin contractility. PMID:17314408

RECURRENT NEONATAL SEIZURES RESULT IN LONG-TERM INCREASE OF NEURONAL NETWORK EXCITABILITY IN THE RAT NEOCORTEX

PubMed Central

Isaeva, Elena; Isaev, Dmytro; Savrasova, Alina; Khazipov, Rustem; Holmes, Gregory L.

2011-01-01

Neonatal seizures are associated with a high likelihood of adverse neurological outcomes, including mental retardation, behavioral disorders, and epilepsy. Early seizures typically involve the neocortex, and post-neonatal epilepsy is often of neocortical origin. However, our understanding of the consequences of neonatal seizures for neocortical function is limited. In the present study, we show that neonatal seizures induced by flurothyl result in markedly enhanced susceptibility of the neocortex to seizure-like activity. This change occurs in young rats studied weeks after the last induced seizure and in adult rats studied months after the initial seizures. Neonatal seizures resulted in reductions in the amplitude of spontaneous inhibitory postsynaptic currents and the frequency of miniature inhibitory postsynaptic currents, and significant increases in the amplitude and frequency of spontaneous excitatory postsynaptic currents (sEPSCs) and in the frequency of miniature excitatory postsynaptic currents (mEPSCs) in pyramidal cells of layer 2/3 of the somatosensory cortex. The selective N-methyl-d-aspartate (NMDA) receptor antagonist d-2-amino-5-phosphon-ovalerate eliminated the differences in amplitude and frequency of sEPSCs and mEPSCs in the control and flurothyl groups, suggesting that NMDA receptors contribute significantly to the enhanced excitability seen in slices from rats that experienced recurrent neonatal seizures. Taken together, our results suggest that recurrent seizures in infancy result in a persistent enhancement of neocortical excitability. PMID:20384780

Tuning cell adhesion on polymeric and nanocomposite surfaces: Role of topography versus superhydrophobicity.

PubMed

Zangi, Sepideh; Hejazi, Iman; Seyfi, Javad; Hejazi, Ehsan; Khonakdar, Hossein Ali; Davachi, Seyed Mohammad

2016-06-01

Development of surface modification procedures which allow tuning the cell adhesion on the surface of biomaterials and devices is of great importance. In this study, the effects of different topographies and wettabilities on cell adhesion behavior of polymeric surfaces are investigated. To this end, an improved phase separation method was proposed to impart various wettabilities (hydrophobic and superhydrophobic) on polypropylene surfaces. Surface morphologies and compositions were characterized by scanning electron microscopy and X-ray photoelectron spectroscopy, respectively. Cell culture was conducted to evaluate the adhesion of 4T1 mouse mammary tumor cells. It was found that processing conditions such as drying temperature is highly influential in cell adhesion behavior due to the formation of an utterly different surface topography. It was concluded that surface topography plays a more significant role in cell adhesion behavior rather than superhydrophobicity since the nano-scale topography highly inhibited the cell adhesion as compared to the micro-scale topography. Such cell repellent behavior could be very useful in many biomedical devices such as those in drug delivery and blood contacting applications as well as biosensors. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid and Localized Mechanical Stimulation and Adhesion Assay: TRPM7 Involvement in Calcium Signaling and Cell Adhesion

PubMed Central

Nishitani, Wagner Shin; Alencar, Adriano Mesquita; Wang, Yingxiao

2015-01-01

A cell mechanical stimulation equipment, based on cell substrate deformation, and a more sensitive method for measuring adhesion of cells were developed. A probe, precisely positioned close to the cell, was capable of a vertical localized mechanical stimulation with a temporal frequency of 207 Hz, and strain magnitude of 50%. This setup was characterized and used to probe the response of Human Umbilical Endothelial Vein Cells (HUVECs) in terms of calcium signaling. The intracellular calcium ion concentration was measured by the genetically encoded Cameleon biosensor, with the Transient Receptor Potential cation channel, subfamily M, member 7 (TRPM7) expression inhibited. As TRPM7 expression also regulates adhesion, a relatively simple method for measuring adhesion of cells was also developed, tested and used to study the effect of adhesion alone. Three adhesion conditions of HUVECs on polyacrylamide gel dishes were compared. In the first condition, the substrate is fully treated with Sulfo-SANPAH crosslinking and fibronectin. The other two conditions had increasingly reduced adhesion: partially treated (only coated with fibronectin, with no use of Sulfo-SANPAH, at 5% of the normal amount) and non-treated polyacrylamide gels. The cells showed adhesion and calcium response to the mechanical stimulation correlated to the degree of gel treatment: highest for fully treated gels and lowest for non-treated ones. TRPM7 inhibition by siRNA on HUVECs caused an increase in adhesion relative to control (no siRNA treatment) and non-targeting siRNA, but a decrease to 80% of calcium response relative to non-targeting siRNA which confirms the important role of TRPM7 in mechanotransduction despite the increase in adhesion. PMID:25946314

Structure activity relationship of synaptic and junctional neurotransmission.

PubMed

Goyal, Raj K; Chaudhury, Arun

2013-06-01

Chemical neurotransmission may include transmission to local or remote sites. Locally, contact between 'bare' portions of the bulbous nerve terminal termed a varicosity and the effector cell may be in the form of either synapse or non-synaptic contact. Traditionally, all local transmissions between nerves and effector cells are considered synaptic in nature. This is particularly true for communication between neurons. However, communication between nerves and other effectors such as smooth muscles has been described as nonsynaptic or junctional in nature. Nonsynaptic neurotransmission is now also increasingly recognized in the CNS. This review focuses on the relationship between structure and function that orchestrate synaptic and junctional neurotransmissions. A synapse is a specialized focal contact between the presynaptic active zone capable of ultrafast release of soluble transmitters and the postsynaptic density that cluster ionotropic receptors. The presynaptic and the postsynaptic areas are separated by the 'closed' synaptic cavity. The physiological hallmark of the synapse is ultrafast postsynaptic potentials lasting milliseconds. In contrast, junctions are juxtapositions of nerve terminals and the effector cells without clear synaptic specializations and the junctional space is 'open' to the extracellular space. Based on the nature of the transmitters, postjunctional receptors and their separation from the release sites, the junctions can be divided into 'close' and 'wide' junctions. Functionally, the 'close' and the 'wide' junctions can be distinguished by postjunctional potentials lasting ~1s and tens of seconds, respectively. Both synaptic and junctional communications are common between neurons; however, junctional transmission is the rule at many neuro-non-neural effectors. Published by Elsevier B.V.

Structure activity relationship of synaptic and junctional neurotransmission

PubMed Central

Goyal, Raj K; Chaudhury, Arun

2013-01-01

Chemical neurotransmission may include transmission to local or remote sites. Locally, contact between ‘bare’ portions of the bulbous nerve terminal termed a varicosity and the effector cell may be in the form of either synapse or non-synaptic contact. Traditionally, all local transmissions between nerves and effector cells are considered synaptic in nature. This is particularly true for communication between neurons. However, communication between nerves and other effectors such as smooth muscles has been described as nonsynaptic or junctional in nature. Nonsynaptic neurotransmission is now also increasing recognized in the CNS. This review focuses on the relationship between structure and function that orchestrate synaptic and junctional neurotransmissions. A synapse is a specialized focal contact between the presynaptic active zone capable for ultrafast release of soluble transmitters and the postsynaptic density that cluster ionotropic receptors. The presynaptic and the postsynaptic areas are separated by the ‘closed’ synaptic cavity. The physiological hallmark of the synapse is ultrafast postsynaptic potentials lasting in milliseconds. In contrast, junctions are juxtapositions of nerve terminals and the effector cells without clear synaptic specializations and the junctional space is ‘open’ to the extracellular space. Based on the nature of the transmitters, postjunctional receptors and their separation from the release sites, the junctions can be divided into ‘close’ and ‘wide’ junctions. Functionally, the ‘close’ and the ‘wide’ junctions can be distinguished by postjunctional potentials lasting ~1 second and 10s of seconds, respectively. Both synaptic and junctional communications are common between neurons; however, junctional transmission is the rule at many neuro-non-neural effectors. PMID:23535140

Iron ion irradiation increases promotes adhesion of monocytic cells to arterial vascular endothelium

NASA Astrophysics Data System (ADS)

Kucik, Dennis; Khaled, Saman; Gupta, Kiran; Wu, Xing; Yu, Tao; Chang, Polly; Kabarowski, Janusz

Radiation causes inflammation, and chronic, low-level vascular inflammation is a risk factor for atherosclerosis. Consistent with this, exposure to radiation from a variety of sources is associated with increased risk of heart disease and stroke. Part of the inflammatory response to radiation is a change in the adhesiveness of the endothelial cells that line the blood vessels, triggering inappropriate accumulation of leukocytes, leading to later, damaging effects of inflammation. Although some studies have been done on the effects of gamma irradiation on vascular endothelium, the response of endothelium to heavy ion radiation likely to be encountered in prolonged space flight has not been determined. We investigated how irradiation of aortic endothelial cells with iron ions affects adhesiveness of cultured aortic endothelial cells for monocytic cells and the consequences of this for development of atherosclerosis. Aortic endothelial cells were irradiated with 600 MeV iron ions at Brookhaven National Laboratory and adhesion-related changes were measured. Cells remained viable for at least 72 hours, and were even able to repair acute damage to cell junctions. We found that iron ion irradiation altered expression levels of specific endothelial cell adhesion molecules. Further, these changes had functional consequences. Using a flow chamber adhesion assay to measure adhesion of monocytic cells to endothelial cells under physiological shear stress, we found that adhesivity of vascular endothelium was enhanced in as little as 24 hours after irradiation. Further, the radiation dose dependence was not monotonic, suggesting that it was not simply the result of endothelial cell damage. We also irradiated aortic arches and carotid arteries of Apolipoprotein-E-deficient mice. Histologic analysis of these mice will be conducted to determine whether effects of radiation on endothelial adhesiveness result in consequences for development of atherosclerosis. (Supported by NSBRI: NCC-9-58-162)

[Effect of Golgi α-mannosidase 2 (GM2) gene knockdown on adhesion abilities of human gastric carcinoma cell line BGC-823 and its mechanism].

PubMed

Zeng, Bo; Zeng, Zhen; Liu, Chang; Yang, Yaying

2017-06-01

Objective To investigate the effect of Golgi α-mannosidase II (GM2) gene knockdown on adhesion abilities of BGC-823 human gastric carcinoma cells. Methods Three plasmid vectors expressing GM2 shRNAs and a negative control plasmid vector were designed, constructed and then transfected into BGC-823 cells by Lipofectamine TM 2000. After transfection, the mRNA and protein levels of GM2 in BGC-823 cells were detected by real-time quantitative PCR (qRT-PCR) and Western blotting to evaluate the transfection efficacy. The best plasmid for GM2 gene knockdown was selected and stably transfected into BGC-823 cells. Adhesion abilities of BGC-823 cells after GM2 gene silencing were observed by cell-cell, cell-matrix and cell-endothelial cell adhesion assays. At the same time, the expressions of E-cadherin, P-selectin, CD44v6 and intercellular adhesion molecule-1 (ICAM-1) proteins were detected by Western blotting after GM2 gene knockdown. Results The expression of GM2 was effectively knockdown in GM2-shRNA-2-transfected BGC-823 cells. Compared with the blank control group and the negative control group, the intercellular adhesion ability of the GM2-shRNA-2-transfected cells increased significantly, while their cell-matrix and cell-endothelium adhesion abilities markedly decreased. In GM2-shRNA-2 transfection group, E-cadherin expression was significantly elevated and the P-selectin expression was significantly reduced, while the expression levels of CD44v6 and ICAM-1 were not obviously changed. Conclusion After GM2 gene knockdown, the intercellular adhesion ability of gastric carcinoma BGC-823 cells is enhanced, while the adhesion abilities with the extracellular matrix and endothelial cells are weakened. The changes might be related to the up-regulated expression of E-cadherin and the down-regulation of P-selectin.

Surfactant functionalization induces robust, differential adhesion of tumor cells and blood cells to charged nanotube-coated biomaterials under flow.

PubMed

Mitchell, Michael J; Castellanos, Carlos A; King, Michael R

2015-07-01

The metastatic spread of cancer cells from the primary tumor to distant sites leads to a poor prognosis in cancers originating from multiple organs. Increasing evidence has linked selectin-based adhesion between circulating tumor cells (CTCs) and endothelial cells of the microvasculature to metastatic dissemination, in a manner similar to leukocyte adhesion during inflammation. Functionalized biomaterial surfaces hold promise as a diagnostic tool to separate CTCs and potentially treat metastasis, utilizing antibody and selectin-mediated interactions for cell capture under flow. However, capture at high purity levels is challenged by the fact that CTCs and leukocytes both possess selectin ligands. Here, a straightforward technique to functionalize and alter the charge of naturally occurring halloysite nanotubes using surfactants is reported to induce robust, differential adhesion of tumor cells and blood cells to nanotube-coated surfaces under flow. Negatively charged sodium dodecanoate-functionalized nanotubes simultaneously enhanced tumor cell capture while negating leukocyte adhesion, both in the presence and absence of adhesion proteins, and can be utilized to isolate circulating tumor cells regardless of biomarker expression. Conversely, diminishing nanotube charge via functionalization with decyltrimethylammonium bromide both abolished tumor cell capture while promoting leukocyte adhesion. Copyright © 2015 Elsevier Ltd. All rights reserved.

Characterizing phenolformaldehyde adhesive cure chemistry within the wood cell wall

Treesearch

Daniel J. Yelle; John Ralph

2016-01-01

Adhesive bonding of wood using phenol-formaldehyde remains the industrial standard in wood product bond durability. Not only does this adhesive infiltrate the cell wall, it also is believed to form primary bonds with wood cell wall polymers, particularly guaiacyl lignin. However, the mechanism by which phenol-formaldehyde adhesive intergrally interacts and bonds to...

Evaluation of adhesive penetration of wood fibre by nanoindentation and microscopy

Treesearch

Christopher G. Hunt; Joseph E. Jakes; Warren Grigsby

2010-01-01

Adhesives used in wood products sometimes infiltrate, or diffuse into the solid material of, wood cell walls, potentially modifying their properties. These changes in cell wall properties are likely to impact the performance of adhesive bonds. While adhesive infiltration has been observed by multiple methods, the effect on cell wall properties is not well understood....

Single-cell force spectroscopy of pili-mediated adhesion

NASA Astrophysics Data System (ADS)

Sullan, Ruby May A.; Beaussart, Audrey; Tripathi, Prachi; Derclaye, Sylvie; El-Kirat-Chatel, Sofiane; Li, James K.; Schneider, Yves-Jacques; Vanderleyden, Jos; Lebeer, Sarah; Dufrêne, Yves F.

2013-12-01

Although bacterial pili are known to mediate cell adhesion to a variety of substrates, the molecular interactions behind this process are poorly understood. We report the direct measurement of the forces guiding pili-mediated adhesion, focusing on the medically important probiotic bacterium Lactobacillus rhamnosus GG (LGG). Using non-invasive single-cell force spectroscopy (SCFS), we quantify the adhesion forces between individual bacteria and biotic (mucin, intestinal cells) or abiotic (hydrophobic monolayers) surfaces. On hydrophobic surfaces, bacterial pili strengthen adhesion through remarkable nanospring properties, which - presumably - enable the bacteria to resist high shear forces under physiological conditions. On mucin, nanosprings are more frequent and adhesion forces larger, reflecting the influence of specific pili-mucin bonds. Interestingly, these mechanical responses are no longer observed on human intestinal Caco-2 cells. Rather, force curves exhibit constant force plateaus with extended ruptures reflecting the extraction of membrane nanotethers. These single-cell analyses provide novel insights into the molecular mechanisms by which piliated bacteria colonize surfaces (nanosprings, nanotethers), and offer exciting avenues in nanomedicine for understanding and controlling the adhesion of microbial cells (probiotics, pathogens).

Adhesion of Epstein–Barr virus-positive natural killer cell lines to cultured endothelial cells stimulated with inflammatory cytokines

PubMed Central

Kanno, H; Watabe, D; Shimizu, N; Sawai, T

2008-01-01

Chronic active Epstein–Barr virus (EBV) infection (CAEBV) is characterized by chronic recurrent infectious mononucleosis-like symptoms. Approximately one-fourth of CAEBV patients develop vascular lesions with infiltration of EBV-positive lymphoid cells. Furthermore, EBV-positive natural killer (NK)/T cell lymphomas often exhibit angiocentric or angiodestructive lesions. These suggest an affinity of EBV-positive NK/T cells to vascular components. In this study, we evaluated the expression of adhesion molecules and cytokines in EBV-positive NK lymphoma cell lines, SNK1 and SNK6, and examined the role of cytokines in the interaction between NK cell lines and endothelial cells. SNKs expressed intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) at much higher levels than those in EBV-negative T cell lines. SNKs produced the larger amount of tumour necrosis factor (TNF)-α, which caused increased expression of ICAM-1 and VCAM-1 in cultured human endothelial cells, than that from EBV-negative T cell lines. Furthermore, SNKs exhibited increased adhesion to cultured endothelial cells stimulated with TNF-α or interleukin (IL)-1β, and the pretreatment of cytokine-stimulated endothelial cells with anti-VCAM-1-antibodies reduced cell adhesion. These indicate that the up-regulated expression of VCAM-1 on cytokine-stimulated endothelial cells would be important for the adhesion of EBV-positive NK cells and might initiate the vascular lesions. PMID:18190605

MiR-126 and miR-126* regulate shear-resistant firm leukocyte adhesion to human brain endothelium

PubMed Central

Cerutti, Camilla; Edwards, Laura J.; de Vries, Helga E.; Sharrack, Basil; Male, David K.; Romero, Ignacio A.

2017-01-01

Leukocyte adhesion to brain endothelial cells, the blood-brain barrier main component, is a critical step in the pathogenesis of neuroinflammatory diseases such as multiple sclerosis (MS). Leukocyte adhesion is mediated mainly by selectins, cell adhesion molecules and chemokines induced by pro-inflammatory cytokines such as TNFα and IFNγ, but the regulation of this process is not fully clear. This study investigated the regulation of firm leukocyte adhesion to human brain endothelium by two different brain endothelial microRNAs (miRs), miR-126 and miR-126*, that are downregulated by TNFα and IFNγ in a human brain endothelial cell line, hCMEC/D3. Using a leukocyte adhesion in vitro assay under shear forces mimicking blood flow, we observed that reduction of endothelial miR-126 and miR-126* enhanced firm monocyte and T cell adhesion to hCMEC/D3 cells, whereas their increased expression partially prevented THP1, Jurkat and primary MS patient-derived PBMC firm adhesion. Furthermore, we observed that miR-126* and miR-126 downregulation increased E-selectin and VCAM1, respectively, while miR-126 overexpression reduced VCAM1 and CCL2 expression by hCMEC/D3 cells, suggesting that these miRs regulate leukocyte adhesion by modulating the expression of adhesion-associated endothelial mRNA targets. Hence, human brain endothelial miR-126 and miR-126* could be used as a therapeutic tool to reduce leukocyte adhesion and thus reduce neuroinflammation. PMID:28358058

Epigallocatechin 3-gallate inhibits 7-ketocholesterol-induced monocyte-endothelial cell adhesion.

PubMed

Yamagata, Kazuo; Tanaka, Noriko; Suzuki, Koichi

2013-07-01

7-Ketocholesterol (7KC) induces monocytic adhesion to endothelial cells, and induces arteriosclerosis while high-density lipoprotein (HDL) inhibits monocytic adhesion to the endothelium. Epigallocatechin 3-gallate (EGCG) was found to have a protective effect against arteriosclerosis. Therefore, the purpose of this study was to examine the possible HDL-like mechanisms of EGCG in endothelial cells by investigating whether EGCG inhibits 7KC-induced monocyte-endothelial cell adhesion by activating HDL-dependent signal transduction pathways. 7KC and/or EGCG were added to human endothelial cells (ISO-HAS), and the adhesion of pro-monocytic U937 cells was examined. The expression of genes associated with HDL effects such as Ca(2+)/calmodulin-dependent kinase II (CaMKKII), liver kinase B (LKD1), PSD-95/Dlg/ZO-1 kinase 1 (PDZK1), phosphatidylinositol 3-kinase (PI3K), intercellular adhesion molecule-1 (ICAM-1), monocyte chemotactic protein-1 (MCP-1), and endothelial nitric oxide synthase (eNOS) was examined by RT-PCR, and ICAM-1 protein expression was evaluated by western blot (WB). Production of reactive oxygen species (ROS) was examined with H2DCFDA. 7KC significantly induced adhesion of U937 cells to human endothelial cells while significantly increasing gene expressions of ICAM-1 and MCP-1 and decreasing eNOS and CaMKKII gene expressions. EGCG inhibited 7KC-induced monocytic adhesion to endothelial cells, and induced expression of eNOS and several genes involved in the CaMKKII pathway. Stimulation of endothelial cells with EGCG produced intracellular ROS, whereas treatment with N-acetylcysteine (NAC) blocked EGCG-induced expression of eNOS and CaMKKII. These results suggest that inhibition of monocyte-endothelial cell adhesion by EGCG is associated with CaMKKII pathway activation by ROS. Inhibition of 7KC-induced monocyte-endothelial cell adhesion induced by EGCG may function similarly to HDL. Copyright © 2013 Elsevier Inc. All rights reserved.

Single cell adhesion assay using computer controlled micropipette.

PubMed

Salánki, Rita; Hős, Csaba; Orgovan, Norbert; Péter, Beatrix; Sándor, Noémi; Bajtay, Zsuzsa; Erdei, Anna; Horvath, Robert; Szabó, Bálint

2014-01-01

Cell adhesion is a fundamental phenomenon vital for all multicellular organisms. Recognition of and adhesion to specific macromolecules is a crucial task of leukocytes to initiate the immune response. To gain statistically reliable information of cell adhesion, large numbers of cells should be measured. However, direct measurement of the adhesion force of single cells is still challenging and today's techniques typically have an extremely low throughput (5-10 cells per day). Here, we introduce a computer controlled micropipette mounted onto a normal inverted microscope for probing single cell interactions with specific macromolecules. We calculated the estimated hydrodynamic lifting force acting on target cells by the numerical simulation of the flow at the micropipette tip. The adhesion force of surface attached cells could be accurately probed by repeating the pick-up process with increasing vacuum applied in the pipette positioned above the cell under investigation. Using the introduced methodology hundreds of cells adhered to specific macromolecules were measured one by one in a relatively short period of time (∼30 min). We blocked nonspecific cell adhesion by the protein non-adhesive PLL-g-PEG polymer. We found that human primary monocytes are less adherent to fibrinogen than their in vitro differentiated descendants: macrophages and dendritic cells, the latter producing the highest average adhesion force. Validation of the here introduced method was achieved by the hydrostatic step-pressure micropipette manipulation technique. Additionally the result was reinforced in standard microfluidic shear stress channels. Nevertheless, automated micropipette gave higher sensitivity and less side-effect than the shear stress channel. Using our technique, the probed single cells can be easily picked up and further investigated by other techniques; a definite advantage of the computer controlled micropipette. Our experiments revealed the existence of a sub-population of strongly fibrinogen adherent cells appearing in macrophages and highly represented in dendritic cells, but not observed in monocytes.

Single Cell Adhesion Assay Using Computer Controlled Micropipette

PubMed Central

Salánki, Rita; Hős, Csaba; Orgovan, Norbert; Péter, Beatrix; Sándor, Noémi; Bajtay, Zsuzsa; Erdei, Anna; Horvath, Robert; Szabó, Bálint

2014-01-01

Cell adhesion is a fundamental phenomenon vital for all multicellular organisms. Recognition of and adhesion to specific macromolecules is a crucial task of leukocytes to initiate the immune response. To gain statistically reliable information of cell adhesion, large numbers of cells should be measured. However, direct measurement of the adhesion force of single cells is still challenging and today’s techniques typically have an extremely low throughput (5–10 cells per day). Here, we introduce a computer controlled micropipette mounted onto a normal inverted microscope for probing single cell interactions with specific macromolecules. We calculated the estimated hydrodynamic lifting force acting on target cells by the numerical simulation of the flow at the micropipette tip. The adhesion force of surface attached cells could be accurately probed by repeating the pick-up process with increasing vacuum applied in the pipette positioned above the cell under investigation. Using the introduced methodology hundreds of cells adhered to specific macromolecules were measured one by one in a relatively short period of time (∼30 min). We blocked nonspecific cell adhesion by the protein non-adhesive PLL-g-PEG polymer. We found that human primary monocytes are less adherent to fibrinogen than their in vitro differentiated descendants: macrophages and dendritic cells, the latter producing the highest average adhesion force. Validation of the here introduced method was achieved by the hydrostatic step-pressure micropipette manipulation technique. Additionally the result was reinforced in standard microfluidic shear stress channels. Nevertheless, automated micropipette gave higher sensitivity and less side-effect than the shear stress channel. Using our technique, the probed single cells can be easily picked up and further investigated by other techniques; a definite advantage of the computer controlled micropipette. Our experiments revealed the existence of a sub-population of strongly fibrinogen adherent cells appearing in macrophages and highly represented in dendritic cells, but not observed in monocytes. PMID:25343359

Comparative Study of Human and Mouse Postsynaptic Proteomes Finds High Compositional Conservation and Abundance Differences for Key Synaptic Proteins

PubMed Central

Bayés, Àlex; Collins, Mark O.; Croning, Mike D. R.; van de Lagemaat, Louie N.; Choudhary, Jyoti S.; Grant, Seth G. N.

2012-01-01

Direct comparison of protein components from human and mouse excitatory synapses is important for determining the suitability of mice as models of human brain disease and to understand the evolution of the mammalian brain. The postsynaptic density is a highly complex set of proteins organized into molecular networks that play a central role in behavior and disease. We report the first direct comparison of the proteome of triplicate isolates of mouse and human cortical postsynaptic densities. The mouse postsynaptic density comprised 1556 proteins and the human one 1461. A large compositional overlap was observed; more than 70% of human postsynaptic density proteins were also observed in the mouse postsynaptic density. Quantitative analysis of postsynaptic density components in both species indicates a broadly similar profile of abundance but also shows that there is higher abundance variation between species than within species. Well known components of this synaptic structure are generally more abundant in the mouse postsynaptic density. Significant inter-species abundance differences exist in some families of key postsynaptic density proteins including glutamatergic neurotransmitter receptors and adaptor proteins. Furthermore, we have identified a closely interacting set of molecules enriched in the human postsynaptic density that could be involved in dendrite and spine structural plasticity. Understanding synapse proteome diversity within and between species will be important to further our understanding of brain complexity and disease. PMID:23071613

Cell-matrix adhesion characterization using multiple shear stress zones in single stepwise microchannel

NASA Astrophysics Data System (ADS)

Kim, Min-Ji; Doh, Il; Bae, Gab-Yong; Cha, Hyuk-Jin; Cho, Young-Ho

2014-08-01

This paper presents a cell chip capable to characterize cell-matrix adhesion by monitoring cell detachment rate. The proposed cell chip can supply multiple levels of shear stress in single stepwise microchannel. As epithelial-mesenchymal transition (EMT), one of hallmarks of cancer metastasis is closely associated to the interaction with extracelluar matrix (ECM), we took advantage of two lung cancer cell models with different adhesion properties to ECM depending their epithelial or mesenchymal properties, including the pair of lung cancer cells with (A549sh) or without E-cadherin expression (A549sh-Ecad), which would be optimal model to examine the alteration of adhesion properties after EMT induction. The cell-matrix adhesion resisting to shear stress appeared to be remarkably differed between lung cancer cells. The detachment rate of epithelial-like H358 and mesenchymal-like H460 cells was 53%-80% and 25%-66% in the shear stress range of 34-60 dyn/cm2, respectively. A549sh-Ecad cells exhibits lower detachment rate (5%-9%) compared to A549sh cells (14%-40%). By direct comparison of adhesion between A549sh and A549sh-Ecad, we demonstrated that A549shE-cad to mimic EMT were more favorable to the ECM attachment under the various levels of shear stress. The present method can be applied to quantitative analysis of tumor cell-ECM adhesion.

Differential Expression of Adhesion-Related Proteins and MAPK Pathways Lead to Suitable Osteoblast Differentiation of Human Mesenchymal Stem Cells Subpopulations.

PubMed

Leyva-Leyva, Margarita; López-Díaz, Annia; Barrera, Lourdes; Camacho-Morales, Alberto; Hernandez-Aguilar, Felipe; Carrillo-Casas, Erika M; Arriaga-Pizano, Lourdes; Calderón-Pérez, Jaime; García-Álvarez, Jorge; Orozco-Hoyuela, Gabriel; Piña-Barba, Cristina; Rojas-Martínez, Augusto; Romero-Díaz, Víktor; Lara-Arias, Jorge; Rivera-Bolaños, Nancy; López-Camarillo, César; Moncada-Saucedo, Nidia; Galván-De los Santos, Alejandra; Meza-Urzúa, Fátima; Villarreal-Gómez, Luis; Fuentes-Mera, Lizeth

2015-11-01

Cellular adhesion enables communication between cells and their environment. Adhesion can be achieved throughout focal adhesions and its components influence osteoblast differentiation of human mesenchymal stem cells (hMSCs). Because cell adhesion and osteoblast differentiation are closely related, this article aimed to analyze the expression profiles of adhesion-related proteins during osteoblastic differentiation of two hMSCs subpopulations (CD105(+) and CD105(-)) and propose a strategy for assembling bone grafts based on its adhesion ability. In vitro experiments of osteogenic differentiation in CD105(-) cells showed superior adhesion efficiency and 2-fold increase of α-actinin expression compared with CD105(+) cells at the maturation stage. Interestingly, levels of activated β1-integrin increased in CD105(-) cells during the process. Additionally, the CD105(-) subpopulation showed 3-fold increase of phosphorylated FAK(Y397) compared to CD105(+) cells. Results also indicate that ERK1/2 was activated during CD105(-) bone differentiation and participation of mitogen-activated protein kinase (MAPK)-p38 in CD105(+) differentiation through a focal adhesion kinase (FAK)-independent pathway. In vivo trial demonstrated that grafts containing CD105(-) showed osteocytes embedded in a mineralized matrix, promoted adequate graft integration, increased host vascular infiltration, and efficient intramembranous repairing. In contrast, grafts containing CD105(+) showed deficient endochondral ossification and fibrocartilaginous tissue. Based on the expression of α-actinin, FAKy,(397) and ERK1/2 activation, we define maturation stage as critical for bone graft assembling. By in vitro assays, CD105(-) subpopulation showed superior adhesion efficiency compared to CD105(+) cells. Considering in vitro and in vivo assays, this study suggests that integration of a scaffold with CD105(-) subpopulation at the maturation stage represents an attractive strategy for clinical use in orthopedic bioengineering.

Wavelet Imaging on Multiple Scales (WIMS) reveals focal adhesion distributions, dynamics and coupling between actomyosin bundle stability

PubMed Central

Toplak, Tim; Palmieri, Benoit; Juanes-García, Alba; Vicente-Manzanares, Miguel; Grant, Martin; Wiseman, Paul W.

2017-01-01

We introduce and use Wavelet Imaging on Multiple Scales (WIMS) as an improvement to fluorescence correlation spectroscopy to measure physical processes and features that occur across multiple length scales. In this study, wavelet transforms of cell images are used to characterize molecular dynamics at the cellular and subcellular levels (i.e. focal adhesions). We show the usefulness of the technique by applying WIMS to an image time series of a migrating osteosarcoma cell expressing fluorescently labelled adhesion proteins, which allows us to characterize different components of the cell ranging from optical resolution scale through to focal adhesion and whole cell size scales. Using WIMS we measured focal adhesion numbers, orientation and cell boundary velocities for retraction and protrusion. We also determine the internal dynamics of individual focal adhesions undergoing assembly, disassembly or elongation. Thus confirming as previously shown, WIMS reveals that the number of adhesions and the area of the protruding region of the cell are strongly correlated, establishing a correlation between protrusion size and adhesion dynamics. We also apply this technique to characterize the behavior of adhesions, actin and myosin in Chinese hamster ovary cells expressing a mutant form of myosin IIB (1935D) that displays decreased filament stability and impairs front-back cell polarity. We find separate populations of actin and myosin at each adhesion pole for both the mutant and wild type form. However, we find these populations move rapidly inwards toward one another in the mutant case in contrast to the cells that express wild type myosin IIB where those populations remain stationary. Results obtained with these two systems demonstrate how WIMS has the potential to reveal novel correlations between chosen parameters that belong to different scales. PMID:29049414

Involvement of lipid rafts in adhesion-induced activation of Met and EGFR.

PubMed

Lu, Ying-Che; Chen, Hong-Chen

2011-10-27

Cell adhesion has been shown to induce activation of certain growth factor receptors in a ligand-independent manner. However, the mechanism for such activation remains obscure. Human epidermal carcinoma A431 cells were used as a model to examine the mechanism for adhesion-induced activation of hepatocyte growth factor receptor Met and epidermal growth factor receptor (EGFR). The cells were suspended and replated on culture dishes under various conditions. The phosphorylation of Met at Y1234/1235 and EGFR at Y1173 were used as indicators for their activation. The distribution of the receptors and lipid rafts on the plasma membrane were visualized by confocal fluorescent microscopy and total internal reflection microscopy. We demonstrate that Met and EGFR are constitutively activated in A431 cells, which confers proliferative and invasive potentials to the cells. The ligand-independent activation of Met and EGFR in A431 cells relies on cell adhesion to a substratum, but is independent of cell spreading, extracellular matrix proteins, and substratum stiffness. This adhesion-induced activation of Met and EGFR cannot be attributed to Src activation, production of reactive oxygen species, and the integrity of the cytoskeleton. In addition, we demonstrate that Met and EGFR are independently activated upon cell adhesion. However, partial depletion of Met and EGFR prevents their activation upon cell adhesion, suggesting that overexpression of the receptors is a prerequisite for their self-activation upon cell adhesion. Although Met and EGFR are largely distributed in 0.04% Triton-insoluble fractions (i.e. raft fraction), their activated forms are detected mainly in 0.04% Triton-soluble fractions (i.e. non-raft fraction). Upon cell adhesion, lipid rafts are accumulated at the cell surface close to the cell-substratum interface, while Met and EGFR are mostly excluded from the membrane enriched by lipid rafts. Our results suggest for the first time that cell adhesion to a substratum may induce a polarized distribution of lipid rafts to the cell-substratum interface, which may allow Met and EGFR to be released from lipid rafts, thus leading to their activation in a ligand-independent manner.

Activation of GPR4 by Acidosis Increases Endothelial Cell Adhesion through the cAMP/Epac Pathway

PubMed Central

Leffler, Nancy R.; Asch, Adam S.; Witte, Owen N.; Yang, Li V.

2011-01-01

Endothelium-leukocyte interaction is critical for inflammatory responses. Whereas the tissue microenvironments are often acidic at inflammatory sites, the mechanisms by which cells respond to acidosis are not well understood. Using molecular, cellular and biochemical approaches, we demonstrate that activation of GPR4, a proton-sensing G protein-coupled receptor, by isocapnic acidosis increases the adhesiveness of human umbilical vein endothelial cells (HUVECs) that express GPR4 endogenously. Acidosis in combination with GPR4 overexpression further augments HUVEC adhesion with U937 monocytes. In contrast, overexpression of a G protein signaling-defective DRY motif mutant (R115A) of GPR4 does not elicit any increase of HUVEC adhesion, indicating the requirement of G protein signaling. Downregulation of GPR4 expression by RNA interference reduces the acidosis-induced HUVEC adhesion. To delineate downstream pathways, we show that inhibition of adenylate cyclase by inhibitors, 2′,5′-dideoxyadenosine (DDA) or SQ 22536, attenuates acidosis/GPR4-induced HUVEC adhesion. Consistently, treatment with a cAMP analog or a Gi signaling inhibitor increases HUVEC adhesiveness, suggesting a role of the Gs/cAMP signaling in this process. We further show that the cAMP downstream effector Epac is important for acidosis/GPR4-induced cell adhesion. Moreover, activation of GPR4 by acidosis increases the expression of vascular adhesion molecules E-selectin, VCAM-1 and ICAM-1, which are functionally involved in acidosis/GPR4-mediated HUVEC adhesion. Similarly, hypercapnic acidosis can also activate GPR4 to stimulate HUVEC adhesion molecule expression and adhesiveness. These results suggest that acidosis/GPR4 signaling regulates endothelial cell adhesion mainly through the Gs/cAMP/Epac pathway and may play a role in the inflammatory response of vascular endothelial cells. PMID:22110680

Exploring the Limits of Cell Adhesion under Shear Stress within Physiological Conditions and beyond on a Chip.

PubMed

Stamp, Melanie E M; Jötten, Anna M; Kudella, Patrick W; Breyer, Dominik; Strobl, Florian G; Geislinger, Thomas M; Wixforth, Achim; Westerhausen, Christoph

2016-10-21

Cell adhesion processes are of ubiquitous importance for biomedical applications such as optimization of implant materials. Here, not only physiological conditions such as temperature or pH, but also topographical structures play crucial roles, as inflammatory reactions after surgery can diminish osseointegration. In this study, we systematically investigate cell adhesion under static, dynamic and physiologically relevant conditions employing a lab-on-a-chip system. We screen adhesion of the bone osteosarcoma cell line SaOs-2 on a titanium implant material for pH and temperature values in the physiological range and beyond, to explore the limits of cell adhesion, e.g., for feverish and acidic conditions. A detailed study of different surface roughness R q gives insight into the correlation between the cells' abilities to adhere and withstand shear flow and the topography of the substrates, finding a local optimum at R q = 22 nm. We use shear stress induced by acoustic streaming to determine a measure for the ability of cell adhesion under an external force for various conditions. We find an optimum of cell adhesion for T = 37 °C and pH = 7.4 with decreasing cell adhesion outside the physiological range, especially for high T and low pH. We find constant detachment rates in the physiological regime, but this behavior tends to collapse at the limits of 41 °C and pH 4.

Endothelial cell regulation of leukocyte infiltration in inflammatory tissues

PubMed Central

Mantovani, A.; Introna, M.; Dejana, E.

1995-01-01

Endothelial cells play an important, active role in the onset and regulation of inflammatory and immune reactions. Through the production of chemokines they attract leukocytes and activate their adhesive receptors. This leads to the anchorage of leukocytes to the adhesive molecules expressed on the endothelial surface. Leukocyte adhesion to endothelial cells is frequently followed by their extravasation. The mechanisms which regulate the passage of leukocytes through endothelial clefts remain to be clarified. Many indirect data suggest that leukocytes might transfer signals to endothelial cells both through the release of active agents and adhesion to the endothelial cell surface. Adhesive molecules (such as PECAM) on the endothelial cell surface might also ‘direct’ leukocytes through the intercellular junction by haptotaxis. The information available on the molecular structure and functional properties of endothelial chemokines, adhesive molecules or junction organization is still fragmentary. Further work is needed to clarify how they interplay in regulating leukocyte infiltration into tissues. PMID:18475659

Inhibition of sickle red cell adhesion and vasoocclusion in the microcirculation by antioxidants.

PubMed

Kaul, Dhananjay K; Liu, Xiao-du; Zhang, Xiaoqin; Ma, Li; Hsia, Carleton J C; Nagel, Ronald L

2006-07-01

In sickle cell anemia (SCA), inflammatory (i.e., intravascular sickling and transient vasoocclusive) events result in chronic endothelial activation. In addition to sickling behavior, sickle (SS) red blood cells exhibit abnormal interaction with the vascular endothelium, which is considered to have an important role in initiation of vasoocclusion. Upregulation of endothelial adhesion molecules caused by oxidants (and cytokines) may lead to increased SS red cell adhesion. We hypothesize that endothelial activation is indispensable in SS red cell adhesion to the endothelium and that antioxidants will have an inhibitory effect on this interaction. We examined the effect of selected antioxidants in ex vivo mesocecum vasculature, a well-established model that allows measurement of hemodynamic parameters and, by intravital microscopy, can allow quantification of adhesion. We tested antioxidant enzymes (SOD and catalase) and an intravascular SOD mimetic, polynitroxyl albumin (PNA), in the presence of platelet-activating factor (PAF); the latter causes endothelial oxidant generation and endothelial activation, which characterize SCA. In ex vivo preparations, PAF not only induced marked endothelial oxidant generation, it also enhanced SS red cell adhesion, resulting in frequent blockage of small-diameter venules. The adhesion, inversely related to venular diameter, and vasoocclusion were markedly inhibited by antioxidants, resulting in improved hemodynamics. PNA, the most effective antioxidant, also abolished SS red cell adhesion in non-PAF-activated preparations. Thus SS red cell adhesion and related vasoocclusion may be ameliorated by antioxidant therapy with a stable and long-acting molecule (e.g., PNA).

Neural cell adhesion molecule mediates initial interactions between spinal cord neurons and muscle cells in culture

PubMed Central

1983-01-01

Previous studies in this laboratory have described a cell surface glycoprotein, called neural cell adhesion molecule or N-CAM, that appears to be a ligand in the adhesion between neural membranes. N-CAM antigenic determinants were also shown to be present on embryonic muscle and an N-CAM-dependent adhesion was demonstrated between retinal cell membranes and muscle cells in short-term assays. The present studies indicate that these antigenic determinants are associated with the N-CAM polypeptide, and that rapid adhesion mediated by this molecule occurs between spinal cord membranes and muscle cells. Detailed examination of the effects of anti-(N-CAM) Fab' fragments in cultures of spinal cord with skeletal muscle showed that the Fab' fragments specifically block adhesion of spinal cord neurites and cells to myotubes. The Fab' did not affect binding of neurites to fibroblasts and collagen substrate, and did not alter myotube morphology. These results indicate that N-CAM adhesion is essential for the in vitro establishment of physical associations between nerve and muscle, and suggest that binding involving N-CAM may be an important early step in synaptogenesis. PMID:6863388

Inhibition of tumor necrosis factor-{alpha}-induced expression of adhesion molecules in human endothelial cells by the saponins derived from roots of Platycodon grandiflorum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Ji Young; Kim, Dong Hee; Kim, Hyung Gyun

2006-01-15

Adhesion molecules play an important role in the development of atherogenesis and are produced by endothelial cells after being stimulated with various inflammatory cytokines. This study examined the effect of saponins that were isolated from the roots of Platycodon grandiflorum A. DC (Campanulaceae), Changkil saponins (CKS), on the cytokine-induced monocyte/human endothelial cell interaction, which is a crucial early event in atherogenesis. CKS significantly inhibited the TNF{alpha}-induced increase in monocyte adhesion to endothelial cells as well as decreased the protein and mRNA expression levels of vascular adhesion molecule-1 and intercellular cell adhesion molecule-1 on endothelial cells. Furthermore, CKS significantly inhibited themore » TNF{alpha}-induced production of intracellular reactive oxygen species (ROS) and activation of NF-{kappa}B by preventing I{kappa}B degradation and inhibiting I{kappa}B kinase activity. Overall, CKS has anti-atherosclerotic and anti-inflammatory activity, which is least in part the result of it reducing the cytokine-induced endothelial adhesion to monocytes by inhibiting intracellular ROS production, NF-{kappa}B activation, and cell adhesion molecule expression in endothelial cells.« less

Simulated microgravity does not alter epithelial cell adhesion to matrix and other molecules

NASA Technical Reports Server (NTRS)

Jessup, J. M.; Brown, K.; Ishii, S.; Ford, R.; Goodwin, T. J.; Spaulding, G.

1994-01-01

Microgravity has advantages for the cultivation of tissues with high fidelity; however, tissue formation requires cellular recognition and adhesion. We tested the hypothesis that simulated microgravity does not affect cell adhesion. Human colorectal carcinoma cells were cultured in the NASA Rotating Wall Vessel (RWV) under low shear stress with randomization of the gravity vector that simulates microgravity. After 6 - 7 days, cells were assayed for binding to various substrates and compared to cells grown in standard tissue culture flasks and static suspension cultures. The RWV cultures bound as well to basement membrane proteins and to Carcinoembryonic Antigen (CEA), an intercellular adhesion molecule, as control cultures did. Thus, microgravity does not alter epithelial cell adhesion and may be useful for tissue engineering.

Structural Requirements for Outside-In and Inside-Out Signaling by Drosophila Neuroglian, a Member of the L1 Family of Cell Adhesion Molecules

PubMed Central

Hortsch, Michael; Homer, Diahann; Malhotra, Jyoti Dhar; Chang, Sherry; Frankel, Jason; Jefford, Gregory; Dubreuil, Ronald R.

1998-01-01

Expression of the Drosophila cell adhesion molecule neuroglian in S2 cells leads to cell aggregation and the intracellular recruitment of ankyrin to cell contact sites. We localized the region of neuroglian that interacts with ankyrin and investigated the mechanism that limits this interaction to cell contact sites. Yeast two-hybrid analysis and expression of neuroglian deletion constructs in S2 cells identified a conserved 36-amino acid sequence that is required for ankyrin binding. Mutation of a conserved tyrosine residue within this region reduced ankyrin binding and extracellular adhesion. However, residual recruitment of ankyrin by this mutant neuroglian molecule was still limited to cell contacts, indicating that the lack of ankyrin binding at noncontact sites is not caused by tyrosine phosphorylation. A chimeric molecule, in which the extracellular domain of neuroglian was replaced with the corresponding domain from the adhesion molecule fasciclin II, also selectively recruited ankyrin to cell contacts. Thus, outside-in signaling by neuroglian in S2 cells depends on extracellular adhesion, but does not depend on any unique property of its extracellular domain. We propose that the recruitment of ankyrin to cell contact sites depends on a physical rearrangement of neuroglian in response to cell adhesion, and that ankyrin binding plays a reciprocal role in stabilizing the adhesive interaction. PMID:9660878

Structural requirements for outside-in and inside-out signaling by Drosophila neuroglian, a member of the L1 family of cell adhesion molecules.

PubMed

Hortsch, M; Homer, D; Malhotra, J D; Chang, S; Frankel, J; Jefford, G; Dubreuil, R R

1998-07-13

Expression of the Drosophila cell adhesion molecule neuroglian in S2 cells leads to cell aggregation and the intracellular recruitment of ankyrin to cell contact sites. We localized the region of neuroglian that interacts with ankyrin and investigated the mechanism that limits this interaction to cell contact sites. Yeast two-hybrid analysis and expression of neuroglian deletion constructs in S2 cells identified a conserved 36-amino acid sequence that is required for ankyrin binding. Mutation of a conserved tyrosine residue within this region reduced ankyrin binding and extracellular adhesion. However, residual recruitment of ankyrin by this mutant neuroglian molecule was still limited to cell contacts, indicating that the lack of ankyrin binding at noncontact sites is not caused by tyrosine phosphorylation. A chimeric molecule, in which the extracellular domain of neuroglian was replaced with the corresponding domain from the adhesion molecule fasciclin II, also selectively recruited ankyrin to cell contacts. Thus, outside-in signaling by neuroglian in S2 cells depends on extracellular adhesion, but does not depend on any unique property of its extracellular domain. We propose that the recruitment of ankyrin to cell contact sites depends on a physical rearrangement of neuroglian in response to cell adhesion, and that ankyrin binding plays a reciprocal role in stabilizing the adhesive interaction.

Molecular mechanisms of mechanotransduction in integrin-mediated cell-matrix adhesion

PubMed Central

Li, Zhenhai; Lee, Hyunjung; Zhu, Cheng

2016-01-01

Cell-matrix adhesion complexes are multi-protein structures linking the extracellular matrix (ECM) to the cytoskeleton. They are essential to both cell motility and function by bidirectionally sensing and transmitting mechanical and biochemical stimulations. Several types of cell-matrix adhesions have been identified and they share many key molecular components, such as integrins and actin-integrin linkers. Mechanochemical coupling between ECM molecules and the actin cytoskeleton has been observed from the single cell to the single molecule level and from immune cells to neuronal cells. However, the mechanisms underlying force regulation of integrin-mediated mechanotransduction still need to be elucidated. In this review article, we focus on integrin-mediated adhesions and discuss force regulation of cell-matrix adhesions and key adaptor molecules, three different force-dependent behaviors, and molecular mechanisms for mechanochemical coupling in force regulation. PMID:27720950

EphA2 promotes cell adhesion and spreading of monocyte and monocyte/macrophage cell lines on integrin ligand-coated surfaces.

PubMed

Saeki, Noritaka; Nishino, Shingo; Shimizu, Tomohiro; Ogawa, Kazushige

2015-01-01

Eph signaling, which arises following stimulation by ephrins, is known to induce opposite cell behaviors such as promoting and inhibiting cell adhesion as well as promoting cell-cell adhesion and repulsion by altering the organization of the actin cytoskeleton and influencing the adhesion activities of integrins. However, crosstalk between Eph/ephrin with integrin signaling has not been fully elucidated in leukocytes, including monocytes and their related cells. Using a cell attachment stripe assay, we have shown that, following stimulation with ephrin-A1, kinase-independent EphA2 promoted cell spreading/elongation as well as adhesion to integrin ligand-coated surfaces in cultured U937 (monocyte) and J774.1 (monocyte/macrophage) cells as well as sublines of these cells expressing dominant negative EphA2 that lacks most of the intracellular region. Moreover, a pull-down assay showed that dominant negative EphA2 is recruited to the β2 integrin/ICAM1 and β2 integrin/VCAM1 molecular complexes in the subline cells following stimulation with ephrin-A1-Fc. Notably, this study is the first comprehensive analysis of the effects of EphA2 receptors on integrin-mediated cell adhesion in monocytic cells. Based on these findings we propose that EphA2 promotes cell adhesion by an unknown signaling pathway that largely depends on the extracellular region of EphA2 and the activation of outside-in integrin signaling.

Quantifying effects of cyclic stretch on cell-collagen substrate adhesiveness of vascular endothelial cells.

PubMed

Omidvar, Ramin; Tafazzoli-Shadpour, Mohammad; Mahmoodi-Nobar, Farbod; Azadi, Shohreh; Khani, Mohammad-Mehdi

2018-05-01

Vascular endothelium is continuously subjected to mechanical stimulation in the form of shear forces due to blood flow as well as tensile forces as a consequence of blood pressure. Such stimuli influence endothelial behavior and regulate cell-tissue interaction for an optimized functionality. This study aimed to quantify influence of cyclic stretch on the adhesive property and stiffness of endothelial cells. The 10% cyclic stretch with frequency of 1 Hz was applied to a layer of endothelial cells cultured on a polydimethylsiloxane substrate. Cell-substrate adhesion of endothelial cells was examined by the novel approach of atomic force microscope-based single-cell force spectroscopy and cell stiffness was measured by atomic force microscopy. Furthermore, the adhesive molecular bonds were evaluated using modified Hertz contact theory. Our results show that overall adhesion of endothelial cells with substrate decreased after cyclic stretch while they became stiffer. Based on the experimental results and theoretical modeling, the decrease in the number of molecular bonds after cyclic stretch was quantified. In conclusion, in vitro cyclic stretch caused alterations in both adhesive capacity and elastic modulus of endothelial cells through mechanotransductive pathways as two major determinants of the function of these cells within the cardiovascular system.

Adhesion kinetics of human primary monocytes, dendritic cells, and macrophages: Dynamic cell adhesion measurements with a label-free optical biosensor and their comparison with end-point assays.

PubMed

Orgovan, Norbert; Ungai-Salánki, Rita; Lukácsi, Szilvia; Sándor, Noémi; Bajtay, Zsuzsa; Erdei, Anna; Szabó, Bálint; Horvath, Robert

2016-09-01

Monocytes, dendritic cells (DCs), and macrophages (MFs) are closely related immune cells that differ in their main functions. These specific functions are, to a considerable degree, determined by the differences in the adhesion behavior of the cells. To study the inherently and essentially dynamic aspects of the adhesion of monocytes, DCs, and MFs, dynamic cell adhesion assays were performed with a high-throughput label-free optical biosensor [Epic BenchTop (BT)] on surfaces coated with either fibrinogen (Fgn) or the biomimetic copolymer PLL-g-PEG-RGD. Cell adhesion profiles typically reached their maximum at ∼60 min after cell seeding, which was followed by a monotonic signal decrease, indicating gradually weakening cell adhesion. According to the biosensor response, cell types could be ordered by increasing adherence as monocytes, MFs, and DCs. Notably, all three cell types induced a larger biosensor signal on Fgn than on PLL-g-PEG-RGD. To interpret this result, the molecular layers were characterized by further exploiting the potentials of the biosensor: by measuring the adsorption signal induced during the surface coating procedure, the authors could estimate the surface density of adsorbed molecules and, thus, the number of binding sites potentially presented for the adhesion receptors. Surfaces coated with PLL-g-PEG-RGD presented less RGD sites, but was less efficient in promoting cell spreading than those coated with Fgn; hence, other binding sites in Fgn played a more decisive role in determining cell adherence. To support the cell adhesion data obtained with the biosensor, cell adherence on Fgn-coated surfaces 30-60 min after cell seeding was measured with three complementary techniques, i.e., with (1) a fluorescence-based classical adherence assay, (2) a shear flow chamber applying hydrodynamic shear stress to wash cells away, and (3) an automated micropipette using vacuum-generated fluid flow to lift cells up. These techniques confirmed the results obtained with the high-temporal-resolution Epic BT, but could only provide end-point data. In contrast, complex, nonmonotonic cell adhesion kinetics measured by the high-throughput optical biosensor is expected to open a window on the hidden background of the immune cell-extracellular matrix interactions.

Suppression of endothelial cell adhesion by XJP-1, a new phenolic compound derived from banana peel.

PubMed

Fu, Rong; Yan, Tianhua; Wang, Qiujuan; Guo, Qinglong; Yao, Hequan; Wu, Xiaoming; Li, Yang

2012-01-01

The adhesion of monocytes to activated vascular endothelial cells is a critical event in the initiation of atherosclerosis. Adhesion is mediated by oxidized low-density lipoprotein (ox-LDL) which up-regulates inflammatory markers on endothelial cells. Here we report that (±) 7, 8-dihydroxy-3-methyl-isochromanone-4 (XJP-1), an inhibitor of ox-LDL-induced adhesion of monocytes to endothelial cells blocks cellular functions which are associated with adhesion. We show that XJP-1 down-regulates ox-LDL-induced over-expression of adhesion molecules (ICAM-1 and VCAM-1) in a dose-dependent manner in human umbilical vein endothelial cells (HUVECs), attenuates ox-LDL-induced up-regulation of low-density lipoprotein receptor (LOX)-1, decreases generation of reactive oxygen species (ROS), blocks translocation of nuclear factor-kappa B (NF-κB) activity, and prevents activation of c-Jun N-terminal kinase (JNK)/p38 pathways in endothelial cells. These findings suggest that XJP-1 may attenuate ox-LDL-induced endothelial adhesion of monocytes by blocking expression of adhesion molecules through suppressing ROS/NF-κB, JNK and p38 pathways. Copyright © 2012 Elsevier Inc. All rights reserved.

Isolation of integrin-based adhesion complexes.

PubMed

Jones, Matthew C; Humphries, Jonathan D; Byron, Adam; Millon-Frémillon, Angélique; Robertson, Joseph; Paul, Nikki R; Ng, Daniel H J; Askari, Janet A; Humphries, Martin J

2015-03-02

The integration of cells with their extracellular environment is facilitated by cell surface adhesion receptors, such as integrins, which play important roles in both normal development and the onset of pathologies. Engagement of integrins with their ligands in the extracellular matrix, or counter-receptors on other cells, initiates the intracellular assembly of a wide variety of proteins into adhesion complexes such as focal contacts, focal adhesions, and fibrillar adhesions. The proteins recruited to these complexes mediate bidirectional signaling across the plasma membrane, and, as such, help to coordinate and/or modulate the multitude of physical and chemical signals to which the cell is subjected. The protocols in this unit describe two approaches for the isolation or enrichment of proteins contained within integrin-associated adhesion complexes, together with their local plasma membrane/cytosolic environments, from cells in culture. In the first protocol, integrin-associated adhesion structures are affinity isolated using microbeads coated with extracellular ligands or antibodies. The second protocol describes the isolation of ventral membrane preparations that are enriched for adhesion complex structures. The protocols permit the determination of adhesion complex components via subsequent downstream analysis by western blotting or mass spectrometry. Copyright © 2015 John Wiley & Sons, Inc.

Desmoglein 3 regulates membrane trafficking of cadherins, an implication in cell-cell adhesion.

PubMed

Moftah, Hanan; Dias, Kasuni; Apu, Ehsanul Hoque; Liu, Li; Uttagomol, Jutamas; Bergmeier, Lesley; Kermorgant, Stephanie; Wan, Hong

2017-05-04

E-cadherin mediated cell-cell adhesion plays a critical role in epithelial cell polarization and morphogenesis. Our recent studies suggest that the desmosomal cadherin, desmoglein 3 (Dsg3) cross talks with E-cadherin and regulates its adhesive function in differentiating keratinocytes. However, the underlying mechanism remains not fully elucidated. Since E-cadherin trafficking has been recognized to be a central determinant in cell-cell adhesion and homeostasis we hypothesize that Dsg3 may play a role in regulating E-cadherin trafficking and hence the cell-cell adhesion. Here we investigated this hypothesis in cells with loss of Dsg3 function through RNAi mediated Dsg3 knockdown or the stable expression of the truncated mutant Dsg3ΔC. Our results showed that loss of Dsg3 resulted in compromised cell-cell adhesion and reduction of adherens junction and desmosome protein expression as well as the cortical F-actin formation. As a consequence, cells failed to polarize but instead displayed aberrant cell flattening. Furthermore, retardation of E-cadherin internalization and recycling was consistently observed in these cells during the process of calcium induced junction assembling. In contrast, enhanced cadherin endocytosis was detected in cells with overexpression of Dsg3 compared to control cells. Importantly, this altered cadherin trafficking was found to be coincided with the reduced expression and activity of Rab proteins, including Rab5, Rab7 and Rab11 which are known to be involved in E-cadherin trafficking. Taken together, our findings suggest that Dsg3 functions as a key in cell-cell adhesion through at least a mechanism of regulating E-cadherin membrane trafficking.

GABAergic Synapses at the Axon Initial Segment of Basolateral Amygdala Projection Neurons Modulate Fear Extinction.

PubMed

Saha, Rinki; Knapp, Stephanie; Chakraborty, Darpan; Horovitz, Omer; Albrecht, Anne; Kriebel, Martin; Kaphzan, Hanoch; Ehrlich, Ingrid; Volkmer, Hansjürgen; Richter-Levin, Gal

2017-01-01

Inhibitory synaptic transmission in the amygdala has a pivotal role in fear learning and its extinction. However, the local circuits formed by GABAergic inhibitory interneurons within the amygdala and their detailed function in shaping these behaviors are not well understood. Here we used lentiviral-mediated knockdown of the cell adhesion molecule neurofascin in the basolateral amygdala (BLA) to specifically remove inhibitory synapses at the axon initial segment (AIS) of BLA projection neurons. Quantitative analysis of GABAergic synapse markers and measurement of miniature inhibitory postsynaptic currents in BLA projection neurons after neurofascin knockdown ex vivo confirmed the loss of GABAergic input. We then studied the impact of this manipulation on anxiety-like behavior and auditory cued fear conditioning and its extinction as BLA related behavioral paradigms, as well as on long-term potentiation (LTP) in the ventral subiculum-BLA pathway in vivo. BLA knockdown of neurofascin impaired ventral subiculum-BLA-LTP. While this manipulation did not affect anxiety-like behavior and fear memory acquisition and consolidation, it specifically impaired extinction. Our findings indicate that modification of inhibitory synapses at the AIS of BLA projection neurons is sufficient to selectively impair extinction behavior. A better understanding of the role of distinct GABAergic synapses may provide novel and more specific targets for therapeutic interventions in extinction-based therapies.

The interaction between LYVE-1 with hyaluronan on the cell surface may play a role in the diversity of adhesion to cancer cells.

PubMed

Du, Yan; Liu, Hua; He, Yiqing; Liu, Yiwen; Yang, Cuixia; Zhou, Muqing; Wang, Wenjuan; Cui, Lian; Hu, Jiajie; Gao, Feng

2013-01-01

Hyaluronan (HA), a simple disaccharide unit, can polymerize and is considered a primary component of the extracellular matrix, which has a wide range of biological functions. In recent years, HA was found on the surface of tumor cells. According to previous reports, differing HA content on the cell surface of tumor cells is closely related to lymph node metastases, but the mechanisms mediating this process remained unclear. This research intended to study the surface content of HA on tumor cells and analyze cell adhesive changes caused by the interaction between HA and its lymphatic endothelial receptor (LYVE-1). We screened and observed high HA content on HS-578T breast cells and low HA content on MCF-7 breast cells through particle exclusion, immunofluorescence and flow cytometry experiments. The expression of LYVE-1, the lymph-vessel specific HA receptor, was consistent with our previous report and enhanced the adhesion of HA(high)-HS-578T cells to COS-7(LYVE-1(+)) through HA in cell static adhesion and dynamic parallel plate flow chamber experiments. MCF-7 breast cells contain little HA on the surface; however, our results showed little adhesion difference between MCF-7 cells and COS-7(LYVE-1(+)) and COS-7(LYVE-1(-)) cells. Similar results were observed concerning the adhesion of HS-578T cells or MCF-7 cells to SVEC4-10 cells. Furthermore, we observed for the first time that the cell surface HA content of high transfer tumor cells was rich, and we visualized the cross-linking of HA cable structures, which may activate LYVE-1 on lymphatic endothelial cells, promoting tumor adhesion. In summary, high-low cell surface HA content of tumor cells through the interaction with LYVE-1 leads to adhesion differences.

Human Freud-2/CC2D1B: a novel repressor of postsynaptic serotonin-1A receptor expression.

PubMed

Hadjighassem, Mahmoud R; Austin, Mark C; Szewczyk, Bernadeta; Daigle, Mireille; Stockmeier, Craig A; Albert, Paul R

2009-08-01

Altered expression of serotonin-1A (5-HT1A) receptors, both presynaptic in the raphe nuclei and post-synaptic in limbic and cortical target areas, has been implicated in mood disorders such as major depression and anxiety. Within the 5-HT1A receptor gene, a powerful dual repressor element (DRE) is regulated by two protein complexes: Freud-1/CC2D1A and a second, unknown repressor. Here we identify human Freud-2/CC2D1B, a Freud-1 homologue, as the second repressor. Freud-2 distribution was examined with Northern and Western blot, reverse transcriptase polymerase chain reaction, and immunohistochemistry/immunofluorescence; Freud-2 function was examined by electrophoretic mobility shift, reporter assay, and Western blot. Freud-2 RNA was widely distributed in brain and peripheral tissues. Freud-2 protein was enriched in the nuclear fraction of human prefrontal cortex and hippocampus but was weakly expressed in the dorsal raphe nucleus. Freud-2 immunostaining was co-localized with 5-HT1A receptors, neuronal and glial markers. In prefrontal cortex, Freud-2 was expressed at similar levels in control and depressed male subjects. Recombinant hFreud-2 protein bound specifically to 5' or 3' human DRE adjacent to the Freud-1 site. Human Freud-2 showed strong repressor activity at the human 5-HT1A or heterologous promoter in human HEK-293 5-HT1A-negative cells and neuronal SK-N-SH cells, a model of postsynaptic 5-HT1A receptor-positive cells. Furthermore, small interfering RNA knockdown of endogenous hFreud-2 expression de-repressed 5-HT1A promoter activity and increased levels of 5-HT1A receptor protein in SK-N-SH cells. Human Freud-2 binds to the 5-HT1A DRE and represses the human 5-HT1A receptor gene to regulate its expression in non-serotonergic cells and neurons.

The Synaptic Cell Adhesion Molecule, SynCAM1, Mediates Astrocyte-to-Astrocyte and Astrocyte-to-GnRH Neuron Adhesiveness in the Mouse Hypothalamus

PubMed Central

Sandau, Ursula S.; Mungenast, Alison E.; McCarthy, Jack; Biederer, Thomas; Corfas, Gabriel

2011-01-01

We previously identified synaptic cell adhesion molecule 1 (SynCAM1) as a component of a genetic network involved in the hypothalamic control of female puberty. Although it is well established that SynCAM1 is a synaptic adhesion molecule, its contribution to hypothalamic function is unknown. Here we show that, in addition to the expected neuronal localization illustrated by its presence in GnRH neurons, SynCAM1 is expressed in hypothalamic astrocytes. Cell adhesion assays indicated that SynCAM is recognized by both GnRH neurons and astrocytes as an adhesive partner and promotes cell-cell adhesiveness via homophilic, extracellular domain-mediated interactions. Alternative splicing of the SynCAM1 primary mRNA transcript yields four mRNAs encoding membrane-spanning SynCAM1 isoforms. Variants 1 and 4 are predicted to be both N and O glycosylated. Hypothalamic astrocytes and GnRH-producing GT1-7 cells express mainly isoform 4 mRNA, and sequential N- and O-deglycosylation of proteins extracted from these cells yields progressively smaller SynCAM1 species, indicating that isoform 4 is the predominant SynCAM1 variant expressed in astrocytes and GT1-7 cells. Neither cell type expresses the products of two other SynCAM genes (SynCAM2 and SynCAM3), suggesting that SynCAM-mediated astrocyte-astrocyte and astrocyte-GnRH neuron adhesiveness is mostly mediated by SynCAM1 homophilic interactions. When erbB4 receptor function is disrupted in astrocytes, via transgenic expression of a dominant-negative erbB4 receptor form, SynCAM1-mediated adhesiveness is severely compromised. Conversely, SynCAM1 adhesive behavior is rapidly, but transiently, enhanced in astrocytes by ligand-dependent activation of erbB4 receptors, suggesting that erbB4-mediated events affecting SynCAM1 function contribute to regulate astrocyte adhesive communication. PMID:21486931

Chemical and physical effects on the adhesion, maturation, and survival of monocytes, macrophages, and foreign body giant cells

NASA Astrophysics Data System (ADS)

Collier, Terry Odell, III

Injury caused by biomedical device implantation initiates inflammatory and wound healing responses. Cells migrate to the site of injury to degrade bacteria and toxins, create new vasculature, and form new and repair injured tissue. Blood-proteins rapidly adsorb onto the implanted material surface and express adhesive ligands which mediate cell adhesion on the material surface. Monocyte-derived macrophages and multi-nucleated foreign body giant cells adhere to the surface and degrade the surface of the material. Due to the role of macrophage and foreign body giant cell on material biocompatibility and biostability, the effects of surface chemistry, surface topography and specific proteins on the maturation and survival of monocytes, macrophages and foreign body giant cells has been investigated. Novel molecularly designed materials were used to elucidate the dynamic interactions which occur between inflammatory cells, proteins and surfaces. The effect of protein and protein adhesion was investigated using adhesive protein depleted serum conditions on RGD-modified and silane modified surfaces. The effects of surface chemistry were investigated using temperature responsive surfaces of poly (N-isopropylacrylamide) and micropatterned surfaces of N-(2 aminoethyl)-3-aminopropyltrimethoxysilane regions on an interpenetrating polymer network of polyacrylamide and poly(ethylene glycol). The physical effects were investigated using polyimide scaffold materials and polyurethane materials with surface modifying end groups. The depletion of immunoglobulin G caused decreased levels of macrophage adhesion, foreign body giant cell formation and increased levels of apoptosis. The temporal nature of macrophage adhesion was observed with changing effectiveness of adherent cell detachment with time, which correlated to increased expression of beta1 integrin receptors on detached macrophages with time. The limited ability of the micropatterned surface, polyimide scaffold and surface modified polyurethane materials to control macrophage adhesion indicates the complexity of macrophage adhesion and protein adsorption onto a surface. These studies have indicated components and adhesive mechanisms which can be utilized to create materials with enhanced resistance to macrophage adhesion and/or degradative abilities.

Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells

PubMed Central

Zhang, Bo; Zuo, Fanglei; Yu, Rui; Zeng, Zhu; Ma, Huiqin; Chen, Shangwu

2015-01-01

Adhesion to host cells is considered important for Lactobacillus plantarum as well as other lactic acid bacteria (LAB) to persist in human gut and thus exert probiotic effects. Here, we sequenced the genome of Lt. plantarum strain NL42 originating from a traditional Chinese dairy product, performed comparative genomic analysis and characterized a novel adhesion factor. The genome of NL42 was highly divergent from its closest neighbors, especially in six large genomic regions. NL42 harbors a total of 42 genes encoding adhesion-associated proteins; among them, cwaA encodes a protein containing multiple domains, including five cell wall surface anchor repeat domains and an LPxTG-like cell wall anchor motif. Expression of cwaA in Lactococcus lactis significantly increased its autoaggregation and hydrophobicity, and conferred the new ability to adhere to human colonic epithelial HT-29 cells by targeting cellular surface proteins, and not carbohydrate moieties, for CwaA adhesion. In addition, the recombinant Lc. lactis inhibited adhesion of Staphylococcus aureus and Escherichia coli to HT-29 cells, mainly by exclusion. We conclude that CwaA is a novel adhesion factor in Lt. plantarum and a potential candidate for improving the adhesion ability of probiotics or other bacteria of interest. PMID:26370773

Patterning N-type and S-type Neuroblastoma Cells with Pluronic F108 and ECM Proteins

PubMed Central

Corey, Joseph M.; Gertz, Caitlyn C.; Sutton, Thomas J.; Chen, Qiaoran; Mycek, Katherine B.; Wang, Bor-Shuen; Martin, Abbey A.; Johnson, Sara L.; Feldman, Eva L.

2009-01-01

Influencing cell shape using micropatterned substrates affects cell behaviors, such as proliferation and apoptosis. Cell shape may also affect these behaviors in human neuroblastoma (NBL) cancer, but to date, no substrate design has effectively patterned multiple clinically important human NBL lines. In this study, we investigated whether Pluronic F108 was an effective anti-adhesive coating for human NBL cells and whether it would localize three NBL lines to adhesive regions of tissue culture plastic or collagen I on substrate patterns. The adhesion and patterning of an S-type line, SH-EP, and two N-type lines, SH-SY5Y and IMR-32, were tested. In adhesion assays, F108 deterred NBL adhesion equally as well as two anti-adhesive organofunctional silanes and far better than bovine serum albumin. Patterned stripes of F108 restricted all three human NBL lines to adhesive stripes of tissue culture plastic. We then investigated four schemes of applying collagen and F108 to different regions of a substrate. Contact with collagen obliterates the ability of F108 to deter NBL adhesion, limiting how both materials can be applied to substrates to produce high fidelity NBL patterning. This patterned substrate design should facilitate investigations of the role of cell shape in NBL cell behavior. PMID:19609877

Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells.

PubMed

Zhang, Bo; Zuo, Fanglei; Yu, Rui; Zeng, Zhu; Ma, Huiqin; Chen, Shangwu

2015-09-15

Adhesion to host cells is considered important for Lactobacillus plantarum as well as other lactic acid bacteria (LAB) to persist in human gut and thus exert probiotic effects. Here, we sequenced the genome of Lt. plantarum strain NL42 originating from a traditional Chinese dairy product, performed comparative genomic analysis and characterized a novel adhesion factor. The genome of NL42 was highly divergent from its closest neighbors, especially in six large genomic regions. NL42 harbors a total of 42 genes encoding adhesion-associated proteins; among them, cwaA encodes a protein containing multiple domains, including five cell wall surface anchor repeat domains and an LPxTG-like cell wall anchor motif. Expression of cwaA in Lactococcus lactis significantly increased its autoaggregation and hydrophobicity, and conferred the new ability to adhere to human colonic epithelial HT-29 cells by targeting cellular surface proteins, and not carbohydrate moieties, for CwaA adhesion. In addition, the recombinant Lc. lactis inhibited adhesion of Staphylococcus aureus and Escherichia coli to HT-29 cells, mainly by exclusion. We conclude that CwaA is a novel adhesion factor in Lt. plantarum and a potential candidate for improving the adhesion ability of probiotics or other bacteria of interest.

Cell adhesion to borate glasses by colloidal probe microscopy.

PubMed

Wiederhorn, Sheldon M; Chae, Young-Hun; Simon, Carl G; Cahn, Jackson; Deng, Yan; Day, Delbert

2011-05-01

The adhesion of osteoblast-like cells to silicate and borate glasses was measured in cell growth medium using colloidal probe microscopy. The probes consisted of silicate and borate glass spheres, 25-50 μm in diameter, attached to atomic force microscope cantilevers. Variables of the study included glass composition and time of contact of the cell to the glasses. Increasing the time of contact from 15 to 900 s increased the force of adhesion. The data could be plotted linearly on a log-log plot of adhesive force versus time. Of the seven glasses tested, five had slopes close to 0.5, suggesting a square root dependence of the adhesive force on the contact time. Such behavior can be interpreted as a diffusion limited process occurring during the early stages of cell attachment. We suggest that the rate limiting step in the adhesion process is the diffusion of integrins resident in the cell membrane to the area of cell attachment. Data presented in this paper support the hypothesis of Hench et al. that strong adhesion depends on the formation of a calcium phosphate reaction layer on the surfaces of the glass. Glasses that did not form a calcium phosphate layer exhibited a weaker adhesive force relative to those glasses that did form a calcium phosphate layer. Published by Elsevier Ltd.

Friction-Controlled Traction Force in Cell Adhesion

PubMed Central

Pompe, Tilo; Kaufmann, Martin; Kasimir, Maria; Johne, Stephanie; Glorius, Stefan; Renner, Lars; Bobeth, Manfred; Pompe, Wolfgang; Werner, Carsten

2011-01-01

The force balance between the extracellular microenvironment and the intracellular cytoskeleton controls the cell fate. We report a new (to our knowledge) mechanism of receptor force control in cell adhesion originating from friction between cell adhesion ligands and the supporting substrate. Adherent human endothelial cells have been studied experimentally on polymer substrates noncovalently coated with fluorescent-labeled fibronectin (FN). The cellular traction force correlated with the mobility of FN during cell-driven FN fibrillogenesis. The experimental findings have been explained within a mechanistic two-dimensional model of the load transfer at focal adhesion sites. Myosin motor activity in conjunction with sliding of FN ligands noncovalently coupled to the surface of the polymer substrates is shown to result in a controlled traction force of adherent cells. We conclude that the friction of adhesion ligands on the supporting substrate is important for mechanotransduction and cell development of adherent cells in vitro and in vivo. PMID:22004739

Actin filaments regulate the adhesion between the plasma membrane and the cell wall of tobacco guard cells.

PubMed

Yu, Qin; Ren, Jing-Jing; Kong, Lan-Jing; Wang, Xiu-Ling

2018-01-01

During the opening and closing of stomata, guard cells undergo rapid and reversible changes in their volume and shape, which affects the adhesion of the plasma membrane (PM) to the cell wall (CW). The dynamics of actin filaments in guard cells are involved in stomatal movement by regulating structural changes and intracellular signaling. However, it is unclear whether actin dynamics regulate the adhesion of the PM to the CW. In this study, we investigated the relationship between actin dynamics and PM-CW adhesion by the hyperosmotic-induced plasmolysis of tobacco guard cells. We found that actin filaments in guard cells were depolymerized during mannitol-induced plasmolysis. The inhibition of actin dynamics by treatment with latrunculin B or jasplakinolide and the disruption of the adhesion between the PM and the CW by treatment with RGDS peptide (Arg-Gly-Asp-Ser) enhanced guard cell plasmolysis. However, treatment with latrunculin B alleviated the RGDS peptide-induced plasmolysis and endocytosis. Our results reveal that the actin depolymerization is involved in the regulation of the PW-CW adhesion during hyperosmotic-induced plasmolysis in tobacco guard cells.

Toll-like receptor (TLR)-1/2 triggering of multiple myeloma cells modulates their adhesion to bone marrow stromal cells and enhances bortezomib-induced apoptosis.

PubMed

Abdi, Jahangir; Mutis, Tuna; Garssen, Johan; Redegeld, Frank A

2014-01-01

In multiple myeloma (MM), the malignant plasma cells usually localize to the bone marrow where they develop drug resistance due to adhesion to stromal cells and various environmental signals. Hence, modulation of this interaction is expected to influence drug sensitivity of MM cells. Toll-like receptor (TLR) ligands have displayed heterogeneous effects on B-cell malignancies and also on MM cells in a few recent studies, but effects on adhesion and drug sensitivity of myeloma cells in the context of bone marrow stromal cells (BMSCs) have never been investigated. In the present study, we explored the modulatory effects of TLR1/2 ligand (Pam3CSK4) on adhesion of human myeloma cells to BMSCs. It is shown that TLR1/2 triggering has opposite effects in different HMCLs on their adhesion to BMSCs. Fravel, L363, UM-6, UM-9 and U266 showed increased adhesion to BMSC in parallel with an increased surface expression of integrin molecules α4 and αVβ3. OPM-1, OPM-2 and NCI-H929 showed a dose-dependent decrease in adhesion upon TLR activation following a downregulation of β7 integrin expression. Importantly, TLR1/2 triggering increased cytotoxic and apoptotic effects of bortezomib in myeloma cells independent of the effect on stromal cell adhesion. Moreover, the apoptosis-enhancing effect of Pam3CSK4 paralleled induction of cleaved caspase-3 protein in FACS analysis suggesting a caspase-dependent mechanism. Our findings uncover a novel role of TLR activation in MM cells in the context of bone marrow microenvironment. Stimulation of TLR1/2 bypasses the protective shield of BMSCs and may be an interesting strategy to enhance drug sensitivity of multiple myeloma cells.

Local palmitoylation cycles define activity-regulated postsynaptic subdomains

PubMed Central

Fukata, Yuko; Dimitrov, Ariane; Boncompain, Gaelle; Vielemeyer, Ole

2013-01-01

Distinct PSD-95 clusters are primary landmarks of postsynaptic densities (PSDs), which are specialized membrane regions for synapses. However, the mechanism that defines the locations of PSD-95 clusters and whether or how they are reorganized inside individual dendritic spines remains controversial. Because palmitoylation regulates PSD-95 membrane targeting, we combined a conformation-specific recombinant antibody against palmitoylated PSD-95 with live-cell super-resolution imaging and discovered subsynaptic nanodomains composed of palmitoylated PSD-95 that serve as elementary units of the PSD. PSD-95 in nanodomains underwent continuous de/repalmitoylation cycles driven by local palmitoylating activity, ensuring the maintenance of compartmentalized PSD-95 clusters within individual spines. Plasma membrane targeting of DHHC2 palmitoyltransferase rapidly recruited PSD-95 to the plasma membrane and proved essential for postsynaptic nanodomain formation. Furthermore, changes in synaptic activity rapidly reorganized PSD-95 nano-architecture through plasma membrane–inserted DHHC2. Thus, the first genetically encoded antibody sensitive to palmitoylation reveals an instructive role of local palmitoylation machinery in creating activity-responsive PSD-95 nanodomains, contributing to the PSD (re)organization. PMID:23836932

Mena binds α5 integrin directly and modulates α5β1 function.

PubMed

Gupton, Stephanie L; Riquelme, Daisy; Hughes-Alford, Shannon K; Tadros, Jenny; Rudina, Shireen S; Hynes, Richard O; Lauffenburger, Douglas; Gertler, Frank B

2012-08-20

Mena is an Ena/VASP family actin regulator with roles in cell migration, chemotaxis, cell-cell adhesion, tumor cell invasion, and metastasis. Although enriched in focal adhesions, Mena has no established function within these structures. We find that Mena forms an adhesion-regulated complex with α5β1 integrin, a fibronectin receptor involved in cell adhesion, motility, fibronectin fibrillogenesis, signaling, and growth factor receptor trafficking. Mena bound directly to the carboxy-terminal portion of the α5 cytoplasmic tail via a 91-residue region containing 13 five-residue "LERER" repeats. In fibroblasts, the Mena-α5 complex was required for "outside-in" α5β1 functions, including normal phosphorylation of FAK and paxillin and formation of fibrillar adhesions. It also supported fibrillogenesis and cell spreading and controlled cell migration speed. Thus, fibroblasts require Mena for multiple α5β1-dependent processes involving bidirectional interactions between the extracellular matrix and cytoplasmic focal adhesion proteins.

Integrative systems and synthetic biology of cell-matrix adhesion sites.

PubMed

Zamir, Eli

2016-09-02

The complexity of cell-matrix adhesion convolves its roles in the development and functioning of multicellular organisms and their evolutionary tinkering. Cell-matrix adhesion is mediated by sites along the plasma membrane that anchor the actin cytoskeleton to the matrix via a large number of proteins, collectively called the integrin adhesome. Fundamental challenges for understanding how cell-matrix adhesion sites assemble and function arise from their multi-functionality, rapid dynamics, large number of components and molecular diversity. Systems biology faces these challenges in its strive to understand how the integrin adhesome gives rise to functional adhesion sites. Synthetic biology enables engineering intracellular modules and circuits with properties of interest. In this review I discuss some of the fundamental questions in systems biology of cell-matrix adhesion and how synthetic biology can help addressing them.

TES is a novel focal adhesion protein with a role in cell spreading.

PubMed

Coutts, Amanda S; MacKenzie, Elaine; Griffith, Elen; Black, Donald M

2003-03-01

Previously, we identified TES as a novel candidate tumour suppressor gene that mapped to human chromosome 7q31.1. In this report we demonstrate that the TES protein is localised at focal adhesions, actin stress fibres and areas of cell-cell contact. TES has three C-terminal LIM domains that appear to be important for focal adhesion targeting. Additionally, the N-terminal region is important for targeting TES to actin stress fibres. Yeast two-hybrid and biochemical analyses yielded interactions with several focal adhesion and/or cytoskeletal proteins including mena, zyxin and talin. The fact that TES localises to regions of cell adhesion suggests that it functions in events related to cell motility and adhesion. In support of this, we demonstrate that fibroblasts stably overexpressing TES have an increased ability to spread on fibronectin.

Synaptic dysfunction and intellectual disability.

PubMed

Valnegri, Pamela; Sala, Carlo; Passafaro, Maria

2012-01-01

Intellectual disability (ID) is a common and highly heterogeneous paediatric disorder with a very severe social impact. Intellectual disability can be caused by environmental and/or genetic factors. Although in the last two decades a number of genes have been discovered whose mutations cause mental retardation, we are still far from identifying the impact of these mutations on brain functions. Many of the genes mutated in ID code for several proteins with a variety of functions: chromatin remodelling, pre-/post-synaptic activity, and intracellular trafficking. The prevailing hypothesis suggests that the ID phenotype could emerge from abnormal cellular processing leading to pre- and/or post-synaptic dysfunction. In this chapter, we focus on the role of small GTPases and adhesion molecules, and we discuss the mechanisms through which they lead to synaptic network dysfunction.

Orai1 as New Therapeutic Target for Inhibiting Breast Tumor Metastasis

DTIC Science & Technology

2009-09-01

includes focal adhesion assembly (formation of focal complex) and focal adhesion disassembly, we used live - cell imaging to quantify the rates of assembly...A and B) Live cell imaging of paxillin-GFP transfected MEF cells in the absence (A) or presence (B) of SKF96365. Scale bar: 10 µm. (C and D...includes focal adhesion assembly (formation of focal complexes) and focal adhesion disassembly, we used live - cell imaging to quantify the rates of focal

Investigation of adhesion and mechanical properties of human glioma cells by single cell force spectroscopy and atomic force microscopy.

PubMed

Andolfi, Laura; Bourkoula, Eugenia; Migliorini, Elisa; Palma, Anita; Pucer, Anja; Skrap, Miran; Scoles, Giacinto; Beltrami, Antonio Paolo; Cesselli, Daniela; Lazzarino, Marco

2014-01-01

Active cell migration and invasion is a peculiar feature of glioma that makes this tumor able to rapidly infiltrate into the surrounding brain tissue. In our recent work, we identified a novel class of glioma-associated-stem cells (defined as GASC for high-grade glioma--HG--and Gasc for low-grade glioma--LG) that, although not tumorigenic, act supporting the biological aggressiveness of glioma-initiating stem cells (defined as GSC for HG and Gsc for LG) favoring also their motility. Migrating cancer cells undergo considerable molecular and cellular changes by remodeling their cytoskeleton and cell interactions with surrounding environment. To get a better understanding about the role of the glioma-associated-stem cells in tumor progression, cell deformability and interactions between glioma-initiating stem cells and glioma-associated-stem cells were investigated. Adhesion of HG/LG-cancer cells on HG/LG-glioma-associated stem cells was studied by time-lapse microscopy, while cell deformability and cell-cell adhesion strengths were quantified by indentation measurements by atomic force microscopy and single cell force spectroscopy. Our results demonstrate that for both HG and LG glioma, cancer-initiating-stem cells are softer than glioma-associated-stem cells, in agreement with their neoplastic features. The adhesion strength of GSC on GASC appears to be significantly lower than that observed for Gsc on Gasc. Whereas, GSC spread and firmly adhere on Gasc with an adhesion strength increased as compared to that obtained on GASC. These findings highlight that the grade of glioma-associated-stem cells plays an important role in modulating cancer cell adhesion, which could affect glioma cell migration, invasion and thus cancer aggressiveness. Moreover this work provides evidence about the importance of investigating cell adhesion and elasticity for new developments in disease diagnostics and therapeutics.

Receptor-like Molecules on Human Intestinal Epithelial Cells Interact with an Adhesion Factor from Lactobacillus reuteri.

PubMed

Matsuo, Yosuke; Miyoshi, Yukihiro; Okada, Sanae; Satoh, Eiichi

2012-01-01

A surface protein of Lactobacillus reuteri, mucus adhesion-promoting protein (MapA), is considered to be an adhesion factor. MapA is expressed in L. reuteri strains and adheres to piglet gastric mucus, collagen type I, and human intestinal epithelial cells such as Caco-2. The aim of this study was to identify molecules that mediate the attachment of MapA from L. reuteri to the intestinal epithelial cell surface by investigating the adhesion of MapA to receptor-like molecules on Caco-2 cells. MapA-binding receptor-like molecules were detected in Caco-2 cell lysates by 2D-PAGE. Two proteins, annexin A13 (ANXA13) and paralemmin (PALM), were identified by MALDI TOF-MS. The results of a pull-down assay showed that MapA bound directly to ANXA13 and PALM. Fluorescence microscopy studies confirmed that MapA binding to ANXA13 and PALM was colocalized on the Caco-2 cell membrane. To evaluate whether ANXA13 and PALM are important for MapA adhesion, ANXA13 and PALM knockdown cell lines were established. The adhesion of MapA to the abovementioned cell lines was reduced compared with that to wild-type Caco-2 cells. These knockdown experiments established the importance of these receptor-like molecules in MapA adhesion.

Galectin-1 induces cell adhesion to the extracellular matrix and apoptosis of non-adherent human colon cancer Colo201 cells.

PubMed

Horiguchi, Natsuko; Arimoto, Kei-ichiro; Mizutani, Atsushi; Endo-Ichikawa, Yoko; Nakada, Hiroshi; Taketani, Shigeru

2003-12-01

To isolate cDNAs for molecules involved in cell adhesion to the extracellular matrix, expression cloning with non-adherent colon cancer Colo201 cells was carried out. Four positive clones were isolated and, when sequenced, one was found to be galectin-1, a beta-galactoside-binding protein. When cultured on fibronectin-, laminin-, and collagen-coated and non-coated dishes, the adherent galectin-1 cDNA-transfected Colo201 cells increased and spread somewhat. Immunofluorescence staining revealed that galectin-1 was expressed inside and outside of Colo201 cells. The adhesion was dependent on the carbohydrate-recognition domain of galectin-1 since lactose inhibited the adhesion and exogenously-added galectin-1 caused the adhesion. PD58059, an inhibitor of mitogen-activated protein kinase, or LY294002, a phosphoinositide 3-OH kinase inhibitor, decreased the adhesion. Furthermore, the expression of galectin-1 in Colo201 cells induced apoptotic cell death, while exogenously-added galectin-1 did not cause apoptosis. These results indicate that galectin-1 plays a role in both cell-matrix interactions and the inhibition of Colo201 cell proliferation, and suggest that galectin-1 expressed in cells could be associated with apoptosis.

A hot water extract of Curcuma longa inhibits adhesion molecule protein expression and monocyte adhesion to TNF-α-stimulated human endothelial cells.

PubMed

Kawasaki, Kengo; Muroyama, Koutarou; Yamamoto, Norio; Murosaki, Shinji

2015-01-01

The recruitment of arterial leukocytes to endothelial cells is an important step in the progression of various inflammatory diseases. Therefore, its modulation is thought to be a prospective target for the prevention or treatment of such diseases. Adhesion molecules on endothelial cells are induced by proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), and contribute to the recruitment of leukocytes. In the present study, we investigated the effect of hot water extract of Curcuma longa (WEC) on the protein expression of adhesion molecules, monocyte adhesion induced by TNF-α in human umbilical vascular endothelial cells (HUVECs). Treatment of HUVECs with WEC significantly suppressed both TNF-α-induced protein expression of adhesion molecules and monocyte adhesion. WEC also suppressed phosphorylation and degradation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) induced by TNF-α in HUVECs, suggesting that WEC inhibits the NF-κB signaling pathway.

Glucocorticoid-induced tumor necrosis factor receptor family-related ligand triggering upregulates vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 and promotes leukocyte adhesion.

PubMed

Lacal, Pedro Miguel; Petrillo, Maria Grazia; Ruffini, Federica; Muzi, Alessia; Bianchini, Rodolfo; Ronchetti, Simona; Migliorati, Graziella; Riccardi, Carlo; Graziani, Grazia; Nocentini, Giuseppe

2013-10-01

The interaction of glucocorticoid-induced tumor necrosis factor receptor-family related (GITR) protein with its ligand (GITRL) modulates different functions, including immune/inflammatory response. These effects are consequent to intracellular signals activated by both GITR and GITRL. Previous results have suggested that lack of GITR expression in GITR(-/-) mice decreases the number of leukocytes within inflamed tissues. We performed experiments to analyze whether the GITRL/GITR system modulates leukocyte adhesion and extravasation. For that purpose, we first evaluated the capability of murine splenocytes to adhere to endothelial cells (EC). Our results indicated that adhesion of GITR(-/-) splenocytes to EC was reduced as compared with wild-type cells, suggesting that GITR plays a role in adhesion and that this effect may be due to GITRL-GITR interaction. Moreover, adhesion was increased when EC were pretreated with an agonist GITR-Fc fusion protein, thus indicating that triggering of GITRL plays a role in adhesion by EC regulation. In a human in vitro model, the adhesion to human EC of HL-60 cells differentiated toward the monocytic lineage was increased by EC pretreatment with agonist GITR-Fc. Conversely, antagonistic anti-GITR and anti-GITRL Ab decreased adhesion, thus further indicating that GITRL triggering increases the EC capability to support leukocyte adhesion. EC treatment with GITR-Fc favored extravasation, as demonstrated by a transmigration assay. Notably, GITRL triggering increased intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression and anti-ICAM-1 and anti-VCAM-1 Abs reversed GITR-Fc effects. Our study demonstrates that GITRL triggering in EC increases leukocyte adhesion and transmigration, suggesting new anti-inflammatory therapeutic approaches based on inhibition of GITRL-GITR interaction.

Separation of integrin-dependent adhesion from morphological changes based on differential PLC specificities.

PubMed

Wooten, D K; Teague, T K; McIntyre, B W

1999-01-01

In normal lymphocytes an inside-out signal up-regulating integrin adhesion is followed by a ligand-mediated outside-in cell spreading signal. Protein kinase C (PKC) inhibition blocks lymphocyte adherence to and spreading on fibronectin. In contrast, putative PLC inhibitors yield distinct differences with respect to adhesion and morphology. The phosphatidylinositol-specific phospholipase C (PLC) inhibitor neomycin blocked spreading of CD3/CD28-activated T cells on fibronectin by disrupting adhesion. Furthermore, when an additional inside-out signal for fibronectin adhesion is unnecessary such as with HPB-ALL T leukemic or phorbol-myristate-acetate-treated normal T cells, neomycin treatment does not alter adhesion or morphology. However, the phosphatidylcholine-specific PLC inhibitor D609 abrogates cell spreading without affecting adhesion to fibronectin in these cells as well as the CD3/CD28-activated T cells. These results strongly suggest that inside-out signaling for the integrin alpha4beta1 in lymphocytes proceeds through phosphatidylinositol-specific PLC and PKC, whereas the outside-in signal utilizes phosphatidylcholine-specific PLC and PKC.

Modeling cell-substrate de-adhesion dynamics under fluid shear

NASA Astrophysics Data System (ADS)

Maan, Renu; Rani, Garima; Menon, Gautam I.; Pullarkat, Pramod A.

2018-07-01

Changes in cell-substrate adhesion are believed to signal the onset of cancer metastasis, but such changes must be quantified against background levels of intrinsic heterogeneity between cells. Variations in cell-substrate adhesion strengths can be probed through biophysical measurements of cell detachment from substrates upon the application of an external force. Here, we investigate, theoretically and experimentally, the detachment of cells adhered to substrates when these cells are subjected to fluid shear. We present a theoretical framework within which we calculate the fraction of detached cells as a function of shear stress for fast ramps as well as the decay in this fraction at fixed shear stress as a function of time. Using HEK and 3T3 fibroblast cells as experimental model systems, we extract characteristic force scales for cell adhesion as well as characteristic detachment times. We estimate force-scales of  ∼500 pN associated to a single focal contact, and characteristic time-scales of s representing cell-spread-area dependent mean first passage times to the detached state at intermediate values of the shear stress. Variations in adhesion across cell types are especially prominent when cell detachment is probed by applying a time-varying shear stress. These methods can be applied to characterizing changes in cell adhesion in a variety of contexts, including metastasis.

Matrix MetalloProteinases (MMPs) andTissue Inhibitors of MetalloProteinases (TIMPs): positive and negative regulators intumor cell adhesion

PubMed Central

Bourboulia, Dimitra; Stetler-Stevenson, William G.

2010-01-01

Cells adhere to one another and/or to matrices that surround them. Regulation of cell-cell (intercellular) and cell-matrix adhesion is tightly controlled in normal cells, however, defects in cell adhesion are common in the majority of humancancers. Multilateral communication among tumor cells with the extracellular matrix (ECM) and neighbor cells is accomplished through adhesion molecules, ECM components, proteolytic enzymes and their endogenous inhibitors. There is sufficient evidence to suggest that reduced adherence is a tumor cell propertyengaged during tumor progression. Tumor cells acquire the ability to change shape, detach and easily move through spaces disorganizing the normal tissue architecture. This property is due to changes in expression levels of adhesion molecules and/or due to elevated levels of secreted proteolytic enzymes, including matrix metalloproteinases (MMPs). Among other roles, MMPsdegrade the ECMand, therefore, prepare the path for tumor cells to migrate, invade and spread to distant secondary areas, where they form metastasis. Tissue Inhibitors of Metalloproteinases or TIMPs control MMP activities and, therefore, minimize matrix degradation. Both MMPs and TIMPs are involved in tissue remodeling and decisively regulate tumor cell progression including tumor angiogenesis. In this review, we describe and discuss data that support the important role of MMPs and TIMPs in cancer cell adhesion and tumor progression. PMID:20470890

Quantifying the effect of electric current on cell adhesion studied by single-cell force spectroscopy.

PubMed

Jaatinen, Leena; Young, Eleanore; Hyttinen, Jari; Vörös, János; Zambelli, Tomaso; Demkó, László

2016-03-20

This study presents the effect of external electric current on the cell adhesive and mechanical properties of the C2C12 mouse myoblast cell line. Changes in cell morphology, viability, cytoskeleton, and focal adhesion structure were studied by standard staining protocols, while single-cell force spectroscopy based on the fluidic force microscopy technology provided a rapid, serial quantification and detailed analysis of cell adhesion and its dynamics. The setup allowed measurements of adhesion forces up to the μN range, and total detachment distances over 40 μm. Force-distance curves have been fitted with a simple elastic model including a cell detachment protocol in order to estimate the Young's modulus of the cells, as well as to reveal changes in the dynamic properties as functions of the applied current dose. While the cell spreading area decreased monotonously with increasing current doses, small current doses resulted only in differences related to cell elasticity. Current doses above 11 As/m(2), however, initiated more drastic changes in cell morphology, viability, cellular structure, as well as in properties related to cell adhesion. The observed differences, eventually leading to cell death toward higher doses, might originate from both the decrease in pH and the generation of reactive oxygen species.

Mathematical modeling of cell adhesion in shear flow: application to targeted drug delivery in inflammation and cancer metastasis.

PubMed

Jadhav, Sameer; Eggleton, Charles D; Konstantopoulos, Konstantinos

2007-01-01

Cell adhesion plays a pivotal role in diverse biological processes that occur in the dynamic setting of the vasculature, including inflammation and cancer metastasis. Although complex, the naturally occurring processes that have evolved to allow for cell adhesion in the vasculature can be exploited to direct drug carriers to targeted cells and tissues. Fluid (blood) flow influences cell adhesion at the mesoscale by affecting the mechanical response of cell membrane, the intercellular contact area and collisional frequency, and at the nanoscale level by modulating the kinetics and mechanics of receptor-ligand interactions. Consequently, elucidating the molecular and biophysical nature of cell adhesion requires a multidisciplinary approach involving the synthesis of fundamentals from hydrodynamic flow, molecular kinetics and cell mechanics with biochemistry/molecular cell biology. To date, significant advances have been made in the identification and characterization of the critical cell adhesion molecules involved in inflammatory disorders, and, to a lesser degree, in cancer metastasis. Experimental work at the nanoscale level to determine the lifetime, interaction distance and strain responses of adhesion receptor-ligand bonds has been spurred by the advent of atomic force microscopy and biomolecular force probes, although our current knowledge in this area is far from complete. Micropipette aspiration assays along with theoretical frameworks have provided vital information on cell mechanics. Progress in each of the aforementioned research areas is key to the development of mathematical models of cell adhesion that incorporate the appropriate biological, kinetic and mechanical parameters that would lead to reliable qualitative and quantitative predictions. These multiscale mathematical models can be employed to predict optimal drug carrier-cell binding through isolated parameter studies and engineering optimization schemes, which will be essential for developing effective drug carriers for delivery of therapeutic agents to afflicted sites of the host.

Loss of the endothelial glycocalyx is associated with increased E-selectin mediated adhesion of lung tumour cells to the brain microvascular endothelium.

PubMed

Rai, Srijana; Nejadhamzeeigilani, Zaynab; Gutowski, Nicholas J; Whatmore, Jacqueline L

2015-09-25

Arrest of metastasising lung cancer cells to the brain microvasculature maybe mediated by interactions between ligands on circulating tumour cells and endothelial E-selectin adhesion molecules; a process likely to be regulated by the endothelial glycocalyx. Using human cerebral microvascular endothelial cells and non-small cell lung cancer (NSCLC) cell lines, we describe how factors secreted by NSCLC cells i.e. cystatin C, cathepsin L, insulin-like growth factor-binding protein 7 (IGFBP7), vascular endothelial growth factor (VEGF) and tumour necrosis factor-alpha (TNF-α), damage the glycocalyx and enhance initial contacts between lung tumour and cerebral endothelial cells. Endothelial cells were treated with tumour secreted-proteins or lung tumour conditioned medium (CM). Surface levels of E-selectin were quantified by ELISA. Adhesion of A549 and SK-MES-1 cells was examined under flow conditions (1 dyne/cm(2)). Alterations in the endothelial glycocalyx were quantified by binding of fluorescein isothiocyanate-linked wheat germ agglutinin (WGA-FITC). A549 and SK-MES-1 CM and secreted-proteins significantly enhanced endothelial surface E-selectin levels after 30 min and 4 h and tumour cell adhesion after 30 min, 4 and 24 h. Both coincided with significant glycocalyx degradation; A549 and SK-MES-1 CM removing 55 ± 12 % and 58 ± 18.7 % of WGA-FITC binding, respectively. Inhibition of E-selectin binding by monoclonal anti-E-selectin antibody completely attenuated tumour cell adhesion. These data suggest that metastasising lung cancer cells facilitate their own adhesion to the brain endothelium by secreting factors that damage the endothelial glycocalyx, resulting in exposure of the previously shielded adhesion molecules and engagement of the E-selectin-mediated adhesion axis.

The actions of volatile anaesthetics on synaptic transmission in the dentate gyrus.

PubMed Central

Richards, C D; White, A E

1975-01-01

1. The action of four volatile anaesthetics on the evoked synaptic potentials of in vitro preparations of the hippocampus were examined. 2. All four anaesthetics (ether, halothane, methoxyflurane and trichloroethylene) depressed the synaptic transmission between the perforant path and the granule cells at concentrations lower than those required to maintain anaesthesia in intact animals. 3. The population excitatory post-synaptic potential (e.p.s.p.) and massed discharge of the cortical cells (population spike) were depressed at concentrations of the anaesthetics lower than those required to depress the compound action potential of the perforant path nerve fibres. None of the anaesthetics studied increased the threshold depolarization required for granule cell discharge. Furthermore, frequency potentiation of the evoked cortical e.p.s.p.s was not impaired by any of the anaesthetics studied. 4. It is concluded that all four anaesthetics depress synaptic transmission in the dentate gyrus either by reducing the amount of transmitter released from each nerve terminal in response to an afferent volley, or by decreasing the sensitivity of the post-synaptic membrane to released transmitted or by both effects together. PMID:1202196

Targeting of adhesion molecules as a therapeutic strategy in multiple myeloma.

PubMed

Neri, Paola; Bahlis, Nizar J

2012-09-01

Multiple myeloma (MM) is a clonal disorder of plasma cells that remains, for the most part, incurable despite the advent of several novel therapeutic agents. Tumor cells in this disease are cradled within the bone marrow (BM) microenvironment by an array of adhesive interactions between the BM cellular residents, the surrounding extracellular matrix (ECM) components such as fibronectin (FN), laminin, vascular cell adhesion molecule-1 (VCAM-1), proteoglycans, collagens and hyaluronan, and a variety of adhesion molecules on the surface of MM cells including integrins, hyaluronan receptors (CD44 and RHAMM) and heparan sulfate proteoglycans. Several signaling responses are activated by these interactions, affecting the survival, proliferation and migration of MM cells. An important consequence of these direct adhesive interactions between the BM/ECM and MM cells is the development of drug resistance. This phenomenon is termed "cell adhesion-mediated drug resistance" (CAM-DR) and it is thought to be one of the major mechanisms by which MM cells escape the cytotoxic effects of therapeutic agents. This review will focus on the adhesion molecules involved in the cross-talk between MM cells and components of the BM microenvironment. The complex signaling networks downstream of these adhesive molecules mediated by direct ligand binding or inside-out soluble factors signaling will also be reviewed. Finally, novel therapeutic strategies targeting these molecules will be discussed. Identification of the mediators of MM-BM interaction is essential to understand MM biology and to elucidate novel therapeutic targets for this disease.

Ligand-induced adhesion to activated endothelium and to vascular cell adhesion molecule-1 in lymphocytes transfected with the N-formyl peptide receptor.

PubMed

Honda, S; Campbell, J J; Andrew, D P; Engelhardt, B; Butcher, B A; Warnock, R A; Ye, R D; Butcher, E C

1994-04-15

Binding of FMLP to the neutrophil N-formyl peptide receptor (FPR) transmits signals through pertussis toxin-sensitive G proteins triggering Ca2+ flux, superoxide production, granule exocytosis, and neutrophil aggregation and adhesion involving the beta 2 (CD18) integrins. Expression of the FPR in mouse fibroblasts or human kidney cells has been shown to confer an N-formyl peptide-inducible Ca2+ flux in transfectants. Here we demonstrate that the transfected receptor can also support ligand-induced alterations in cellular adhesion. We established stable transfectants of mouse L1-2 pre-B cells with cDNA for human FPR (L1-2 FPR cells). The transfectants bind N-formyl-Nle-Leu-Phe-Nle-Tyr-Lys-fluorescein with 1.4 x 10(5) sites per cell and a dissociation constant of 3.3 nM. Stimulation with FMLP induces a transient Ca2+ flux. FMLP also triggers adhesion of L1-2 FPR cells to TNF-alpha- or LPS-activated bEnd3 cells (mouse brain-derived endothelial cells) and to purified mouse VCAM-1. Binding is inhibited by Abs to VCAM-1 and to the alpha-chain of its lymphocyte receptor (the alpha 4 beta 1 integrin, VLA-4). Stimulation with FMLP does not induce a change in cell surface expression of alpha 4. Induced adhesion to VCAM-1 is rapid, detectable at the earliest times measurable (30 to 60 s after FMLP addition), and is inhibited by pertussis toxin. We conclude that FPR can mediate integrin activation not only in neutrophils but also in lymphocytes, and can trigger rapid adhesion via lymphocyte alpha 4 beta 1. The adhesion of lymphocytes is critical to their migration and targeting; our results suggest the possibility of manipulating adhesive responses through expression of chemoattractant receptors in lymphoid cells engineered for cellular therapy, allowing targeted adhesion and potentially migration in response to locally administered ligands.

Biological role of site-specific O-glycosylation in cell adhesion activity and phosphorylation of osteopontin.

PubMed

Oyama, Midori; Kariya, Yoshinobu; Kariya, Yukiko; Matsumoto, Kana; Kanno, Mayumi; Yamaguchi, Yoshiki; Hashimoto, Yasuhiro

2018-05-09

Osteopontin (OPN) is an extracellular glycosylated phosphoprotein that promotes cell adhesion by interacting with several integrin receptors. We previously reported that an OPN mutant lacking five O-glycosylation sites (Thr 134 /Thr 138 /Thr 143 /Thr 147 /Thr 152 ) in the threonine/proline-rich region increased cell adhesion activity and phosphorylation compared with the wild type. However, the role of O-glycosylation in cell adhesion activity and phosphorylation of OPN remains to be clarified. Here, we show that site-specific O-glycosylation in the threonine/proline-rich region of OPN affects its cell adhesion activity and phosphorylation independently and/or synergistically. Using site-directed mutagenesis, we found that OPN mutants with substitution sets of Thr 134 /Thr 138 or Thr 143 /Thr 147 /Thr 152 had decreased and increased cell adhesion activity, respectively. In contrast, the introduction of a single mutation into the O-glycosylation sites had no effect on OPN cell adhesion activity. An adhesion assay using function-blocking antibodies against αvβ3 and β1 integrins, as well as αvβ3 integrin-overexpressing A549 cells, revealed that site-specific O-glycosylation affected the association of OPN with the two integrins. Phosphorylation analyses using phos-tag and LC-MS/MS indicated that phosphorylation levels and sites were influenced by the O-glycosylation status, although the number of O-glycosylation sites was not correlated with the phosphorylation level in OPN. Furthermore, a correlation analysis between phosphorylation level and cell adhesion activity in OPN mutants with the site-specific O-glycosylation showed that they were not always correlated. These results provide conclusive evidence of a novel regulatory mechanism of cell adhesion activity and phosphorylation of OPN by site-specific O-glycosylation. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Development of assay system for immunoglobulin production regulatory factors using whole cell cultures of mouse splenocytes.

PubMed

Takasugi, M; Tamura, Y; Tachibana, H; Sugano, M; Yamada, K

2001-01-01

We tried to establish an assay system for screening and assessment of immunoregulatory factors using whole cell cultures of mouse splenocytes and found that splenic adhesive cells markedly increased immunogobulin (Ig) production of splenocytes. In the absence of adhesive cells, lipopolysaccharides, pokeweed mitogen, and phytohemagglutinin stimulated the production of IgA, IgG, and IgM in a class-dependent manner. Adhesive cells increased more markedly Ig production of splenocytes stimulated with these mitogens. When mouse splenocytes were cultured with milk proteins in the absence of adhesive cells, lactoferrin, beta-lactoglobulin, alpha-casein, and beta-casein stimulated IgA and IgG production. Adhesive cells increased IgA production of splenocytes stimulated with milk proteins, especially. These results suggest that the assay system is useful for assessment of Ig production-regulating factors.

A Novel Nectin-mediated Cell Adhesion Apparatus That Is Implicated in Prolactin Receptor Signaling for Mammary Gland Development*

PubMed Central

Kitayama, Midori; Mizutani, Kiyohito; Maruoka, Masahiro; Mandai, Kenji; Sakakibara, Shotaro; Ueda, Yuki; Komori, Takahide; Shimono, Yohei; Takai, Yoshimi

2016-01-01

Mammary gland development is induced by the actions of various hormones to form a structure consisting of collecting ducts and milk-secreting alveoli, which comprise two types of epithelial cells known as luminal and basal cells. These cells adhere to each other by cell adhesion apparatuses whose roles in hormone-dependent mammary gland development remain largely unknown. Here we identified a novel cell adhesion apparatus at the boundary between the luminal and basal cells in addition to desmosomes. This apparatus was formed by the trans-interaction between the cell adhesion molecules nectin-4 and nectin-1, which were expressed in the luminal and basal cells, respectively. Nectin-4 of this apparatus further cis-interacted with the prolactin receptor in the luminal cells to enhance the prolactin-induced prolactin receptor signaling for alveolar development with lactogenic differentiation. Thus, a novel nectin-mediated cell adhesion apparatus regulates the prolactin receptor signaling for mammary gland development. PMID:26757815

FRET measurements of cell-traction forces and nano-scale clustering of adhesion ligands varied by substrate stiffness.

PubMed

Kong, Hyun Joon; Polte, Thomas R; Alsberg, Eben; Mooney, David J

2005-03-22

The mechanical properties of cell adhesion substrates regulate cell phenotype, but the mechanism of this relation is currently unclear. It may involve the magnitude of traction force applied by the cell, and/or the ability of the cells to rearrange the cell adhesion molecules presented from the material. In this study, we describe a FRET technique that can be used to evaluate the mechanics of cell-material interactions at the molecular level and simultaneously quantify the cell-based nanoscale rearrangement of the material itself. We found that these events depended on the mechanical rigidity of the adhesion substrate. Furthermore, both the proliferation and differentiation of preosteoblasts (MC3T3-E1) correlated to the magnitude of force that cells generate to cluster the cell adhesion ligands, but not the extent of ligand clustering. Together, these data demonstrate the utility of FRET in analyzing cell-material interactions, and suggest that regulation of phenotype with substrate stiffness is related to alterations in cellular traction forces.

Role of flexural stiffness of leukocyte microvilli in adhesion dynamics

NASA Astrophysics Data System (ADS)

Wu, Tai-Hsien; Qi, Dewei

2018-03-01

Previous work reported that microvillus deformation has an important influence on dynamics of cell adhesion. However, the existing studies were limited to the extensional deformation of microvilli and did not consider the effects of their bending deformation on cell adhesion. This Rapid Communication investigates the effects of flexural stiffness of microvilli on the rolling process related to adhesion of leukocytes by using a lattice-Boltzmann lattice-spring method (LLM) combined with adhesive dynamics (AD) simulations. The simulation results reveal that the flexural stiffness of microvilli and their bending deformation have a profound effect on rolling velocity and adhesive forces. As the flexural stiffness of the microvilli decreases, their bending angles increase, resulting in an increase in the number of receptor-ligand bonds and adhesive bonding force and a decrease in the rolling velocity of leukocytes. The effects of flexural stiffness on deformation and adhesion represent crucial factors involved in cell adhesion.

Increased leukocyte adhesion to vascular endothelium in preeclampsia is inhibited by antioxidants.

PubMed

Ryu, Seongho; Huppmann, Alison R; Sambangi, Nirmala; Takacs, Peter; Kauma, Scott W

2007-04-01

To test the hypothesis that plasma from women with preeclampsia increases leukocyte adhesion to vascular endothelial cells and that antioxidants inhibit this effect. Plasma from 12 women with severe preeclampsia and 12 with normal pregnancy was tested in an in vitro leukocyte-endothelium adhesion assay in the presence or absence of vitamin E, vitamin C, or N-acetylcysteine. Preeclamptic plasma significantly increased monocyte (U937 cells) and T-cell (Jurkat) adhesion to human umbilical vein (HUVEC) and microvascular endothelial cells, compared with normal pregnant plasma. The antioxidants vitamin E, vitamin C, and N-acetylcysteine significantly inhibited monocyte adhesion to HUVEC in the presence of preeclamptic but not normal pregnant plasma. Increased adhesion in response to preeclamptic plasma was not mediated through a protein kinase C (PKC) mechanism, because the PKC inhibitor bisindolylmaleimide I had no effect on adhesion in the presence of preeclamptic plasma. Severe preeclampsia is associated with increased leukocyte-endothelium adhesion and clinically useful antioxidants can inhibit this effect.

Intermediate conductance calcium-activated potassium channels modulate summation of parallel fiber input in cerebellar Purkinje cells.

PubMed

Engbers, Jordan D T; Anderson, Dustin; Asmara, Hadhimulya; Rehak, Renata; Mehaffey, W Hamish; Hameed, Shahid; McKay, Bruce E; Kruskic, Mirna; Zamponi, Gerald W; Turner, Ray W

2012-02-14

Encoding sensory input requires the expression of postsynaptic ion channels to transform key features of afferent input to an appropriate pattern of spike output. Although Ca(2+)-activated K(+) channels are known to control spike frequency in central neurons, Ca(2+)-activated K(+) channels of intermediate conductance (KCa3.1) are believed to be restricted to peripheral neurons. We now report that cerebellar Purkinje cells express KCa3.1 channels, as evidenced through single-cell RT-PCR, immunocytochemistry, pharmacology, and single-channel recordings. Furthermore, KCa3.1 channels coimmunoprecipitate and interact with low voltage-activated Cav3.2 Ca(2+) channels at the nanodomain level to support a previously undescribed transient voltage- and Ca(2+)-dependent current. As a result, subthreshold parallel fiber excitatory postsynaptic potentials (EPSPs) activate Cav3 Ca(2+) influx to trigger a KCa3.1-mediated regulation of the EPSP and subsequent after-hyperpolarization. The Cav3-KCa3.1 complex provides powerful control over temporal summation of EPSPs, effectively suppressing low frequencies of parallel fiber input. KCa3.1 channels thus contribute to a high-pass filter that allows Purkinje cells to respond preferentially to high-frequency parallel fiber bursts characteristic of sensory input.

Annexin A6 contributes to the invasiveness of breast carcinoma cells by influencing the organization and localization of functional focal adhesions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sakwe, Amos M., E-mail: asakwe@mmc.edu; Koumangoye, Rainelli; Guillory, Bobby

2011-04-01

The interaction of annexin A6 (AnxA6) with membrane phospholipids and either specific extracellular matrix (ECM) components or F-actin suggests that it may influence cellular processes associated with rapid plasma membrane reorganization such as cell adhesion and motility. Here, we examined the putative roles of AnxA6 in adhesion-related cellular processes that contribute to breast cancer progression. We show that breast cancer cells secrete annexins via the exosomal pathway and that the secreted annexins are predominantly cell surface-associated. Depletion of AnxA6 in the invasive BT-549 breast cancer cells is accompanied by enhanced anchorage-independent cell growth but cell-cell cohesion, cell adhesion/spreading onto collagenmore » type IV or fetuin-A, cell motility and invasiveness were strongly inhibited. To explain the loss in adhesion/motility, we show that vinculin-based focal adhesions in the AnxA6-depleted BT-549 cells are elongated and randomly distributed. These focal contacts are also functionally defective because the activation of focal adhesion kinase and the phosphoinositide-3 kinase/Akt pathway were strongly inhibited while the MAP kinase pathway remained constitutively active. Compared with normal human breast tissues, reduced AnxA6 expression in breast carcinoma tissues correlates with enhanced cell proliferation. Together this suggests that reduced AnxA6 expression contributes to breast cancer progression by promoting the loss of functional cell-cell and/or cell-ECM contacts and anchorage-independent cell proliferation.« less

Why are enteric ganglia so small? Role of differential adhesion of enteric neurons and enteric neural crest cells.

PubMed Central

Rollo, Benjamin N.; Zhang, Dongcheng; Simkin, Johanna E.; Menheniott, Trevelyan R.; Newgreen, Donald F.

2015-01-01

The avian enteric nervous system (ENS) consists of a vast number of unusually small ganglia compared to other peripheral ganglia. Each ENS ganglion at mid-gestation has a core of neurons and a shell of mesenchymal precursor/glia-like enteric neural crest (ENC) cells. To study ENS cell ganglionation we isolated midgut ENS cells by HNK-1 fluorescence-activated cell sorting (FACS) from E5 and E8 quail embryos, and from E9 chick embryos. We performed cell-cell aggregation assays which revealed a developmentally regulated functional increase in ENS cell adhesive function, requiring both Ca 2+ -dependent and independent adhesion. This was consistent with N-cadherin and NCAM labelling. Neurons sorted to the core of aggregates, surrounded by outer ENC cells, showing that neurons had higher adhesion than ENC cells. The outer surface of aggregates became relatively non-adhesive, correlating with low levels of NCAM and N-cadherin on this surface of the outer non-neuronal ENC cells. Aggregation assays showed that ENS cells FACS selected for NCAM-high and enriched for enteric neurons formed larger and more coherent aggregates than unsorted ENS cells. In contrast, ENS cells of the NCAM-low FACS fraction formed small, disorganised aggregates.  This suggests a novel mechanism for control of ENS ganglion morphogenesis where i) differential adhesion of ENS neurons and ENC cells controls the core/shell ganglionic structure and ii) the ratio of neurons to ENC cells dictates the equilibrium ganglion size by generation of an outer non-adhesive surface. PMID:26064478

Leukocyte-inspired biodegradable particles that selectively and avidly adhere to inflamed endothelium in vitro and in vivo

NASA Astrophysics Data System (ADS)

Sakhalkar, Harshad S.; Dalal, Milind K.; Salem, Aliasger K.; Ansari, Ramin; Fu, Jie; Kiani, Mohammad F.; Kurjiaka, David T.; Hanes, Justin; Shakesheff, Kevin M.; Goetz, Douglas J.

2003-12-01

We exploited leukocyte-endothelial cell adhesion chemistry to generate biodegradable particles that exhibit highly selective accumulation on inflamed endothelium in vitro and in vivo. Leukocyte-endothelial cell adhesive particles exhibit up to 15-fold higher adhesion to inflamed endothelium, relative to noninflamed endothelium, under in vitro flow conditions similar to that present in blood vessels, a 6-fold higher adhesion to cytokine inflamed endothelium relative to non-cytokine-treated endothelium in vivo, and a 10-fold enhancement in adhesion to trauma-induced inflamed endothelium in vivo due to the addition of a targeting ligand. The leukocyte-inspired particles have adhesion efficiencies similar to that of leukocytes and were shown to target each of the major inducible endothelial cell adhesion molecules (E-selectin, P-selectin, vascular cell adhesion molecule 1, and intercellular adhesion molecule 1) that are up-regulated at sites of pathological inflammation. The potential for targeted drug delivery to inflamed endothelium has significant implications for the improved treatment of an array of pathologies, including cardiovascular disease, arthritis, inflammatory bowel disease, and cancer.

LGR5 receptor promotes cell-cell adhesion in stem cells and colon cancer cells via the IQGAP1-Rac1 pathway.

PubMed

Carmon, Kendra S; Gong, Xing; Yi, Jing; Wu, Ling; Thomas, Anthony; Moore, Catherine M; Masuho, Ikuo; Timson, David J; Martemyanov, Kirill A; Liu, Qingyun J

2017-09-08

Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) is a bona fide marker of adult stem cells in several epithelial tissues, most notably in the intestinal crypts, and is highly up-regulated in many colorectal, hepatocellular, and ovarian cancers. LGR5 activation by R-spondin (RSPO) ligands potentiates Wnt/β-catenin signaling in vitro ; however, deletion of LGR5 in stem cells has little or no effect on Wnt/β-catenin signaling or cell proliferation in vivo Remarkably, modulation of LGR5 expression has a major impact on the actin cytoskeletal structure and cell adhesion in the absence of RSPO stimulation, but the molecular mechanism is unclear. Here, we show that LGR5 interacts with IQ motif-containing GTPase-activating protein 1 (IQGAP1), an effector of Rac1/CDC42 GTPases, in the regulation of actin cytoskeleton dynamics and cell-cell adhesion. Specifically, LGR5 decreased levels of IQGAP1 phosphorylation at Ser-1441/1443, leading to increased binding of Rac1 to IQGAP1 and thus higher levels of cortical F-actin and enhanced cell-cell adhesion. LGR5 ablation in colon cancer cells and crypt stem cells resulted in loss of cortical F-actin, reduced cell-cell adhesion, and disrupted localization of adhesion-associated proteins. No evidence of LGR5 coupling to any of the four major subtypes of heterotrimeric G proteins was found. These findings suggest that LGR5 primarily functions via the IQGAP1-Rac1 pathway to strengthen cell-cell adhesion in normal adult crypt stem cells and colon cancer cells. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Quantitative measurements of intercellular adhesion between a macrophage and cancer cells using a cup-attached AFM chip.

PubMed

Kim, Hyonchol; Yamagishi, Ayana; Imaizumi, Miku; Onomura, Yui; Nagasaki, Akira; Miyagi, Yohei; Okada, Tomoko; Nakamura, Chikashi

2017-07-01

Intercellular adhesion between a macrophage and cancer cells was quantitatively measured using atomic force microscopy (AFM). Cup-shaped metal hemispheres were fabricated using polystyrene particles as a template, and a cup was attached to the apex of the AFM cantilever. The cup-attached AFM chip (cup-chip) approached a murine macrophage cell (J774.2), the cell was captured on the inner concave of the cup, and picked up by withdrawing the cup-chip from the substrate. The cell-attached chip was advanced towards a murine breast cancer cell (FP10SC2), and intercellular adhesion between the two cells was quantitatively measured. To compare cell adhesion strength, the work required to separate two adhered cells (separation work) was used as a parameter. Separation work was almost 2-fold larger between a J774.2 cell and FP10SC2 cell than between J774.2 cell and three additional different cancer cells (4T1E, MAT-LyLu, and U-2OS), two FP10SC2 cells, or two J774.2 cells. FP10SC2 was established from 4T1E as a highly metastatic cell line, indicates separation work increased as the malignancy of cancer cells became higher. One possible explanation of the strong adhesion of macrophages to cancer cells observed in this study is that the measurement condition mimicked the microenvironment of tumor-associated macrophages (TAMs) in vivo, and J774.2 cells strongly expressed CD204, which is a marker of TAMs. The results of the present study, which were obtained by measuring cell adhesion strength quantitatively, indicate that the fabricated cup-chip is a useful tool for measuring intercellular adhesion easily and quantitatively. Copyright © 2017 Elsevier B.V. All rights reserved.

Bacteria as living patchy colloids: Phenotypic heterogeneity in surface adhesion

PubMed Central

Hermes, Michiel; Schwarz-Linek, Jana; Poon, Wilson C. K.

2018-01-01

Understanding and controlling the surface adhesion of pathogenic bacteria is of urgent biomedical importance. However, many aspects of this process remain unclear (for example, microscopic details of the initial adhesion and possible variations between individual cells). Using a new high-throughput method, we identify and follow many single cells within a clonal population of Escherichia coli near a glass surface. We find strong phenotypic heterogeneities: A fraction of the cells remain in the free (planktonic) state, whereas others adhere with an adhesion strength that itself exhibits phenotypic heterogeneity. We explain our observations using a patchy colloid model; cells bind with localized, adhesive patches, and the strength of adhesion is determined by the number of patches: Nonadherers have no patches, weak adherers bind with a single patch only, and strong adherers bind via a single or multiple patches. We discuss possible implications of our results for controlling bacterial adhesion in biomedical and other applications. PMID:29719861

The role of cytoskeleton and adhesion proteins in the resistance to photodynamic therapy. Possible therapeutic interventions.

PubMed

Di Venosa, Gabriela; Perotti, Christian; Batlle, Alcira; Casas, Adriana

2015-08-01

It is known that Photodynamic Therapy (PDT) induces changes in the cytoskeleton, the cell shape, and the adhesion properties of tumour cells. In addition, these targets have also been demonstrated to be involved in the development of PDT resistance. The reversal of PDT resistance by manipulating the cell adhesion process to substrata has been out of reach. Even though the existence of cell adhesion-mediated PDT resistance has not been reported so far, it cannot be ruled out. In addition to its impact on the apoptotic response to photodamage, the cytoskeleton alterations are thought to be associated with the processes of metastasis and invasion after PDT. In this review, we will address the impact of photodamage on the microfilament and microtubule cytoskeleton components and its regulators on PDT-treated cells as well as on cell adhesion. We will also summarise the impact of PDT on the surviving and resistant cells and their metastatic potential. Possible strategies aimed at taking advantage of the changes induced by PDT on actin, tubulin and cell adhesion proteins by targeting these molecules will also be discussed.

Effects on in vitro and in vivo angiogenesis induced by small peptides carrying adhesion sequences.

PubMed

Conconi, Maria Teresa; Ghezzo, Francesca; Dettin, Monica; Urbani, Luca; Grandi, Claudio; Guidolin, Diego; Nico, Beatrice; Di Bello, Carlo; Ribatti, Domenico; Parnigotto, Pier Paolo

2010-07-01

It is well known that tumor growth is strictly dependent on neo-vessel formation inside the tumor mass and that cell adhesion is required to allow EC proliferation and migration inside the tumor. In this work, we have evaluated the in vitro and in vivo effects on angiogenesis of some peptides, originally designed to promote cell adhesion on biomaterials, containing RGD motif mediating cell adhesion via integrin receptors [RGD, GRGDSPK, and (GRGDSP)(4)K] or the heparin-binding sequence of human vitronectin that interacts with HSPGs [HVP(351-359)]. Cell adhesion, proliferation, migration, and capillary-like tube formation in Matrigel were determined on HUVECs, whereas the effects on in vivo angiogenesis were evaluated using the CAM assay. (GRGDSP)(4)K linear sequence inhibited cell adhesion, decreased cell proliferation, migration and morphogenesis in Matrigel, and induced anti-angiogenic responses on CAM at higher degree than that determined after incubation with RGD or GRGDSPK. Moreover, it counteracted both in vitro and in vivo the pro-angiogenic effects induced by the Fibroblast growth factor (FGF-2). On the other hand, HVP was not able to affect cell adhesion and appeared less effective than (GRGDSP)(4)K. Our data indicate that the activity of RGD-containing peptides is related to their adhesive properties, and their effects are modulated by the number of cell adhesion motifs and the aminoacidic residues next to these sequences. The anti-angiogenic properties of (GRGDSP)(4)K seem to depend on its interaction with integrins, whereas the effects of HVP may be partially due to an impairment of HSPGs/FGF-2.

Up-regulation of Paxillin and Focal Adhesion Signaling follows Dystroglycan Complex deletions and promotes a Hypertensive State of Differentiation

PubMed Central

Sen, Shamik; Tewari, Manorama; Zajac, Allison; Barton, Elisabeth; Sweeney, H. Lee; Discher, Dennis E.

2010-01-01

Anchorage to matrix is mediated for many cells not only by integrin-based focal adhesions but also by a parallel assembly of integral and peripheral membrane proteins known as the Dystroglycan Complex. Deficiencies in either dystrophin (mdx mice) or γ-sarcoglycan (γSG−/− mice) components of the Dystroglycan Complex lead to upregulation of numerous focal adhesion proteins, and the phosphoprotein paxillin proves to be among the most prominent. In mdx muscle, paxillin-Y31 and Y118 are both hyper-phosphorylated as are key sites in focal adhesion kinase (FAK) and the stretch-stimulatable pro-survival MAPK pathway, whereas γSG−/− muscle exhibits more erratic hyper-phosphorylation. In cultured myotubes, cell tension generated by myosin-II appears required for localization of paxillin to adhesions while vinculin appears more stably integrated. Over-expression of wild-type (WT) paxillin has no obvious effect on focal adhesion density or the physical strength of adhesion, but WT and a Y118F mutant promote contractile sarcomere formation whereas a Y31F mutant shows no effect, implicating Y31 in striation. Self-peeling of cells as well as Atomic Force Microscopy (AFM) probing of cells with or without myosin II inhibition indicate an increase in cell tension within paxillin-overexpressing cells. However, prednisolone, a first-line glucocorticoid for muscular dystrophies, decreases cell tension without affecting paxillin at adhesions, suggesting a non-linear relationship between paxillin and cell tension. Hypertension that results from upregulation of integrin adhesions is thus a natural and treatable outcome of dystroglycan complex down-regulation. PMID:20663583

Designing a Binding Interface for Control of Cancer Cell Adhesion via 3D Topography and Metabolic Oligosaccharide Engineering

PubMed Central

Du, Jian; Che, Pao-Lin; Wang, Zhi-Yun; Aich, Udayanath; Yarema, Kevin J.

2011-01-01

This study combines metabolic oligosaccharide engineering (MOE), a technology where the glycocalyx of living cells is endowed with chemical features not normally found in sugars, with custom-designed three dimensional biomaterial substrates to enhance the adhesion of cancer cells and control their morphology and gene expression. Specifically, Ac5ManNTGc, a thiol-bearing analogue of N-acetyl-d-mannosamine (ManNAc) was used to introduce thiolated sialic acids into the glycocalyx of human Jurkat T-lymphoma derived cells. In parallel 2D films and 3D electrospun nanofibrous scaffolds were prepared from polyethersulfone (PES) and (as controls) left unmodified or aminated. Alternately, the materials were malemided or gold-coated to provide bioorthogonal binding partners for the thiol groups newly expressed on the cell surface. Cell attachment was modulated by both the topography of the substrate surface and by the chemical compatibility of the binding interface between the cell and the substrate; a substantial increase in binding for normally non-adhesive Jurkat line for 3D scaffold compared to 2D surfaces with an added degree of adhesion resulting from chemoselective binding to malemidede-derivatived or gold-coated surfaces. In addition, the morphology of the cells attached to the 3D scaffolds via MOE-mediated adhesion was dramatically altered and the expression of genes involved in cell adhesion changed in a time-dependent manner. This study showed that cell adhesion could be enhanced, gene expression modulated, and cell fate controlled by introducing the 3D topograhical cues into the growth substrate and by creating a glycoengineered binding interface where the chemistry of both the cell surface and biomaterials scaffold was controlled to facilitate a new mode of carbohydrate-mediated adhesion. PMID:21549424

Mycophenolate mofetil increases adhesion capacity of tumor cells in vitro.

PubMed

Blaheta, Roman A; Bogossian, Harilaos; Beecken, Wolf-Dietrich; Jonas, Dietger; Hasenberg, Christoph; Makarevic, Jasmina; Ogbomo, Henry; Bechstein, Wolf O; Oppermann, Elsie; Leckel, Kerstin; Cinatl, Jindrich

2003-12-27

The immunosuppressive drug mycophenolate mofetil (MMF) reduces expression of the heterophilic binding elements intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 and thereby prevents attachment of alloactivated leukocytes to donor endothelium. The authors speculated that MMF might further diminish receptors of the immunoglobulin superfamily which, however, act as homophilic binding elements. Because decrease of homophilic adhesion receptors correlates with tumor dissemination and metastasis, MMF could trigger development or recurrence of neoplastic tumors. The authors analyzed the influence of MMF on homotypic adhesion receptors and its consequence for tumor cell attachment to an endothelial cell monolayer. Neuroblastoma (NB) cells, which self-aggregate by means of the homophilic-binding element neural cell adhesion molecule (NCAM), were used. Effects of MMF on the 140- and 180-kDa NCAM isoforms were investigated quantitatively by flow cytometry, Western blot, and reverse-transcriptase (RT) polymerase chain reaction (PCR). The relevance of NCAM for tumor cell binding was proven by treating NB with NCAM antisense oligonucleotides. MMF profoundly increased the number of adherent NB cells, with a maximum effect at 0.1 microM, compared with controls. Decrease of NCAM on the cell surface was detected by flow cytometry. Western blot and RT-PCR demonstrated reduced protein and RNA levels of the 140- and 180-kDa isoforms. Treatment of NB cells with NCAM antisense oligonucleotides showed that reduced NCAM expression leads to enhanced tumor cell adhesion. MMF decreases NCAM receptors, which is associated with enhanced tumor cell invasiveness. The authors conclude that an MMF-based immunosuppressive regimen might increase the risk of tumor metastasis if this process is predominantly conveyed by means of homophilic adhesion proteins.

CD44 in cancer progression: adhesion, migration and growth regulation.

PubMed

Marhaba, R; Zöller, M

2004-03-01

It is well established that the large array of functions that a tumour cell has to fulfil to settle as a metastasis in a distant organ requires cooperative activities between the tumour and the surrounding tissue and that several classes of molecules are involved, such as cell-cell and cell-matrix adhesion molecules and matrix degrading enzymes, to name only a few. Furthermore, metastasis formation requires concerted activities between tumour cells and surrounding cells as well as matrix elements and possibly concerted activities between individual molecules of the tumour cell itself. Adhesion molecules have originally been thought to be essential for the formation of multicellular organisms and to tether cells to the extracellular matrix or to neighbouring cells. CD44 transmembrane glycoproteins belong to the families of adhesion molecules and have originally been described to mediate lymphocyte homing to peripheral lymphoid tissues. It was soon recognized that the molecules, under selective conditions, may suffice to initiate metastatic spread of tumour cells. The question remained as to how a single adhesion molecule can fulfil that task. This review outlines that adhesion is by no means a passive task. Rather, ligand binding, as exemplified for CD44 and other similar adhesion molecules, initiates a cascade of events that can be started by adherence to the extracellular matrix. This leads to activation of the molecule itself, binding to additional ligands, such as growth factors and matrix degrading enzymes, complex formation with additional transmembrane molecules and association with cytoskeletal elements and signal transducing molecules. Thus, through the interplay of CD44 with its ligands and associating molecules CD44 modulates adhesiveness, motility, matrix degradation, proliferation and cell survival, features that together may well allow a tumour cell to proceed through all steps of the metastatic cascade.

Curcumin inhibits activation induced by urban particulate material or titanium dioxide nanoparticles in primary human endothelial cells

PubMed Central

Montiel-Dávalos, Angélica; Silva Sánchez, Guadalupe Jazmin; Huerta-García, Elizabeth; Rueda-Romero, Cristhiam; Soca Chafre, Giovanny; Mitre-Aguilar, Irma B.; Alfaro-Moreno, Ernesto; Pedraza-Chaverri, José

2017-01-01

Curcumin has protective effects against toxic agents and shows preventive properties for various diseases. Particulate material with an aerodynamic diameter of ≤10 μm (PM10) and titanium dioxide nanoparticles (TiO2-NPs) induce endothelial dysfunction and activation. We explored whether curcumin is able to attenuate different events related to endothelial activation. This includes adhesion, expression of adhesion molecules and oxidative stress induced by PM10 and TiO2-NPs. Human umbilical vein endothelial cells (HUVEC) were treated with 1, 10 and 100 μM curcumin for 1 h and then exposed to PM10 at 3 μg/cm2 or TiO2-NPs at 10 μg/cm2. Cell adhesion was evaluated by co-culture with U937 human myelomonocytic cells. Adhesion molecules expression was measured by flow cytometry after 3 or 24 h of exposure. Oxidative stress was determined by 2,7-dichlorodihydrofluorescein (H2DCF) oxidation. PM10 and TiO2-NPs induced the adhesion of U937 cells and the expression of E- and P-selectins, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and platelet-endothelial cell adhesion molecule-1 (PECAM-1). The expression of E- and P-selectins matched the adhesion of monocytes to HUVEC after 3 h. In HUVEC treated with 1 or 10 μM curcumin, the expression of adhesion molecules and monocytes adhesion was significantly diminished. Curcumin also partially reduced the H2DCF oxidation induced by PM10 and TiO2-NPs. Our results suggest an anti-inflammatory and antioxidant role by curcumin attenuating the activation caused on endothelial cells by exposure to particles. Therefore, curcumin could be useful in the treatment of diseases where an inflammatory process and endothelial activation are involved. PMID:29244817

Marine Bacterial Polysaccharide EPS11 Inhibits Cancer Cell Growth via Blocking Cell Adhesion and Stimulating Anoikis

PubMed Central

Cao, Ruobing; Jin, Weihua; Shan, Yeqi; Wang, Ju; Liu, Ge; Kuang, Shan

2018-01-01

Tumor cells that acquire metastatic potential have developed resistance to anoikis, a cell death process, after detachment from their primary site to the second organ. In this study, we investigated the molecular mechanisms of a novel marine bacterial polysaccharide EPS11 which exerts its cytotoxic effects through affecting cancer cell adhesion and anoikis. Firstly, we found that EPS11 could significantly affect cell proliferation and block cell adhesion in A549 cells. We further demonstrated that the expression of several cell adhesion associated proteins is downregulated and the filiform structures of cancer cells are destroyed after EPS11 treatment. Interestingly, the destruction of filiform structures in A549 cells by EPS11 is in a dose-dependent manner, and the inhibitory tendency is very consistent with that observed in the cell adhesion assay, which confirms that filiform structures play important roles in modulating cell adhesion. Moreover, we showed that EPS11 induces apoptosis of A549 cells through stimulating βIII-tubulin associated anoikis: (i) EPS11 inhibits the expression of βIII-tubulin in both transcription and translation levels; and (ii) EPS11 treatment dramatically decreases the phosphorylation of protein kinase B (PKB or AKT), a critical downstream effector of βIII-tubulin. Importantly, EPS11 evidently inhibits the growth of A549-derived tumor xenografts in vivo. Thus, our results suggest that EPS11 may be a potential candidate for human non-small cell lung carcinoma treatment via blocking filiform structure mediated adhesion and stimulating βIII-tubulin associated anoikis. PMID:29518055

Material- and feature-dependent effects on cell adhesion to micro injection moulded medical polymers.

PubMed

Choi, Seong Ying; Habimana, Olivier; Flood, Peter; Reynaud, Emmanuel G; Rodriguez, Brian J; Zhang, Nan; Casey, Eoin; Gilchrist, Michael D

2016-09-01

Two polymers, polymethylmethacrylate (PMMA) and cyclic olefin copolymer (COC), containing a range of nano- to micron- roughness surfaces (Ra 0.01, 0.1, 0.4, 1.0, 2.0, 3.2 and 5.0μm) were fabricated using electrical discharge machining (EDM) and replicated using micro injection moulding (μIM). Polymer samples were characterized using optical profilometry, atomic force microscopy (AFM) and water surface contact angle. Cell adhesion tests were carried out using bacterial Pseudomonas fluorescens and mammalian Madin-Darby Canine Kidney (MDCK) cells to determine the effect of surface hydrophobicity, surface roughness and stiffness. It is found that there are features which gave insignificant differences (feature-dependent effect) in cell adhesion, albeit a significant difference in the physicochemical properties (material-dependent effect) of substrata. In bacterial cell adhesion, the strongest feature-dependence is found at Ra 0.4μm surfaces, with material-dependent effects strongest at Ra 0.01μm. Ra 0.1μm surfaces exhibited strongest feature-dependent effects and Ra 5.0μm has strongest material-dependent effects on mammalian cell adhesion. Bacterial cell adhesion is found to be favourable to hydrophobic surfaces (COC), with the lowest adhesion at Ra 0.4μm for both materials. Mammalian cell adhesion is lowest in Ra 0.1μm and highest in Ra 1.0μm, and generally favours hydrophilic surfaces (PMMA). These findings can be used as a basis for developing medical implants or microfluidic devices using micro injection moulding for diagnostic purposes, by tuning the cell adhesion on different areas containing different surface roughnesses on the diagnostic microfluidic devices or medical implants. Copyright © 2016 Elsevier B.V. All rights reserved.

Tumor necrosis factor alpha (TNF-alpha)-induced cell adhesion to human endothelial cells is under dominant control of one TNF receptor type, TNF-R55

PubMed Central

1993-01-01

Tumor necrosis factor alpha (TNF-alpha) is a pleiotropic cytokine triggering cell responses through two distinct membrane receptors. Stimulation of leukocyte adhesion to the endothelium is one of the many TNF-alpha activities and is explained by the upregulation of adhesion molecules on the endothelial cell surface. Human umbilical vein endothelial cells (HUVEC) were isolated, cultured, and demonstrated to express both TNF receptor types, TNF-R55 and TNF-R75. Cell adhesion to HUVEC was studied using the HL60, U937, and MOLT-4 cell lines. HUVEC were activated by either TNF-alpha, binding to both TNF-R55 and TNF- R75, and by receptor type-specific agonists, binding exclusively to TNF- R55 or to TNF-R75. The TNF-alpha-induced cell adhesion to HUVEC was found to be controlled almost exclusively by TNF-R55. This finding correlated with the exclusive activity of TNF-R55 in the TNF-alpha- dependent regulation of the expression of the intercellular adhesion molecule type 1 (ICAM-1), E-selectin, and vascular cell adhesion molecule type 1 (VCAM-1). The CD44 adhesion molecule in HUVEC was also found to be upregulated through TNF-R55. However, both TNF-R55 and TNF- R75 upregulate alpha 2 integrin expression in HUVEC. The predominant role of TNF-R55 in TNF-alpha-induced adhesion in HUVEC may correlate with its specific control of NF-kappa B activation, since kappa B elements are known to be present in ICAM-1, E-selectin, and VCAM-1 gene regulatory sequences. PMID:8386742

Desmoglein 3 regulates membrane trafficking of cadherins, an implication in cell-cell adhesion

PubMed Central

Moftah, Hanan; Dias, Kasuni; Apu, Ehsanul Hoque; Liu, Li; Uttagomol, Jutamas; Bergmeier, Lesley; Kermorgant, Stephanie; Wan, Hong

2017-01-01

ABSTRACT E-cadherin mediated cell-cell adhesion plays a critical role in epithelial cell polarization and morphogenesis. Our recent studies suggest that the desmosomal cadherin, desmoglein 3 (Dsg3) cross talks with E-cadherin and regulates its adhesive function in differentiating keratinocytes. However, the underlying mechanism remains not fully elucidated. Since E-cadherin trafficking has been recognized to be a central determinant in cell-cell adhesion and homeostasis we hypothesize that Dsg3 may play a role in regulating E-cadherin trafficking and hence the cell-cell adhesion. Here we investigated this hypothesis in cells with loss of Dsg3 function through RNAi mediated Dsg3 knockdown or the stable expression of the truncated mutant Dsg3ΔC. Our results showed that loss of Dsg3 resulted in compromised cell-cell adhesion and reduction of adherens junction and desmosome protein expression as well as the cortical F-actin formation. As a consequence, cells failed to polarize but instead displayed aberrant cell flattening. Furthermore, retardation of E-cadherin internalization and recycling was consistently observed in these cells during the process of calcium induced junction assembling. In contrast, enhanced cadherin endocytosis was detected in cells with overexpression of Dsg3 compared to control cells. Importantly, this altered cadherin trafficking was found to be coincided with the reduced expression and activity of Rab proteins, including Rab5, Rab7 and Rab11 which are known to be involved in E-cadherin trafficking. Taken together, our findings suggest that Dsg3 functions as a key in cell-cell adhesion through at least a mechanism of regulating E-cadherin membrane trafficking. PMID:27254775

Polymer surface functionalities that control human embryoid body cell adhesion revealed by high throughput surface characterization of combinatorial material microarrays

PubMed Central

Yang, Jing; Mei, Ying; Hook, Andrew L.; Taylor, Michael; Urquhart, Andrew J.; Bogatyrev, Said R.; Langer, Robert; Anderson, Daniel G.; Davies, Martyn C.; Alexander, Morgan R.

2010-01-01

High throughput materials discovery using combinatorial polymer microarrays to screen for new biomaterials with new and improved function is established as a powerful strategy. Here we combine this screening approach with high throughput surface characterisation (HT-SC) to identify surface structure-function relationships. We explore how this combination can help to identify surface chemical moieties that control protein adsorption and subsequent cellular response. The adhesion of human embryoid body (hEB) cells to a large number (496) of different acrylate polymers synthesized in a microarray format is screened using a high throughput procedure. To determine the role of the polymer surface properties on hEB cell adhesion, detailed HT-SC of these acrylate polymers is carried out using time of flight secondary ion mass spectrometry (ToF SIMS), x-ray photoelectron spectroscopy (XPS), pico litre drop sessile water contact angle (WCA) measurement and atomic force microscopy (AFM). A structure-function relationship is identified between the ToF SIMS analysis of the surface chemistry after a fibronectin (Fn) pre-conditioning step and the cell adhesion to each spot using the multivariate analysis technique partial least squares (PLS) regression. Secondary ions indicative of the adsorbed Fn correlate with increased cell adhesion whereas glycol and other functionalities from the polymers are identified that reduce cell adhesion. Furthermore, a strong relationship between the ToF SIMS spectra of bare polymers and the cell adhesion to each spot is identified using PLS regression. This identifies a role for both the surface chemistry of the bare polymer and the pre-adsorbed Fn, as-represented in the ToF SIMS spectra, in controlling cellular adhesion. In contrast, no relationship is found between cell adhesion and wettability, surface roughness, elemental or functional surface composition. The correlation between ToF SIMS data of the surfaces and the cell adhesion demonstrates the ability of identifying surface moieties that control protein adsorption and subsequent cell adhesion using ToF SIMS and multivariate analysis. PMID:20832108

Single Cell Force Spectroscopy for Quantification of Cellular Adhesion on Surfaces

NASA Astrophysics Data System (ADS)

Christenson, Wayne B.

Cell adhesion is an important aspect of many biological processes. The atomic force microscope (AFM) has made it possible to quantify the forces involved in cellular adhesion using a technique called single cell force spectroscopy (SCFS). AFM based SCFS offers versatile control over experimental conditions for probing directly the interaction between specific cell types and specific proteins, surfaces, or other cells. Transmembrane integrins are the primary proteins involved in cellular adhesion to the extra cellular matix (ECM). One of the chief integrins involved in the adhesion of leukocyte cells is alpha Mbeta2 (Mac-1). The experiments in this dissertation quantify the adhesion of Mac-1 expressing human embryonic kidney (HEK Mac-1), platelets, and neutrophils cells on substrates with different concentrations of fibrinogen and on fibrin gels and multi-layered fibrinogen coated fibrin gels. It was shown that multi-layered fibrinogen reduces the adhesion force of these cells considerably. A novel method was developed as part of this research combining total internal reflection microscopy (TIRFM) with SCFS allowing for optical microscopy of HEK Mac-1 cells interacting with bovine serum albumin (BSA) coated glass after interacting with multi-layered fibrinogen. HEK Mac-1 cells are able to remove fibrinogen molecules from the multi-layered fibrinogen matrix. An analysis methodology for quantifying the kinetic parameters of integrin-ligand interactions from SCFS experiments is proposed, and the kinetic parameters of the Mac-1 fibrinogen bond are quantified. Additional SCFS experiments quantify the adhesion of macrophages and HEK Mac-1 cells on functionalized glass surfaces and normal glass surfaces. Both cell types show highest adhesion on a novel functionalized glass surface that was prepared to induce macrophage fusion. These experiments demonstrate the versatility of AFM based SCFS, and how it can be applied to address many questions in cellular biology offering quantitative insights.

Binding constant of cell adhesion receptors and substrate-immobilized ligands depends on the distribution of ligands

NASA Astrophysics Data System (ADS)

Li, Long; Hu, Jinglei; Xu, Guangkui; Song, Fan

2018-01-01

Cell-cell adhesion and the adhesion of cells to tissues and extracellular matrix, which are pivotal for immune response, tissue development, and cell locomotion, depend sensitively on the binding constant of receptor and ligand molecules anchored on the apposing surfaces. An important question remains of whether the immobilization of ligands affects the affinity of binding with cell adhesion receptors. We have investigated the adhesion of multicomponent membranes to a flat substrate coated with immobile ligands using Monte Carlo simulations of a statistical mesoscopic model with biologically relevant parameters. We find that the binding of the adhesion receptors to ligands immobilized on the substrate is strongly affected by the ligand distribution. In the case of ligand clusters, the receptor-ligand binding constant can be significantly enhanced due to the less translational entropy loss of lipid-raft domains in the model cell membranes upon the formation of additional complexes. For ligands randomly or uniformly immobilized on the substrate, the binding constant is rather decreased since the receptors localized in lipid-raft domains have to pay an energetic penalty in order to bind ligands. Our findings help to understand why cell-substrate adhesion experiments for measuring the impact of lipid rafts on the receptor-ligand interactions led to contradictory results.

Basal Lamina Mimetic Nanofibrous Peptide Networks for Skeletal Myogenesis

NASA Astrophysics Data System (ADS)

Yasa, I. Ceren; Gunduz, Nuray; Kilinc, Murat; Guler, Mustafa O.; Tekinay, Ayse B.

2015-11-01

Extracellular matrix (ECM) is crucial for the coordination and regulation of cell adhesion, recruitment, differentiation and death. Therefore, equilibrium between cell-cell and cell-matrix interactions and matrix-associated signals are important for the normal functioning of cells, as well as for regeneration. In this work, we describe importance of adhesive signals for myoblast cells’ growth and differentiation by generating a novel ECM mimetic peptide nanofiber scaffold system. We show that not only structure but also composition of bioactive signals are important for cell adhesion, growth and differentiation by mimicking the compositional and structural properties of native skeletal muscle basal lamina. We conjugated laminin-derived integrin binding peptide sequence, “IKVAV”, and fibronectin-derived well known adhesive sequence, “RGD”, into peptide nanostructures to provide adhesive and myogenic cues on a nanofibrous morphology. The myogenic and adhesive signals exhibited a synergistic effect on model myoblasts, C2C12 cells. Our results showed that self-assembled peptide nanofibers presenting laminin derived epitopes support adhesion, growth and proliferation of the cells and significantly promote the expression of skeletal muscle-specific marker genes. The functional peptide nanofibers used in this study present a biocompatible and biodegradable microenvironment, which is capable of supporting the growth and differentiation of C2C12 myoblasts into myotubes.

Cytotoxic-T-lymphocyte antigen 4 receptor signaling for lymphocyte adhesion is mediated by C3G and Rap1.

PubMed

Kloog, Yoel; Mor, Adam

2014-03-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders.

Basal Lamina Mimetic Nanofibrous Peptide Networks for Skeletal Myogenesis

PubMed Central

Yasa, I. Ceren; Gunduz, Nuray; Kilinc, Murat; Guler, Mustafa O.; Tekinay, Ayse B.

2015-01-01

Extracellular matrix (ECM) is crucial for the coordination and regulation of cell adhesion, recruitment, differentiation and death. Therefore, equilibrium between cell-cell and cell-matrix interactions and matrix-associated signals are important for the normal functioning of cells, as well as for regeneration. In this work, we describe importance of adhesive signals for myoblast cells’ growth and differentiation by generating a novel ECM mimetic peptide nanofiber scaffold system. We show that not only structure but also composition of bioactive signals are important for cell adhesion, growth and differentiation by mimicking the compositional and structural properties of native skeletal muscle basal lamina. We conjugated laminin-derived integrin binding peptide sequence, “IKVAV”, and fibronectin-derived well known adhesive sequence, “RGD”, into peptide nanostructures to provide adhesive and myogenic cues on a nanofibrous morphology. The myogenic and adhesive signals exhibited a synergistic effect on model myoblasts, C2C12 cells. Our results showed that self-assembled peptide nanofibers presenting laminin derived epitopes support adhesion, growth and proliferation of the cells and significantly promote the expression of skeletal muscle-specific marker genes. The functional peptide nanofibers used in this study present a biocompatible and biodegradable microenvironment, which is capable of supporting the growth and differentiation of C2C12 myoblasts into myotubes. PMID:26555958

Osteoblast adhesion to orthopaedic implant alloys: Effects of cell adhesion molecules and diamond-like carbon coating

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kornu, R.; Kelly, M.A.; Smith, R.L.

1996-11-01

In total joint arthroplasty, long-term outcomes depend in part on the biocompatibility of implant alloys. This study analyzed effects of surface finish and diamond-like carbon coating on osteoblast cell adhesion to polished titanium-aluminum-vanadium and polished or grit-blasted cobalt-chromium-molybdenum alloys. Osteoblast binding was tested in the presence and absence of the cell adhesion proteins fibronectin, laminin, fibrinogen, and vitronectin and was quantified by measurement of DNA content. Although adherence occurred in serum-free medium, maximal osteoblast binding required serum and was similar for titanium and cobalt alloys at 2 and 12 hours. With the grit-blasted cobalt alloy, cell binding was reduced 48%more » (p < 0.05) by 24 hours. Coating the alloys with diamond-like carbon did not alter osteoblast adhesion, whereas fibronectin pretreatment increased cell binding 2.6-fold (p < 0.05). In contrast, fibrinogen, vitronectin, and laminin did not enhance cell adhesion. These results support the hypothesis that cell adhesion proteins can modify cell binding to orthopaedic alloys. Although osteoblast binding was not affected by the presence of diamond-like carbon, this coating substance may influence other longer term processes, such as bone formation, and deserves further study. 40 refs., 4 figs.« less

Mena binds α5 integrin directly and modulates α5β1 function

PubMed Central

Riquelme, Daisy; Hughes-Alford, Shannon K.; Tadros, Jenny; Rudina, Shireen S.; O.Hynes, Richard; Lauffenburger, Douglas

2012-01-01

Mena is an Ena/VASP family actin regulator with roles in cell migration, chemotaxis, cell–cell adhesion, tumor cell invasion, and metastasis. Although enriched in focal adhesions, Mena has no established function within these structures. We find that Mena forms an adhesion-regulated complex with α5β1 integrin, a fibronectin receptor involved in cell adhesion, motility, fibronectin fibrillogenesis, signaling, and growth factor receptor trafficking. Mena bound directly to the carboxy-terminal portion of the α5 cytoplasmic tail via a 91-residue region containing 13 five-residue “LERER” repeats. In fibroblasts, the Mena–α5 complex was required for “outside-in” α5β1 functions, including normal phosphorylation of FAK and paxillin and formation of fibrillar adhesions. It also supported fibrillogenesis and cell spreading and controlled cell migration speed. Thus, fibroblasts require Mena for multiple α5β1-dependent processes involving bidirectional interactions between the extracellular matrix and cytoplasmic focal adhesion proteins. PMID:22908313

Extracellular galectin-1 enhances adhesion to and invasion of oral epithelial cells by Porphyromonas gingivalis.

PubMed

Tamai, Riyoko; Kobayashi-Sakamoto, Michiyo; Kiyoura, Yusuke

2018-03-15

Galectin-1 and galectin-3 are C-type lectin receptors that bind to lipopolysaccharide in the cell wall of gram-negative bacteria. In this study, we investigated the effects of galectin-1 and galectin-3 on adhesion to and invasion of the human gingival epithelial cell line Ca9-22 by Porphyromonas gingivalis, a periodontal pathogenic gram-negative bacterium. Recombinant galectin-1, but not galectin-3, enhanced P. gingivalis adhesion and invasion, although both galectins bound similarly to P. gingivalis. Flow cytometry also revealed that Ca9-22 cells express low levels of galectin-1 and moderate levels of galectin-3. Ca9-22 cells in which galectin-3 was knocked-down did not exhibit enhanced P. gingivalis adhesion and invasion. Similarly, specific antibodies to galectin-1 and galectin-3 did not inhibit P. gingivalis adhesion and invasion. These results suggest that soluble galectin-1, but not galectin-3, may exacerbate periodontal disease by enhancing the adhesion to and invasion of host cells by periodontal pathogenic bacteria.

Balance between cell-substrate adhesion and myosin contraction determines the frequency of motility initiation in fish keratocytes.

PubMed

Barnhart, Erin; Lee, Kun-Chun; Allen, Greg M; Theriot, Julie A; Mogilner, Alex

2015-04-21

Cells are dynamic systems capable of spontaneously switching among stable states. One striking example of this is spontaneous symmetry breaking and motility initiation in fish epithelial keratocytes. Although the biochemical and mechanical mechanisms that control steady-state migration in these cells have been well characterized, the mechanisms underlying symmetry breaking are less well understood. In this work, we have combined experimental manipulations of cell-substrate adhesion strength and myosin activity, traction force measurements, and mathematical modeling to develop a comprehensive mechanical model for symmetry breaking and motility initiation in fish epithelial keratocytes. Our results suggest that stochastic fluctuations in adhesion strength and myosin localization drive actin network flow rates in the prospective cell rear above a critical threshold. Above this threshold, high actin flow rates induce a nonlinear switch in adhesion strength, locally switching adhesions from gripping to slipping and further accelerating actin flow in the prospective cell rear, resulting in rear retraction and motility initiation. We further show, both experimentally and with model simulations, that the global levels of adhesion strength and myosin activity control the stability of the stationary state: The frequency of symmetry breaking decreases with increasing adhesion strength and increases with increasing myosin contraction. Thus, the relative strengths of two opposing mechanical forces--contractility and cell-substrate adhesion--determine the likelihood of spontaneous symmetry breaking and motility initiation.

Drug-induced in vitro inhibition of neutrophil-endothelial cell adhesion.

PubMed Central

Pellegatta, F.; Lu, Y.; Radaelli, A.; Zocchi, M. R.; Ferrero, E.; Chierchia, S.; Gaja, G.; Ferrero, M. E.

1996-01-01

1. Leukocyte-endothelial cell interactions play an important role during ischaemia-reperfusion events. Adhesion molecules are specifically implicated in this interaction process. 2. Since defibrotide has been shown to be an efficient drug in reducing damage due to ischaemia-reperfusion in many experimental models, we analysed the effect of defibrotide in vitro on leukocyte adhesion to endothelial cells in basal conditions and after their stimulation. 3. In basal conditions, defibrotide (1000 micrograms ml-1) partially inhibited leukocyte adhesion to endothelial cells by 17.3% +/- 3.6 (P < 0.05), and after endothelial cell stimulation (TNF-alpha, 500 u ml-1) or after leukocyte stimulation (fMLP, 10(-7) M), it inhibited leukocyte adhesion by 26.5% +/- 3.4 and 32.4% +/- 1.8, respectively (P < 0.05). 4. In adhesion blockage experiments, the use of the monoclonal antibody anti-CD31 (5 micrograms ml-1) did not demonstrate a significant inhibitory effect whereas use of the monoclonal antibody anti-LFA-1 (5 micrograms ml-1) significantly interfered with the effect of defibrotide. 5. This result was confirmed in NIH/3T3-ICAM-1 transfected cells. 6. We conclude that defibrotide is able to interfere with leukocyte adhesion to endothelial cells mainly in activated conditions and that the ICAM-1/LFA-1 adhesion system is involved in the defibrotide mechanism of action. PMID:8762067

Old and sticky—adhesive mechanisms in the living fossil Nautilus pompilius (Mollusca, Cephalopoda)

PubMed Central

von Byern, Janek; Wani, Ryoji; Schwaha, Thomas; Grunwald, Ingo; Cyran, Norbert

2012-01-01

Nautiloidea is the oldest group within the cephalopoda, and modern Nautilus differs much in its outer morphology from all other recent species; its external shell and pinhole camera eye are the most prominent distinguishing characters. A further unique feature of Nautilus within the cephalopods is the lack of suckers or hooks on the tentacles. Instead, the animals use adhesive structures present on the digital tentacles. Earlier studies focused on the general tentacle morphology and put little attention on the adhesive gland system. Our results show that the epithelial parts on the oral adhesive ridge contain three secretory cell types (columnar, goblet, and cell type 1) that differ in shape and granule size. In the non-adhesive aboral epithelium, two glandular cell types (cell types 2 and 3) are present; these were not mentioned in any earlier study and differ from the cells in the adhesive area. The secretory material of all glandular cell types consists mainly of neutral mucopolysaccharide units, whereas one cell type in the non-adhesive epithelium also reacts positive for acidic mucopolysaccharides. The present data indicate that the glue in Nautilus consists mainly of neutral mucopolysaccharides. The glue seems to be a viscous carbohydrate gel, as known from another cephalopod species. De-attachment is apparently effectuated mechanically, i.e., by muscle contraction of the adhesive ridges and tentacle retraction. PMID:22221553

Pten Knockdown in vivo Increases Excitatory Drive onto Dentate Granule Cells

PubMed Central

Luikart, Bryan W.; Schnell, Eric; Washburn, Eric K.; Bensen, AeSoon L.; Tovar, Kenneth R.; Westbrook, Gary L.

2011-01-01

Some cases of autism spectrum disorder (ASD) have mutations in the lipid phosphatase, Pten (phosphatase and tensin homolog on chromosome 10). Tissue specific deletion of Pten in the hippocampus and cortex of mice causes anatomical and behavioral abnormalities similar to human autism. However, the impact of reductions in Pten on synaptic and circuit function remains unexplored. We used in vivo stereotaxic injections of lentivirus expressing an shRNA to knockdown Pten in mouse neonatal and young adult dentate granule cells. We then assessed the morphology and synaptic physiology between two weeks and four months later. Confocal imaging of the hippocampus revealed a marked increase in granule cell size and an increase in dendritic spine density. The onset of morphological changes occurred earlier in neonatal mice than in young adults. We used whole-cell recordings from granule cells in acute slices to assess synaptic function following Pten knockdown. Consistent with the increase in dendritic spines, the frequency of excitatory miniature and spontaneous postsynaptic currents increased. However, there was little or no effect on inhibitory postsynaptic currents. Thus Pten knockdown results in an imbalance between excitatory and inhibitory synaptic activity. Because reductions in Pten affected mature granule cells as well as developing granule cells, we suggest that the disruption of circuit function by Pten hypofunction may be ongoing well beyond early development. PMID:21411674

Frequency-dependent glycinergic inhibition modulates plasticity in hippocampus.

PubMed

Keck, Tara; Lillis, Kyle P; Zhou, Yu-Dong; White, John A

2008-07-16

Previous studies have demonstrated the presence of functional glycine receptors (GlyRs) in hippocampus. In this work, we examine the baseline activity and activity-dependent modulation of GlyRs in region CA1. We find that strychnine-sensitive GlyRs are open in the resting CA1 pyramidal cell, creating a state of tonic inhibition that "shunts" the magnitude of EPSPs evoked by electrical stimulation of the Schaffer collateral inputs. This GlyR-mediated shunting conductance is independent of the presynaptic stimulation rate; however, pairs of presynaptic and postsynaptic action potentials, repeated at frequencies above 5 Hz, reduce the GlyR-mediated conductance and increase peak EPSP magnitudes to levels at least 20% larger than those seen with presynaptic stimulation alone. We refer to this phenomenon as rate-dependent efficacy (RDE). Exogenous GlyR agonists (glycine, taurine) block RDE by preventing the closure of postsynaptic GlyRs. The GlyR antagonist strychnine blocks postsynaptic GlyRs under all conditions, occluding RDE. During RDE, GlyRs are less responsive to local glycine application, suggesting that a reduction in the number or sensitivity of membrane-inserted GlyRs underlies RDE. By extending the RDE induction protocol to include 500 paired presynaptic and postsynaptic spikes, we can induce long-term synaptic depression (LTD). Manipulations that lead to reduced functionality of GlyRs, either pharmacologically or through RDE, also lead to increased LTD. This result suggests that RDE contributes to long-term synaptic plasticity in the hippocampus.

Pindolol antagonises G-protein activation at both pre- and postsynaptic serotonin 5-HT1A receptors: a.

PubMed

Newman-Tancredi, A; Chaput, C; Touzard, M; Millan, M J

2001-04-01

The arylalkylamine, pindolol, may potentiate the clinical actions of antidepressant agents. Although it is thought to act via blockade of 5-HT1A autoreceptors, its efficacy at these sites remains controversial. Herein, we evaluated the actions of pindolol at 5-HT1A autoreceptors and specific populations of postsynaptic 5-HT1A receptors employing [35S]GTPgammaS autoradiography, a measure of receptor-mediated G-protein activation. Both 8-OH-DPAT (1 microM) and 5-HT (10 microM) elicited a pronounced increase in [35S]GTPyS binding in the dorsal raphe nucleus, which contains serotonergic cell bodies bearing 5-HT1A autoreceptors. Pindolol abolished their actions. In the dentate gyrus, lateral septum and entorhinal cortex, structures enriched in postsynaptic 5-HT1A receptors, 8-OH-DPAT (1 microM) and 5-HT (10 microM) also elicited a marked increase in [35S]GTPgammaS binding which was likewise blocked by pindolol. The antagonism of 5-HT-induced [35S]GTPgammaS labelling in the dentate gyrus was shown to be concentration-dependent, yielding a pIC50 of 5.82. Pindolol did not, itself, affect [35S]GTPgammaS binding in any brain region examined. In conclusion, these data suggest that, as characterised by [35S]GTPgammaS autoradiography, and compared with 5-HT and 8-OH-DPAT, pindolol possesses low efficacy at both pre- and postsynaptic 5-HT1A receptors.

Neural cell adhesion molecule-deficient beta-cell tumorigenesis results in diminished extracellular matrix molecule expression and tumour cell-matrix adhesion.

PubMed

Håkansson, Joakim; Xian, Xiaojie; He, Liqun; Ståhlberg, Anders; Nelander, Sven; Samuelsson, Tore; Kubista, Mikael; Semb, Henrik

2005-01-01

To understand by which mechanism neural cell adhesion molecule (N-CAM) limits beta tumour cell disaggregation and dissemination, we searched for potential downstream genes of N-CAM during beta tumour cell progression by gene expression profiling. Here, we show that N-CAM-deficient beta-cell tumorigenesis is associated with changes in the expression of genes involved in cell-matrix adhesion and cytoskeletal dynamics, biological processes known to affect the invasive and metastatic behaviour of tumour cells. The extracellular matrix (ECM) molecules emerged as the primary target, i.e. N-CAM deficiency resulted in down-regulated mRNA expression of a broad range of ECM molecules. Consistent with this result, deficient deposition of major ECM stromal components, such as fibronectin, laminin 1 and collagen IV, was observed. Moreover, N-CAM-deficient tumour cells displayed defective matrix adhesion. These results offer a potential mechanism for tumour cell disaggregation during N-CAM-deficient beta tumour cell progression. Prospective consequences of these findings for the role of N-CAM in beta tumour cell dissemination are discussed.

The Drosophila cell adhesion molecule Neuroglian regulates Lissencephaly-1 localisation in circulating immunosurveillance cells.

PubMed

Williams, Michael J

2009-03-25

When the parasitoid wasp Leptopilina boulardi lays its eggs in Drosophila larvae phagocytic cells called plasmatocytes and specialized cells known as lamellocytes encapsulate the egg. This requires these circulating immunosurveillance cells (haemocytes) to change from a non-adhesive to an adhesive state enabling them to bind to the invader. Interestingly, attachment of leukocytes, platelets, and insect haemocytes requires the same adhesion complexes as epithelial and neuronal cells. Here evidence is presented showing that the Drosophila L1-type cell adhesion molecule Neuroglian (Nrg) is required for haemocytes to encapsulate L. boulardi wasp eggs. The amino acid sequence FIGQY containing a conserved phosphorylated tyrosine is found in the intracellular domain of all L1-type cell adhesion molecules. This conserved tyrosine is phosphorylated at the cell periphery of plasmatocytes and lamellocytes prior to parasitisation, but dephosphorylated after immune activation. Intriguingly, another pool of Nrg located near the nucleus of plasmatocytes remains phosphorylated after parasitisation. In mammalian neuronal cells phosphorylated neurofascin, another L1-type cell adhesion molecule interacts with a nucleokinesis complex containing the microtubule binding protein lissencephaly-1 (Lis1) 1. Interestingly in plasmatocytes from Nrg mutants the nucleokinesis regulating protein Lissencephaly-1 (Lis1) fails to localise properly around the nucleus and is instead found diffuse throughout the cytoplasm and at unidentified perinuclear structures. After attaching to the wasp egg control plasmatocytes extend filopodia laterally from their cell periphery; as well as extending lateral filopodia plasmatocytes from Nrg mutants also extend many filopodia from their apical surface. The Drosophila cellular adhesion molecule Neuroglian is expressed in haemocytes and its activity is required for the encapsulation of L. boularli eggs. At the cell periphery of haemocytes Neuroglian may be involved in cell-cell interactions, while at the cell centre Neuroglian regulates the localisation of the nucleokinesis complex protein lissencephaly-1.

Inhibition of endothelial receptor expression and of T-cell ligand activity by mycophenolate mofetil.

PubMed

Blaheta, R A; Leckel, K; Wittig, B; Zenker, D; Oppermann, E; Harder, S; Scholz, M; Weber, S; Schuldes, H; Encke, A; Markus, B H

1998-12-01

The novel immunosuppressive drug mycophenolate mofetil (CellCept, MMF) blocks DNA-synthesis by the inhibition of the enzyme inosine monophosphate dehydrogenase (IMDH). IMDH is also involved in the synthesis of adhesion receptors which are known to play an important role in the regulation of cell-cell contacts. Therefore, application of MMF might lead to a reduction of cellular infiltrates in the course of transplant rejection. To evaluate the therapeutic value of MMF, we investigated to what extent MMF blocks T-lymphocyte infiltration in vitro with regard to (a) adhesion to endothelial cells, (b) horizontal migration along these cells and (c) penetration through the endothelial cells. The results demonstrated a strong inhibition of both CD4+ and CD8+ T-cell adhesion and penetration by MMF. The ID50 value for CD4+ T-cell adhesion was calculated to be 0.03 microM and the ID50 value for CD4+ T-cell penetration 1.21 microM. MMF did not significantly influence the horizontal migration of T-lymphocytes along the human vascular endothelial cell (HUVEC) borders. FACS-analysis revealed a diminished E-selectin and P-selectin expression on endothelial cell membranes in the presence of MMF. Although MMF did not interfere with the synthesis of T-cell adhesion ligands, the binding activity of lymphocytic leucocyte function associated antigen 1 (LFA-1), very late antigen 4 (VLA-4) and PSGL-1 (P-selectin glycoprotein ligand 1) to immobilized intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and P-selectin was impaired. Moreover, MMF prevented VLA-4 and PSGL-1 receptor accumulation on the membranes of T-cell pseudopodia. It can be concluded that MMF possesses potent infiltration blocking properties. MMF evoked down-regulation of specific endothelial membrane molecules and the loss of protein localization in the lymphocyte protrusions might be predominantly responsible for the observed blockade of cell adhesion and penetration.

The Drosophila cell adhesion molecule Neuroglian regulates Lissencephaly-1 localisation in circulating immunosurveillance cells

PubMed Central

Williams, Michael J

2009-01-01

Background When the parasitoid wasp Leptopilina boulardi lays its eggs in Drosophila larvae phagocytic cells called plasmatocytes and specialized cells known as lamellocytes encapsulate the egg. This requires these circulating immunosurveillance cells (haemocytes) to change from a non-adhesive to an adhesive state enabling them to bind to the invader. Interestingly, attachment of leukocytes, platelets, and insect haemocytes requires the same adhesion complexes as epithelial and neuronal cells. Results Here evidence is presented showing that the Drosophila L1-type cell adhesion molecule Neuroglian (Nrg) is required for haemocytes to encapsulate L. boulardi wasp eggs. The amino acid sequence FIGQY containing a conserved phosphorylated tyrosine is found in the intracellular domain of all L1-type cell adhesion molecules. This conserved tyrosine is phosphorylated at the cell periphery of plasmatocytes and lamellocytes prior to parasitisation, but dephosphorylated after immune activation. Intriguingly, another pool of Nrg located near the nucleus of plasmatocytes remains phosphorylated after parasitisation. In mammalian neuronal cells phosphorylated neurofascin, another L1-type cell adhesion molecule interacts with a nucleokinesis complex containing the microtubule binding protein lissencephaly-1 (Lis1) [1]. Interestingly in plasmatocytes from Nrg mutants the nucleokinesis regulating protein Lissencephaly-1 (Lis1) fails to localise properly around the nucleus and is instead found diffuse throughout the cytoplasm and at unidentified perinuclear structures. After attaching to the wasp egg control plasmatocytes extend filopodia laterally from their cell periphery; as well as extending lateral filopodia plasmatocytes from Nrg mutants also extend many filopodia from their apical surface. Conclusion The Drosophila cellular adhesion molecule Neuroglian is expressed in haemocytes and its activity is required for the encapsulation of L. boularli eggs. At the cell periphery of haemocytes Neuroglian may be involved in cell-cell interactions, while at the cell centre Neuroglian regulates the localisation of the nucleokinesis complex protein lissencephaly-1. PMID:19320973

Protein adhesives

Treesearch

Charles R. Frihart; Linda F. Lorenz

2018-01-01

Nature uses a wide variety of chemicals for providing adhesion internally (e.g., cell to cell) and externally (e.g., mussels to ships and piers). This adhesive bonding is chemically and mechanically complex, involving a variety of proteins, carbohydrates, and other compounds.Consequently,the effect of protein structures on adhesive properties is only partially...

Increased soluble vascular cell adhesion molecule-1 plasma levels and soluble intercellular adhesion molecule-1 during antiretroviral therapy interruption and retention of elevated soluble vascular cellular adhesion molecule-1 levels following resumption of antiretroviral therapy.

PubMed

Papasavvas, Emmanouil; Azzoni, Livio; Pistilli, Maxwell; Hancock, Aidan; Reynolds, Griffin; Gallo, Cecile; Ondercin, Joe; Kostman, Jay R; Mounzer, Karam; Shull, Jane; Montaner, Luis J

2008-06-19

We investigated the effect of short viremic episodes on soluble markers associated with endothelial stress and cardiovascular disease risk in chronically HIV-1-infected patients followed during continuous antiretroviral therapy, antiretroviral therapy interruption and antiretroviral therapy resumption. We assessed changes in plasma levels of von Willebrand factor, soluble vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 by enzyme-linked immunosorbent assay, as well as T-cell activation (CD8+/CD38+, CD8+/HLA-DR+ and CD3+/CD95+) by flow cytometry, in 36 chronically HIV-1-infected patients participating in a randomized study. Patients were divided into the following three groups: a, on continuous antiretroviral therapy; b, on a 6-week antiretroviral therapy interruption; or c, on antiretroviral therapy interruption extended to the achievement of viral set point. Although all measurements remained stable over a 40-week follow-up on antiretroviral therapy, plasma levels of soluble vascular cell adhesion molecule-1 (P < 0.0001) and soluble intercellular adhesion molecule-1 (P = 0.003) increased during treatment interruption in correlation with viral rebound and T-cell activation. No significant changes in von Willebrand factor were observed in any of the groups. After resuming antiretroviral therapy, soluble vascular cell adhesion molecule-1 levels remained elevated even after achievement of viral suppression to less than 50 copies/ml. The prompt rise in plasma soluble vascular cell adhesion molecule-1 and soluble intercellular adhesion molecule-1 upon viral rebound suggests an acute increase in endothelial stress upon treatment interruption, which may persists after viral resuppression of virus. Thus, viral replication during short-term treatment interruption may increase the overall cardiovascular risk during and beyond treatment interruption.

Surface free energy predominates in cell adhesion to hydroxyapatite through wettability.

PubMed

Nakamura, Miho; Hori, Naoko; Ando, Hiroshi; Namba, Saki; Toyama, Takeshi; Nishimiya, Nobuyuki; Yamashita, Kimihiro

2016-05-01

The initial adhesion of cells to biomaterials is critical in the regulation of subsequent cell behaviors. The purpose of this study was to investigate a mechanism through which the surface wettability of biomaterials can be improved and determine the effects of biomaterial surface characteristics on cellular behaviors. We investigated the surface characteristics of various types of hydroxyapatite after sintering in different atmospheres and examined the effects of various surface characteristics on cell adhesion to study cell-biomaterial interactions. Sintering atmosphere affects the polarization capacity of hydroxyapatite by changing hydroxide ion content and grain size. Compared with hydroxyapatite sintered in air, hydroxyapatite sintered in saturated water vapor had a higher polarization capacity that increased surface free energy and improved wettability, which in turn accelerated cell adhesion. We determined the optimal conditions of hydroxyapatite polarization for the improvement of surface wettability and acceleration of cell adhesion. Copyright © 2016 Elsevier B.V. All rights reserved.

Unitary synaptic connections among substantia nigra pars reticulata neurons

PubMed Central

Wilson, Charles J.

2016-01-01

Neurons in substantia nigra pars reticulata (SNr) are synaptically coupled by local axon collaterals, providing a potential mechanism for local signal processing. Because SNr neurons fire spontaneously, these synapses are constantly active. To investigate their properties, we recorded spontaneous inhibitory postsynaptic currents (sIPSCs) from SNr neurons in brain slices, in which afferents from upstream nuclei are severed, and the cells fire rhythmically. The sIPSC trains contained a mixture of periodic and aperiodic events. Autocorrelation analysis of sIPSC trains showed that a majority of cells had one to four active unitary inputs. The properties of the unitary IPSCs (uIPSCs) were analyzed for cells with one unitary input, using a model of periodic presynaptic firing and stochastic synaptic transmission. The inferred presynaptic firing rates and coefficient of variation of interspike intervals (ISIs) corresponded well with direct measurements of spiking in SNr neurons. Methods were developed to estimate the success probability, amplitude distributions, and kinetics of the uIPSCs, while removing the contribution from aperiodic sIPSCs. The sIPSC amplitudes were not increased upon release from halorhodopsin silencing, suggesting that most synapses were not depressed at the spontaneous firing rate. Gramicidin perforated-patch recordings indicated that the average reversal potential of spontaneous inhibitory postsynaptic potentials was −64 mV. Because of the change in driving force across the ISI, the unitary inputs are predicted to have a larger postsynaptic impact when they arrive late in the ISI. Simulations of network activity suggest that this very sparse inhibitory coupling may act to desynchronize the activity of SNr neurons while having only a small effect on firing rate. PMID:26961101

Early changes in synaptic connectivity following progressive photoreceptor degeneration in RCS rats.

PubMed

Cuenca, Nicolás; Pinilla, Isabel; Sauvé, Yves; Lund, Raymond

2005-09-01

The Royal College of Surgeons (RCS) rat has a retinal pigment epithelial cell defect that causes progressive loss of photoreceptors. Although it is extensively used in retinal degeneration and repair studies, how photoreceptor degeneration affects retinal circuitry has not been fully explored. This study examined the changes in synaptic connectivity between photoreceptors and their target cells using immunocytochemistry and correlated these changes with retinal function using the electroretinogram (ERG). Immunostaining with bassoon and synaptophysin (as presynaptic markers) and metabotropic glutamate receptor (mGluR6, a postsynaptic marker for ON-bipolar dendrites) was already impaired at postnatal day (P) 21 and progressively lost with infrequent pairing of presynaptic and postsynaptic elements at P60. By P90 to P120, staining became increasingly patchy and was eventually restricted to sparsely and irregularly distributed foci in which the normal pairing of presynaptic and postsynaptic markers was lost. ERG results showed that mixed scotopic a-waves and b-waves were already reduced by P21 but not oscillatory potentials. While cone-driven responses (photopic b-wave) reached normal levels at P30, they were impaired by P60 but could still be recorded at P120, although with reduced amplitude; rod responses never reached normal amplitudes. Thus, only cone-driven activity attained normal levels, but declined rapidly thereafter. In conclusion, the synaptic markers associated with photoreceptors and processes of bipolar and horizontal cells show abnormalities prior to significant photoreceptor loss. These changes are paralleled with the deterioration of specific aspects of ERG responsiveness with age. Besides providing information on the effects of photoreceptor dysfunction and loss on connection patterns in the retina, the work addresses the more general issue of how disorder of input neurons affects downstream circuitry.

Receptor-like Molecules on Human Intestinal Epithelial Cells Interact with an Adhesion Factor from Lactobacillus reuteri

PubMed Central

MATSUO, Yosuke; MIYOSHI, Yukihiro; OKADA, Sanae; SATOH, Eiichi

2012-01-01

A surface protein of Lactobacillus reuteri, mucus adhesion-promoting protein (MapA), is considered to be an adhesion factor. MapA is expressed in L. reuteri strains and adheres to piglet gastric mucus, collagen type I, and human intestinal epithelial cells such as Caco-2. The aim of this study was to identify molecules that mediate the attachment of MapA from L. reuteri to the intestinal epithelial cell surface by investigating the adhesion of MapA to receptor-like molecules on Caco-2 cells. MapA-binding receptor-like molecules were detected in Caco-2 cell lysates by 2D-PAGE. Two proteins, annexin A13 (ANXA13) and paralemmin (PALM), were identified by MALDI TOF-MS. The results of a pull-down assay showed that MapA bound directly to ANXA13 and PALM. Fluorescence microscopy studies confirmed that MapA binding to ANXA13 and PALM was colocalized on the Caco-2 cell membrane. To evaluate whether ANXA13 and PALM are important for MapA adhesion, ANXA13 and PALM knockdown cell lines were established. The adhesion of MapA to the abovementioned cell lines was reduced compared with that to wild-type Caco-2 cells. These knockdown experiments established the importance of these receptor-like molecules in MapA adhesion. PMID:24936355

Mechanisms for Flow-Enhanced Cell Adhesion

PubMed Central

Zhu, Cheng; Yago, Tadayuki; Lou, Jizhong; Zarnitsyna, Veronika I.; McEver, Rodger P.

2009-01-01

Cell adhesion is mediated by specific receptor—ligand bonds. In several biological systems, increasing flow has been observed to enhance cell adhesion despite the increasing dislodging fluid shear forces. Flow-enhanced cell adhesion includes several aspects: flow augments the initial tethering of flowing cells to a stationary surface, slows the velocity and increases the regularity of rolling cells, and increases the number of rollingly adherent cells. Mechanisms for this intriguing phenomenon may include transport-dependent acceleration of bond formation and force-dependent deceleration of bond dissociation. The former includes three distinct transport modes: sliding of cell bottom on the surface, Brownian motion of the cell, and rotational diffusion of the interacting molecules. The latter involves a recently demonstrated counterintuitive behavior called catch bonds where force prolongs rather than shortens the lifetimes of receptor—ligand bonds. In this article, we summarize our recently published data that used dimensional analysis and mutational analysis to elucidate the above mechanisms for flow-enhanced leukocyte adhesion mediated by L-selectinligand interactions. PMID:18299992

Le(x) glycan mediates homotypic adhesion of embryonal cells independently from E-cadherin: a preliminary note.

PubMed

Handa, Kazuko; Takatani-Nakase, Tomoka; Larue, Lionel; Stemmler, Marc P; Kemler, Rolf; Hakomori, Sen-itiroh

2007-06-22

Le(x) glycan and E-cadherin (Ecad) are co-expressed at embryonal stem (ES) cells and embryonal carcinoma (EC) cells. While the structure and function of Ecad mediating homotypic adhesion of these cells have been well established, evidence that Le(x) glycan also mediates such adhesion is weak, despite the fact that Le(x) oligosaccharide inhibits the compaction process. To provide stronger evidence, we knocked out Ecad gene in EC and ES cells to establish F9 Ecad (-/-) and D3M Ecad (-/-) cells, which highly express Le(x) glycan but do not express Ecad at all. Both F9 Ecad (-/-) and D3M Ecad (-/-) cells displayed strong autoaggregation in the presence of Ca(2+), while PYS-2 cells, which express trace amount of Ecad and undetectable level of Le(x) glycan, did not display autoaggregation. In addition, F9 Ecad (-/-) and D3M Ecad (-/-) cells displayed strong adhesion to plates coated with Le(x) glycosphingolipid (III(3)FucnLc4Cer), in dose-dependent manner, in the presence of Ca(2+). Thus, ES or EC cells display autoaggregation and strong adhesion to Le(x)-coated plates in the absence of Ecad, further supporting the notion of Le(x) self-recognition (i.e., Le(x)-to-Le(x) interaction) in cell adhesion.

Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs): Positive and negative regulators in tumor cell adhesion.

PubMed

Bourboulia, Dimitra; Stetler-Stevenson, William G

2010-06-01

Cells adhere to one another and/or to matrices that surround them. Regulation of cell-cell (intercellular) and cell-matrix adhesion is tightly controlled in normal cells, however, defects in cell adhesion are common in the majority of human cancers. Multilateral communication among tumor cells with the extracellular matrix (ECM) and neighbor cells is accomplished through adhesion molecules, ECM components, proteolytic enzymes and their endogenous inhibitors. There is sufficient evidence to suggest that reduced adherence is a tumor cell property engaged during tumor progression. Tumor cells acquire the ability to change shape, detach and easily move through spaces disorganizing the normal tissue architecture. This property is due to changes in expression levels of adhesion molecules and/or due to elevated levels of secreted proteolytic enzymes, including matrix metalloproteinases (MMPs). Among other roles, MMPs degrade the ECM and, therefore, prepare the path for tumor cells to migrate, invade and spread to distant secondary areas, where they form metastasis. Tissue inhibitors of metalloproteinases or TIMPs control MMP activities and, therefore, minimize matrix degradation. Both MMPs and TIMPs are involved in tissue remodeling and decisively regulate tumor cell progression including tumor angiogenesis. In this review, we describe and discuss data that support the important role of MMPs and TIMPs in cancer cell adhesion and tumor progression. Published by Elsevier Ltd.

Extract of corn silk (stigma of Zea mays) inhibits the tumour necrosis factor-alpha- and bacterial lipopolysaccharide-induced cell adhesion and ICAM-1 expression.

PubMed

Habtemariam, S

1998-05-01

Treatment of human endothelial cells with cytokines such as tumour necrosis factor-alpha (TNF) or E. coli lipopolysaccharide (LPS) induces the expression of several adhesion molecules and enhances leukocyte adhesion to endothelial cell surface. Interfering with this leukocyte adhesion or adhesion molecules upregulation is an important therapeutic target for the treatment of bacterial sepsis and various inflammatory diseases. In the course of screening marketed European anti-inflammatory herbal drugs for TNF antagonistic activity, a crude ethanolic extract of corn silk (stigma of Zea mays) exhibited significant activity. The extract at concentrations of 9-250 micrograms/ml effectively inhibited the TNF- and LPS-induced adhesiveness of EAhy 926 endothelial cells to monocytic U937 cells. Similar concentration ranges of corn silk extract did also block the TNF and LPS but not the phorbol 12-myristate 13-acetate-induced ICAM-1 expression on EAhy 926 endothelial cell surface. The extract did not alter the production of TNF by LPS-activated macrophages and failed to inhibit the cytotoxic activity of TNF. It is concluded that corn silk possesses important therapeutic potential for TNF- and LPS-mediated leukocyte adhesion and trafficking.

PAK4 promotes kinase-independent stabilization of RhoU to modulate cell adhesion

PubMed Central

Dart, Anna E.; Box, Gary M.; Court, William; Gale, Madeline E.; Brown, John P.; Pinder, Sarah E.; Eccles, Suzanne A.

2015-01-01

P21-activated kinase 4 (PAK4) is a Cdc42 effector protein thought to regulate cell adhesion disassembly in a kinase-dependent manner. We found that PAK4 expression is significantly higher in high-grade human breast cancer patient samples, whereas depletion of PAK4 modifies cell adhesion dynamics of breast cancer cells. Surprisingly, systematic analysis of PAK4 functionality revealed that PAK4-driven adhesion turnover is neither dependent on Cdc42 binding nor kinase activity. Rather, reduced expression of PAK4 leads to a concomitant loss of RhoU expression. We report that RhoU is targeted for ubiquitination by the Rab40A–Cullin 5 complex and demonstrate that PAK4 protects RhoU from ubiquitination in a kinase-independent manner. Overexpression of RhoU rescues the PAK4 depletion phenotype, whereas loss of RhoU expression reduces cell adhesion turnover and migration. These data support a new kinase-independent mechanism for PAK4 function, where an important role of PAK4 in cellular adhesions is to stabilize RhoU protein levels. Thus, PAK4 and RhoU cooperate to drive adhesion turnover and promote cell migration. PMID:26598620

Adhesion behavior of endothelial progenitor cells to endothelial cells in simple shear flow

NASA Astrophysics Data System (ADS)

Gong, Xiao-Bo; Li, Yu-Qing; Gao, Quan-Chao; Cheng, Bin-Bin; Shen, Bao-Rong; Yan, Zhi-Qiang; Jiang, Zong-Lai

2011-12-01

The adhesion of endothelial progenitor cells (EPCs) on endothelial cells (ECs) is one of the critical physiological processes for the regenesis of vascular vessels and the prevention of serious cardiovascular diseases. Here, the rolling and adhesion behavior of EPCs on ECs was studied numerically. A two-dimensional numerical model was developed based on the immersed boundary method for simulating the rolling and adhesion of cells in a channel flow. The binding force arising from the catch bond of a receptor and ligand pair was modeled with stochastic Monte Carlo method and Hookean spring model. The effect of tumor necrosis factor alpha (TNF- α) on the expression of the number of adhesion molecules in ECs was analyzed experimentally. A flow chamber system with CCD camera was set up to observe the top view of the rolling of EPCs on the substrate cultivated with ECs. Numerical results prove that the adhesion of EPC on ECs is closely related to membrane stiffness of the cell and shear rate of the flow. It also suggests that the adhesion force between EPC and EC by P-selectin glycoprotein ligand-1 only is not strong enough to bond the cell onto vessel walls unless contributions of other catch bond are considered. Experimental results demonstrate that TNF- α enhanced the expressions of VCAM, ICAM, P-selectin and E-selectin in ECs, which supports the numerical results that the rolling velocity of EPC on TNF- α treated EC substrate decreases obviously compared with its velocity on the untreated one. It is found that because the adhesion is affected by both the rolling velocity and the deformability of the cell, an optimal stiffness of EPC may exist at a given shear rate of flow for achieving maximum adhesion rates.

Mesalamine modulates intercellular adhesion through inhibition of p-21 activated kinase-1

PubMed Central

Khare, Vineeta; Lyakhovich, Alex; Dammann, Kyle; Lang, Michaela; Borgmann, Melanie; Tichy, Boris; Pospisilova, Sarka; Luciani, Gloria; Campregher, Christoph; Evstatiev, Rayko; Pflueger, Maren; Hundsberger, Harald; Gasche, Christoph

2013-01-01

Mesalamine (5-ASA) is widely used for the treatment of ulcerative colitis, a remitting condition characterized by chronic inflammation of the colon. Knowledge about the molecular and cellular targets of 5-ASA is limited and a clear understanding of its activity in intestinal homeostasis and interference with neoplastic progression is lacking. We sought to identify molecular pathways interfered by 5-ASA, using CRC cell lines with different genetic background. Microarray was performed for gene expression profile of 5-ASA-treated and untreated cells (HCT116 and HT29). Filtering and analysis of data identified three oncogenic pathways interfered by 5-ASA: MAPK/ERK pathway, cell adhesion and β-catenin/Wnt signaling. PAK1 emerged as a consensus target of 5-ASA, orchestrating these pathways. We further investigated the effect of 5-ASA on cell adhesion. 5-ASA increased cell adhesion which was measured by cell adhesion assay and transcellular-resistance measurement. Moreover, 5-ASA treatment restored membranous expression of adhesion molecules E-cadherin and β-catenin. Role of PAK1 as a mediator of mesalamine activity was validated in vitro and in vivo. Inhibition of PAK1 by RNA interference also increased cell adhesion. PAK1 expression was elevated in APCmin polyps and 5-ASA treatment reduced its expression. Our data demonstrates novel pharmacological mechanism of mesalamine in modulation of cell adhesion and role of PAK1 in APCmin polyposis. We propose that inhibition of PAK1 expression by 5-ASA can impede with neoplastic progression in colorectal carcinogenesis. The mechanism of PAK1 inhibition and induction of membranous translocation of adhesion proteins by 5-ASA might be independent of its known anti-inflammatory action. PMID:23146664

Mesalamine modulates intercellular adhesion through inhibition of p-21 activated kinase-1.

PubMed

Khare, Vineeta; Lyakhovich, Alex; Dammann, Kyle; Lang, Michaela; Borgmann, Melanie; Tichy, Boris; Pospisilova, Sarka; Luciani, Gloria; Campregher, Christoph; Evstatiev, Rayko; Pflueger, Maren; Hundsberger, Harald; Gasche, Christoph

2013-01-15

Mesalamine (5-ASA) is widely used for the treatment of ulcerative colitis, a remitting condition characterized by chronic inflammation of the colon. Knowledge about the molecular and cellular targets of 5-ASA is limited and a clear understanding of its activity in intestinal homeostasis and interference with neoplastic progression is lacking. We sought to identify molecular pathways interfered by 5-ASA, using CRC cell lines with different genetic background. Microarray was performed for gene expression profile of 5-ASA-treated and untreated cells (HCT116 and HT29). Filtering and analysis of data identified three oncogenic pathways interfered by 5-ASA: MAPK/ERK pathway, cell adhesion and β-catenin/Wnt signaling. PAK1 emerged as a consensus target of 5-ASA, orchestrating these pathways. We further investigated the effect of 5-ASA on cell adhesion. 5-ASA increased cell adhesion which was measured by cell adhesion assay and transcellular-resistance measurement. Moreover, 5-ASA treatment restored membranous expression of adhesion molecules E-cadherin and β-catenin. Role of PAK1 as a mediator of mesalamine activity was validated in vitro and in vivo. Inhibition of PAK1 by RNA interference also increased cell adhesion. PAK1 expression was elevated in APC(min) polyps and 5-ASA treatment reduced its expression. Our data demonstrates novel pharmacological mechanism of mesalamine in modulation of cell adhesion and role of PAK1 in APC(min) polyposis. We propose that inhibition of PAK1 expression by 5-ASA can impede with neoplastic progression in colorectal carcinogenesis. The mechanism of PAK1 inhibition and induction of membranous translocation of adhesion proteins by 5-ASA might be independent of its known anti-inflammatory action. Copyright © 2012 Elsevier Inc. All rights reserved.

Modeling and measurement of vesicle pools at the cone ribbon synapse: changes in release probability are solely responsible for voltage-dependent changes in release

PubMed Central

Thoreson, Wallace B.; Van Hook, Matthew J.; Parmelee, Caitlyn; Curto, Carina

2015-01-01

Post-synaptic responses are a product of quantal amplitude (Q), size of the releasable vesicle pool (N), and release probability (P). Voltage-dependent changes in presynaptic Ca2+ entry alter post-synaptic responses primarily by changing P but have also been shown to influence N. With simultaneous whole cell recordings from cone photoreceptors and horizontal cells in tiger salamander retinal slices, we measured N and P at cone ribbon synapses by using a train of depolarizing pulses to stimulate release and deplete the pool. We developed an analytical model that calculates the total pool size contributing to release under different stimulus conditions by taking into account the prior history of release and empirically-determined properties of replenishment. The model provided a formula that calculates vesicle pool size from measurements of the initial post-synaptic response and limiting rate of release evoked by a train of pulses, the fraction of release sites available for replenishment, and the time constant for replenishment. Results of the model showed that weak and strong depolarizing stimuli evoked release with differing probabilities but the same size vesicle pool. Enhancing intraterminal Ca2+ spread by lowering Ca2+ buffering or applying BayK8644 did not increase PSCs evoked with strong test steps showing there is a fixed upper limit to pool size. Together, these results suggest that light-evoked changes in cone membrane potential alter synaptic release solely by changing release probability. PMID:26541100

Structure, Distribution, and Function of Neuronal/Synaptic Spinules and Related Invaginating Projections

PubMed Central

Petralia, Ronald S.; Wang, Ya-Xian; Mattson, Mark P.; Yao, Pamela J.

2015-01-01

Neurons and especially their synapses often project long thin processes that can invaginate neighboring neuronal or glial cells. These “invaginating projections” can occur in almost any combination of postsynaptic, presynaptic, and glial processes. Invaginating projections provide a precise mechanism for one neuron to communicate or exchange material exclusively at a highly localized site on another neuron, e.g., to regulate synaptic plasticity. The best-known types are postsynaptic projections called “spinules” that invaginate into presynaptic terminals. Spinules seem to be most prevalent at large very active synapses. Here, we present a comprehensive review of all kinds of invaginating projections associated with both neurons in general and more specifically with synapses; we describe them in all animals including simple, basal metazoans. These structures may have evolved into more elaborate structures in some higher animal groups exhibiting greater synaptic plasticity. In addition to classic spinules and filopodial invaginations, we describe a variety of lesser-known structures such as amphid microvilli, spinules in giant mossy terminals and en marron/brush synapses, the highly specialized fish retinal spinules, the trophospongium, capitate projections, and fly gnarls, as well as examples in which the entire presynaptic or postsynaptic process is invaginated. These various invaginating projections have evolved to modify the function of a particular synapse, or to channel an effect to one specific synapse or neuron, without affecting those nearby. We discuss how they function in membrane recycling, nourishment, and cell signaling and explore how they might change in aging and disease. PMID:26007200

MiR-9 is involved in TGF-β1-induced lung cancer cell invasion and adhesion by targeting SOX7.

PubMed

Han, Lichun; Wang, Wei; Ding, Wei; Zhang, Lijian

2017-09-01

MicroRNA (miR)-9 plays different roles in different cancer types. Here, we investigated the role of miR-9 in non-small-cell lung cancer (NSCLC) cell invasion and adhesion in vitro and explored whether miR-9 was involved in transforming growth factor-beta 1 (TGF-β1)-induced NSCLC cell invasion and adhesion by targeting SOX7. The expression of miR-9 and SOX7 in human NSCLC tissues and cell lines was examined by reverse transcription-quantitative polymerase chain reaction. Gain-of-function and loss-of-function experiments were performed on A549 and HCC827 cells to investigate the effect of miR-9 and SOX7 on NSCLC cell invasion and adhesion in the presence or absence of TGF-β1. Transwell-Matrigel assay and cell adhesion assay were used to examine cell invasion and adhesion abilities. Luciferase reporter assay was performed to determine whether SOX7 was a direct target of miR-9. We found miR-9 was up-regulated and SOX7 was down-regulated in human NSCLC tissues and cell lines. Moreover, SOX7 expression was negatively correlated with miR-9 expression. miR-9 knockdown or SOX7 overexpression could suppress TGF-β1-induced NSCLC cell invasion and adhesion. miR-9 directly targets the 3' untranslated region of SOX7, and SOX7 protein expression was down-regulated by miR-9. TGF-β1 induced miR-9 expression in NSCLC cells. miR-9 up-regulation led to enhanced NSCLC cell invasion and adhesion; however, these effects could be attenuated by SOX7 overexpression. We concluded that miR-9 expression was negatively correlated with SOX7 expression in human NSCLC. miR-9 was up-regulated by TGF-β1 and contributed to TGF-β1-induced NSCLC cell invasion and adhesion by directly targeting SOX7. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Tracking the pathway of diesel exhaust particles from the nose to the brain by X-ray florescence analysis

NASA Astrophysics Data System (ADS)

Matsui, Yasuto; Sakai, Nobumitsu; Tsuda, Akira; Terada, Yasuko; Takaoka, Masaki; Fujimaki, Hidekazu; Uchiyama, Iwao

2009-08-01

Studies have shown that exposure to nano-sized particles (< 50 nm) result in their translocation to the central nervous system through the olfactory nerve. Translocation commonly occurs via inhalation, ingestion and skin uptake. Little information is available on the specific pathway of cellular localization of nano-sized particles in the olfactory bulb. The nano-sized particles entrance into the postsynaptics cell is of particular interest because the mitral cell projects to the central nucleus of the amygdala and the piriform cortex. Therefore, our objective in this follow-up study has been to determine whether or not the mitral cells project nano-sized particles to the brain. Nano-sized particles in this study were generated using diesel exhaust. Lab mice were exposed for a period of 4 weeks. We employed synchrotron radiation (SPring-8, Japan) to determine the concentration levels of metal in the olfactory neuron pathway. Metal levels were assayed by mapping, using X-ray fluorescence analysis. The major metal components measured in the filter that collected the inhaled diesel exhaust particles were calcium, copper, iron, nickel and zinc. Our studies reveal an increase in the amount of nano-sized particles in the glomerular layer as well as in the neurons in the olfactory epithelium. Higher levels of nickel and iron were found in the olfactory epithelium's lamina propria mucosae in comparison to that in the control group. Higher levels of iron also were observed in the glomerular layer. Our studies do not clarify the specifics of metal adhesion and detachment. This remains to be one of the key issues requiring further clarification.

Upregulation of ADAM12 contributes to accelerated cell proliferation and cell adhesion-mediated drug resistance (CAM-DR) in Non-Hodgkin's Lymphoma.

PubMed

Yin, Haibing; Zhong, Fei; Ouyang, Yu; Wang, Qiru; Ding, Linlin; He, Song

2017-10-01

ADAM12 is a member of a disintegrin and metalloproteinase family and has been reported to participate in the development of variety of tumors. However, the role of ADAM12 in Non-Hodgkin Lymphoma (NHL) has not been investigated. The present study was undertaken to determine the expression and biologic function of ADAM12 in human NHL. First, we constructed a model of cell adhesion in NHL, the mRNA, and protein level of ADAM12 in suspension and the adhesion model was analyzed by RT-PCR and western blot. Then, flow cytometry assay and western blot were used to investigate the mechanism of ADAM12 in the proliferation of NHL cells. In vitro, after using siRNA interfering ADAM12 expression, we performed adhesion assay and cell viability assay to determine the effect of ADAM12 on adhesive rate and drug sensitivity. ADAM12 was lowly expressed in suspended cells and highly expressed in adherent NHL cells. In addition, ADAM12 was positively correlated with the proliferation and apoptosis of NHL cells by regulating the expression of p-AKT and p-GSK-3β. Furthermore, ADAM12 promoted cell adhesion-mediated drug resistance (CAM-DR) in DLBCL via AKT signaling pathway. Our data support a role for ADAM12 in NHL cell proliferation, adhesion, and drug resistance, and it may pave the way for a novel therapeutic approach for CAM-DR in NHL.

Redundant control of migration and adhesion by ERM proteins in vascular smooth muscle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baeyens, Nicolas; Latrache, Iman; Yerna, Xavier

Highlights: •The three ERM proteins are expressed in vascular smooth muscle cell. •ERM depletion inhibited PDGF-evoked migration redundantly. •ERM depletion increased cell adhesion redundantly. •ERM depletion did not affect PDGF-evoked Ca signal, Rac1 activation, proliferation. •ERM proteins control PDGF-induced migration by regulating adhesion. -- Abstract: Ezrin, radixin, and moesin possess a very similar structure with a C-terminal actin-binding domain and a N-terminal FERM interacting domain. They are known to be involved in cytoskeleton organization in several cell types but their function in vascular smooth muscle cells (VSMC) is still unknown. The aim of this study was to investigate the rolemore » of ERM proteins in cell migration induced by PDGF, a growth factor involved in pathophysiological processes like angiogenesis or atherosclerosis. We used primary cultured VSMC obtained from rat aorta, which express the three ERM proteins. Simultaneous depletion of the three ERM proteins with specific siRNAs abolished the effects of PDGF on cell architecture and migration and markedly increased cell adhesion and focal adhesion size, while these parameters were only slightly affected by depletion of ezrin, radixin or moesin alone. Rac1 activation, cell proliferation, and Ca{sup 2+} signal in response to PDGF were unaffected by ERM depletion. These results indicate that ERM proteins exert a redundant control on PDGF-induced VSMC migration by regulating focal adhesion turn-over and cell adhesion to substrate.« less

Monitoring of integrin-mediated adhesion of human ovarian cancer cells to model protein surfaces by quartz crystal resonators: evaluation in the impedance analysis mode.

PubMed

Li, Jing; Thielemann, Christiane; Reuning, Ute; Johannsmann, Diethelm

2005-01-15

The quartz crystal microbalance (QCM) was used to monitor specific, integrin-mediated adhesion of human ovarian cancer cells to distinct extracellular matrix (ECM) proteins immobilized on gold-coated quartz crystal resonators. The QCM was operated in the impedance analysis mode, where frequency shift as well as bandwidth are accessible on a broad range of overtones. The increase in bandwidth caused by covering the quartz resonator with cells was reproducible and largely independent of overtone order, whereas the frequency shift displayed some variability. Thus the bandwidth proved to be the more robust parameter for sensing cell adhesive events. The bandwidth increased in proportion to the number of seeded cells to the quartz crystal as long as the number was below 150,000 cells/ml. Comparing the resonance parameters on different harmonics, one finds that viscoelastic modeling with homogeneous layer systems cannot reproduce the results: lateral heterogeneity has to be taken into account. The differences in adhesive strength of human ovarian cancer cells towards selected ECM proteins monitored by QCM was in good agreement with data obtained by conventional cell adhesion assays. Strong cell adhesion was observed to the ECM proteins vitronectin (VN) and fibronectin (FN), while only weak attachment occurred on laminin. In order to prove specific, integrin alphavbeta3-mediated cell adhesion to its ligands FN and VN, the cyclic integrin alphavbeta3-directed peptide c(RGDfV) was used as competitor and significantly reversed cell adhesion. Since integrin interaction with ECM proteins is dependent on the presence of bivalent cations, cell detachment was also seen after treatment of cell monolayers with the chelator ethylene-dinitro-tetra-acetic acid (EDTA). The QCM technique is a reliable method to monitor cell adsorption to ECM-pretreated surfaces in real time. It may be an alternative tool for screening specific and selective antagonists of integrin/ECM interaction.

Strychnine-blocked glycine receptor is removed from synapses by a shift in insertion/degradation equilibrium.

PubMed

Rasmussen, Hanne; Rasmussen, Trine; Triller, Antoine; Vannier, Christian

2002-02-01

The long-term inhibition by strychnine of glycine receptor activity in neurons provokes the receptor's selective intracellular accumulation and disappearance from synapses. This could result either from a disruption of the postsynaptic anchoring of the receptor or from an arrest of its exocytic transport. In this study we combined biochemical and fluorescence microscopy analyses to determine on a short time scale the fate of the strychnine-inactivated glycine receptor. Quantification of the cellular content of receptor showed that the rapid accumulation depends on protein synthesis. Cell surface biotinylation of neurons demonstrated that strychnine did not accelerate the turnover rate of the receptor. Labeling of endosomes indicated that, in strychnine-treated cells, the accumulated receptor is not blocked in the endosomal transport pathway. Taken together, these results indicate that strychnine does not destabilize the postsynaptic receptor but triggers its disappearance from synapses by a nondegradative sequestration of newly synthesized molecules in a nonendocytic compartment. (C)2002 Elsevier Science (USA).

Cholesteryl butyrate solid lipid nanoparticles inhibit the adhesion and migration of colon cancer cells

PubMed Central

Minelli, R; Serpe, L; Pettazzoni, P; Minero, V; Barrera, G; Gigliotti, CL; Mesturini, R; Rosa, AC; Gasco, P; Vivenza, N; Muntoni, E; Fantozzi, R; Dianzani, U; Zara, GP; Dianzani, C

2012-01-01

BACKGROUND AND PURPOSE Cholesteryl butyrate solid lipid nanoparticles (cholbut SLN) provide a delivery system for the anti-cancer drug butyrate. These SLN inhibit the adhesion of polymorphonuclear cells to the endothelium and may act as anti-inflammatory agents. As cancer cell adhesion to endothelium is crucial for metastasis dissemination, here we have evaluated the effect of cholbut SLN on adhesion and migration of cancer cells. EXPERIMENTAL APPROACH Cholbut SLN was incubated with a number of cancer cell lines or human umbilical vein endothelial cells (HUVEC) and adhesion was quantified by a computerized micro-imaging system. Migration was detected by the scratch ‘wound-healing’ assay and the Boyden chamber invasion assay. Expression of ERK and p38 MAPK was analysed by Western blot. Expression of the mRNA for E-cadherin and claudin-1 was measured by RT-PCR. KEY RESULTS Cholbut SLN inhibited HUVEC adhesiveness to cancer cell lines derived from human colon–rectum, breast, prostate cancers and melanoma. The effect was concentration and time-dependent and exerted on both cancer cells and HUVEC. Moreover, these SLN inhibited migration of cancer cells and substantially down-modulated ERK and p38 phosphorylation. The anti-adhesive effect was additive to that induced by the triggering of B7h, which is another stimulus inhibiting both ERK and p38 phosphorylation, and cell adhesiveness. Furthermore, cholbut SLN induced E-cadherin and inhibited claudin-1 expression in HUVEC. CONCLUSION AND IMPLICATIONS These results suggest that cholbut SLN could act as an anti-metastastic agent and they add a new mechanism to the anti-tumour activity of this multifaceted preparation of butyrate. PMID:22049973

Enhanced cell adhesion on bioinert ceramics mediated by the osteogenic cell membrane enzyme alkaline phosphatase.

PubMed

Aminian, Alieh; Shirzadi, Bahareh; Azizi, Zahra; Maedler, Kathrin; Volkmann, Eike; Hildebrand, Nils; Maas, Michael; Treccani, Laura; Rezwan, Kurosch

2016-12-01

Functional bone and dental implant materials are required to guide cell response, offering cues that provide specific instructions to cells at the implant/tissue interface while maintaining full biocompatibility as well as the desired structural requirements and functions. In this work we investigate the influence of covalently immobilized alkaline phosphatase (ALP), an enzyme involved in bone mineralization, on the first contact and initial cell adhesion. To this end, ALP is covalently immobilized by carbodiimide-mediated chemoligation on two highly bioinert ceramics, alpha-alumina (Al2O3) and yttria-stabilized zirconia (Y-TZP) that are well-established for load-bearing applications. The physicochemical surface properties are evaluated by profilometry, zeta potential and water contact angle measurements. The initial cell adhesion of human osteoblasts (HOBs), human osteoblast-like cells (MG-63) and mesenchymal stromal cells (hMSCs) was investigated. Cell adhesion was assessed at serum free condition via quantification of percentage of adherent cells, adhesion area and staining of the focal adhesion protein vinculin. Our findings show that after ALP immobilization, the Al2O3 and Y-TZP surfaces gained a negative charge and their hydrophilicity was increased. In the presence of surface-immobilized ALP, a higher cell adhesion, more pronounced cell spreading and a higher number of focal contact points were found. Thereby, this work gives evidence that surface functionalization with ALP can be utilized to modify inert materials for biological conversion and faster bone regeneration on inert and potentially load-bearing implant materials. Copyright © 2016 Elsevier B.V. All rights reserved.

Intracellular targeting of annexin A2 inhibits tumor cell adhesion, migration, and in vivo grafting.

PubMed

Staquicini, Daniela I; Rangel, Roberto; Guzman-Rojas, Liliana; Staquicini, Fernanda I; Dobroff, Andrey S; Tarleton, Christy A; Ozbun, Michelle A; Kolonin, Mikhail G; Gelovani, Juri G; Marchiò, Serena; Sidman, Richard L; Hajjar, Katherine A; Arap, Wadih; Pasqualini, Renata

2017-06-26

Cytoskeletal-associated proteins play an active role in coordinating the adhesion and migration machinery in cancer progression. To identify functional protein networks and potential inhibitors, we screened an internalizing phage (iPhage) display library in tumor cells, and selected LGRFYAASG as a cytosol-targeting peptide. By affinity purification and mass spectrometry, intracellular annexin A2 was identified as the corresponding binding protein. Consistently, annexin A2 and a cell-internalizing, penetratin-fused version of the selected peptide (LGRFYAASG-pen) co-localized and specifically accumulated in the cytoplasm at the cell edges and cell-cell contacts. Functionally, tumor cells incubated with LGRFYAASG-pen showed disruption of filamentous actin, focal adhesions and caveolae-mediated membrane trafficking, resulting in impaired cell adhesion and migration in vitro. These effects were paralleled by a decrease in the phosphorylation of both focal adhesion kinase (Fak) and protein kinase B (Akt). Likewise, tumor cells pretreated with LGRFYAASG-pen exhibited an impaired capacity to colonize the lungs in vivo in several mouse models. Together, our findings demonstrate an unrecognized functional link between intracellular annexin A2 and tumor cell adhesion, migration and in vivo grafting. Moreover, this work uncovers a new peptide motif that binds to and inhibits intracellular annexin A2 as a candidate therapeutic lead for potential translation into clinical applications.

Dynamic Adhesion of Umbilical Cord Blood Endothelial Progenitor Cells under Laminar Shear Stress

PubMed Central

Angelos, Mathew G.; Brown, Melissa A.; Satterwhite, Lisa L.; Levering, Vrad W.; Shaked, Natan T.; Truskey, George A.

2010-01-01

Late outgrowth endothelial progenitor cells (EPCs) represent a promising cell source for rapid reendothelialization of damaged vasculature after expansion ex vivo and injection into the bloodstream. We characterized the dynamic adhesion of umbilical-cord-blood-derived EPCs (CB-EPCs) to surfaces coated with fibronectin. CB-EPC solution density affected the number of adherent cells and larger cells preferentially adhered at lower cell densities. The number of adherent cells varied with shear stress, with the maximum number of adherent cells and the shear stress at maximum adhesion depending upon fluid viscosity. CB-EPCs underwent limited rolling, transiently tethering for short distances before firm arrest. Immediately before arrest, the instantaneous velocity decreased independent of shear stress. A dimensional analysis indicated that adhesion was a function of the net force on the cells, the ratio of cell diffusion to sliding speed, and molecular diffusivity. Adhesion was not limited by the settling rate and was highly specific to α5β1 integrin. Total internal reflection fluorescence microscopy showed that CB-EPCs produced multiple contacts of α5β1 with the surface and the contact area grew during the first 20 min of attachment. These results demonstrate that CB-EPC adhesion from blood can occur under physiological levels of shear stress. PMID:21112278

Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions

NASA Astrophysics Data System (ADS)

Doyle, Andrew D.; Carvajal, Nicole; Jin, Albert; Matsumoto, Kazue; Yamada, Kenneth M.

2015-11-01

The physical properties of two-dimensional (2D) extracellular matrices (ECMs) modulate cell adhesion dynamics and motility, but little is known about the roles of local microenvironmental differences in three-dimensional (3D) ECMs. Here we generate 3D collagen gels of varying matrix microarchitectures to characterize their regulation of 3D adhesion dynamics and cell migration. ECMs containing bundled fibrils demonstrate enhanced local adhesion-scale stiffness and increased adhesion stability through balanced ECM/adhesion coupling, whereas highly pliable reticular matrices promote adhesion retraction. 3D adhesion dynamics are locally regulated by ECM rigidity together with integrin/ECM association and myosin II contractility. Unlike 2D migration, abrogating contractility stalls 3D migration regardless of ECM pore size. We find force is not required for clustering of activated integrins on 3D native collagen fibrils. We propose that efficient 3D migration requires local balancing of contractility with ECM stiffness to stabilize adhesions, which facilitates the detachment of activated integrins from ECM fibrils.

PKC-Dependent Human Monocyte Adhesion Requires AMPK and Syk Activation

PubMed Central

Chang, Mei-Ying; Huang, Duen-Yi; Ho, Feng-Ming; Huang, Kuo-Chin; Lin, Wan-Wan

2012-01-01

PKC plays a pivotal role in mediating monocyte adhesion; however, the underlying mechanisms of PKC-mediated cell adhesion are still unclear. In this study, we elucidated the signaling network of phorbol ester PMA-stimulated human monocyte adhesion. Our results with pharmacological inhibitors suggested the involvement of AMPK, Syk, Src and ERK in PKC-dependent adhesion of THP-1 monocytes to culture plates. Biochemical analysis further confirmed the ability of PMA to activate these kinases, as well as the involvement of AMPK-Syk-Src signaling in this event. Direct protein interaction between AMPK and Syk, which requires the kinase domain of AMPK and linker region of Syk, was observed following PMA stimulation. Notably, we identified Syk as a novel downstream target of AMPK; AICAR can induce Syk phosphorylation at Ser178 and activation of this kinase. However, activation of AMPK alone, either by stimulation with AICAR or by overexpression, is not sufficient to induce monocyte adhesion. Studies further demonstrated that PKC-mediated ERK signaling independent of AMPK activation is also involved in cell adhesion. Moreover, AMPK, Syk, Src and ERK signaling were also required for PMA to induce THP-1 cell adhesion to endothelial cells as well as to induce adhesion response of human primary monocytes. Taken together, we propose a bifurcated kinase signaling pathway involved in PMA-mediated adhesion of monocytes. PKC can activate LKB1/AMPK, leading to phosphorylation and activation of Syk, and subsequent activation of Src and FAK. In addition, PKC-dependent ERK activation induces a coordinated signal for cytoskeleton rearrangement and cell adhesion. For the first time we demonstrate Syk as a novel substrate target of AMPK, and shed new light on the role of AMPK in monocyte adhesion, in addition to its well identified functions in energy homeostasis. PMID:22848421

Peptide array-based interaction assay of solid-bound peptides and anchorage-dependant cells and its effectiveness in cell-adhesive peptide design.

PubMed

Kato, Ryuji; Kaga, Chiaki; Kunimatsu, Mitoshi; Kobayashi, Takeshi; Honda, Hiroyuki

2006-06-01

Peptide array, the designable peptide library covalently synthesized on cellulose support, was applied to assay peptide-cell interaction, between solid-bound peptides and anchorage-dependant cells, to study objective peptide design. As a model case, cell-adhesive peptides that could enhance cell growth as tissue engineering scaffold material, was studied. On the peptide array, the relative cell-adhesion ratio of NIH/3T3 cells was 2.5-fold higher on the RGDS (Arg-Gly-Asp-Ser) peptide spot as compared to the spot with no peptide, thus indicating integrin-mediated peptide-cell interaction. Such strong cell adhesion mediated by the RGDS peptide was easily disrupted by single residue substitution on the peptide array, thus indicating that the sequence recognition accuracy of cells was strictly conserved in our optimized scheme. The observed cellular morphological extension with active actin stress-fiber on the RGD motif-containing peptide supported our strategy that peptide array-based interaction assay of solid-bound peptide and anchorage-dependant cells (PIASPAC) could provide quantitative data on biological peptide-cell interaction. The analysis of 180 peptides obtained from fibronectin type III domain (no. 1447-1629) yielded 18 novel cell-adhesive peptides without the RGD motif. Taken together with the novel candidates, representative rules of ineffective amino acid usage were obtained from non-effective candidate sequences for the effective designing of cell-adhesive peptides. On comparing the amino acid usage of the top 20 and last 20 peptides from the 180 peptides, the following four brief design rules were indicated: (i) Arg or Lys of positively charged amino acids (except His) could enhance cell adhesion, (ii) small hydrophilic amino acids are favored in cell-adhesion peptides, (iii) negatively charged amino acids and small amino acids (except Gly) could reduce cell adhesion, and (iv) Cys and Met could be excluded from the sequence combination since they have less influence on the peptide design. Such rules that are indicative of the nature of the functional peptide sequence can be obtained only by the mass comparison analysis of PIASPAC using peptide array. By following such indicative rules, numerous amino acid combinations can be effectively screened for further examination of novel peptide design.

Effect of salivary secretory IgA on the adhesion of Candida albicans to polystyrene.

PubMed

San Millán, R; Elguezabal, N; Regúlez, P; Moragues, M D; Quindós, G; Pontón, J

2000-09-01

Attachment of Candida albicans to plastic materials of dental prostheses or to salivary macromolecules adsorbed on their surface is believed to be a critical event in the development of denture stomatitis. In an earlier study, it was shown that adhesion of C. albicans to polystyrene, a model system to study the adhesion of C. albicans to plastic materials, can be partially inhibited with an mAb directed against cell wall polysaccharides of C. albicans. In the present study, the role of whole saliva in the adhesion of C. albicans to polystyrene has been investigated, and three mAbs directed against epitopes of cell wall mannoproteins have been used to mimic the inhibitory effect observed with salivary secretory IgA (sIgA) on the adhesion of C. albicans to polystyrene. In the absence of whole saliva, adherence of C. albicans 3153 increased with germination. However, the presence of whole saliva enhanced the adhesion to polystyrene of C. albicans 3153 yeast cells but decreased the adhesion of germinated cells. The enhancement of adhesion of yeast cells to polystyrene mediated by saliva was confirmed with an agerminative mutant of C. albicans 3153. The inhibition of the adhesion of C. albicans 3153 germ tubes to polystyrene was due to the salivary sIgA since sIgA-depleted saliva enhanced the adhesion of C. albicans 3153 to polystyrene. The inhibitory effect mediated by sIgA was not related to the inhibition of germination but to the blockage of adhesins expressed on the cell wall surface of the germ tubes. The three mAbs studied reduced the adhesion of C. albicans 3153 to polystyrene at levels equivalent to those for purified sIgA. The highest reduction in the adhesion was obtained with the IgA mAb N3B. The best results were obtained when the three mAbs were combined. The results suggest that whole saliva plays a different role in the adhesion of C. albicans to polystyrene depending on the morphological phase of C. albicans. These results may give new insights into the conflicting role of saliva in the adhesion of C. albicans to plastic materials of dental prostheses.

Adhesion signaling promotes protease‑driven polyploidization of glioblastoma cells.

PubMed

Mercapide, Javier; Lorico, Aurelio

2014-11-01

An increase in ploidy (polyploidization) causes genomic instability in cancer. However, the determinants for the increased DNA content of cancer cells have not yet been fully elucidated. In the present study, we investigated whether adhesion induces polyploidization in human U87MG glioblastoma cells. For this purpose, we employed expression vectors that reported transcriptional activation by signaling networks implicated in cancer. Signaling activation induced by intercellular integrin binding elicited both extracellular signal‑regulated kinase (ERK) and Notch target transcription. Upon the prolonged activation of both ERK and Notch target transcription induced by integrin binding to adhesion protein, cell cultures accumulated polyploid cells, as determined by cell DNA content distribution analysis and the quantification of polynucleated cells. This linked the transcriptional activation induced by integrin adhesion to the increased frequency of polyploidization. Accordingly, the inhibition of signaling decreased the extent of polyploidization mediated by protease‑driven intracellular invasion. Therefore, the findings of this study indicate that integrin adhesion induces polyploidization through the stimulation of glioblastoma cell invasiveness.

Reelin promotes the adhesion and drug resistance of multiple myeloma cells via integrin β1 signaling and STAT3

PubMed Central

Lv, Meng; Liang, Xiaodong; Dai, Hui; Qin, Xiaodan; Zhang, Yan; Hao, Jie; Sun, Xiuyuan; Yin, Yanhui; Huang, Xiaojun; Zhang, Jun; Lu, Jin; Ge, Qing

2016-01-01

Reelin is an extracellular matrix (ECM) protein that is essential for neuron migration and positioning. The expression of reelin in multiple myeloma (MM) cells and its association with cell adhesion and survival were investigated. Overexpression, siRNA knockdown, and the addition of recombinant protein of reelin were used to examine the function of reelin in MM cells. Clinically, high expression of reelin was negatively associated with progression-free survival and overall survival. Functionally, reelin promoted the adhesion of MM cells to fibronectin via activation of α5β1 integrin. The resulting phosphorylation of Focal Adhesion Kinase (FAK) led to the activation of Src/Syk/STAT3 and Akt, crucial signaling molecules involved in enhancing cell adhesion and protecting cells from drug-induced cell apoptosis. These findings indicate reelin's important role in the activation of integrin-β1 and STAT3/Akt pathways in multiple myeloma and highlight the therapeutic potential of targeting reelin/integrin/FAK axis. PMID:26848618

Reelin promotes the adhesion and drug resistance of multiple myeloma cells via integrin β1 signaling and STAT3.

PubMed

Lin, Liang; Yan, Fan; Zhao, Dandan; Lv, Meng; Liang, Xiaodong; Dai, Hui; Qin, Xiaodan; Zhang, Yan; Hao, Jie; Sun, Xiuyuan; Yin, Yanhui; Huang, Xiaojun; Zhang, Jun; Lu, Jin; Ge, Qing

2016-03-01

Reelin is an extracellular matrix (ECM) protein that is essential for neuron migration and positioning. The expression of reelin in multiple myeloma (MM) cells and its association with cell adhesion and survival were investigated. Overexpression, siRNA knockdown, and the addition of recombinant protein of reelin were used to examine the function of reelin in MM cells. Clinically, high expression of reelin was negatively associated with progression-free survival and overall survival. Functionally, reelin promoted the adhesion of MM cells to fibronectin via activation of α5β1 integrin. The resulting phosphorylation of Focal Adhesion Kinase (FAK) led to the activation of Src/Syk/STAT3 and Akt, crucial signaling molecules involved in enhancing cell adhesion and protecting cells from drug-induced cell apoptosis. These findings indicate reelin's important role in the activation of integrin-β1 and STAT3/Akt pathways in multiple myeloma and highlight the therapeutic potential of targeting reelin/integrin/FAK axis.

Fractalkine and CX3CR1 Mediate a Novel Mechanism of Leukocyte Capture, Firm Adhesion, and Activation under Physiologic Flow

PubMed Central

Fong, Alan M.; Robinson, Lisa A.; Steeber, Douglas A.; Tedder, Thomas F.; Yoshie, Osamu; Imai, Toshio; Patel, Dhavalkumar D.

1998-01-01

Leukocyte migration into sites of inflammation involves multiple molecular interactions between leukocytes and vascular endothelial cells, mediating sequential leukocyte capture, rolling, and firm adhesion. In this study, we tested the role of molecular interactions between fractalkine (FKN), a transmembrane mucin-chemokine hybrid molecule expressed on activated endothelium, and its receptor (CX3CR1) in leukocyte capture, firm adhesion, and activation under physiologic flow conditions. Immobilized FKN fusion proteins captured resting peripheral blood mononuclear cells at physiologic wall shear stresses and induced firm adhesion of resting monocytes, resting and interleukin (IL)-2–activated CD8+ T lymphocytes and IL-2–activated NK cells. FKN also induced cell shape change in firmly adherent monocytes and IL-2–activated lymphocytes. CX3CR1-transfected K562 cells, but not control K562 cells, firmly adhered to FKN-expressing ECV-304 cells (ECV-FKN) and tumor necrosis factor α–activated human umbilical vein endothelial cells. This firm adhesion was not inhibited by pertussis toxin, EDTA/EGTA, or antiintegrin antibodies, indicating that the firm adhesion was integrin independent. In summary, FKN mediated the rapid capture, integrin-independent firm adhesion, and activation of circulating leukocytes under flow. Thus, FKN and CX3CR1 mediate a novel pathway for leukocyte trafficking. PMID:9782118

Cytotoxic-T-Lymphocyte Antigen 4 Receptor Signaling for Lymphocyte Adhesion Is Mediated by C3G and Rap1

PubMed Central

Kloog, Yoel

2014-01-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders. PMID:24396067

Continuum-level modelling of cellular adhesion and matrix production in aggregates.

PubMed

Geris, Liesbet; Ashbourn, Joanna M A; Clarke, Tim

2011-05-01

Key regulators in tissue-engineering processes such as cell culture and cellular organisation are the cell-cell and cell-matrix interactions. As mathematical models are increasingly applied to investigate biological phenomena in the biomedical field, it is important, for some applications, that these models incorporate an adequate description of cell adhesion. This study describes the development of a continuum model that represents a cell-in-gel culture system used in bone-tissue engineering, namely that of a cell aggregate embedded in a hydrogel. Cell adhesion is modelled through the use of non-local (integral) terms in the partial differential equations. The simulation results demonstrate that the effects of cell-cell and cell-matrix adhesion are particularly important for the survival and growth of the cell population and the production of extracellular matrix by the cells, concurring with experimental observations in the literature.

The role of basal cells in adhesion of columnar epithelium to airway basement membrane.

PubMed

Evans, M J; Plopper, C G

1988-08-01

In this report, we present a new concept of the role of the basal cell in airway epithelium. Previously, the basal cell was thought to be the progenitor cell for the columnar epithelium. However, several studies have shown that this concept may not be correct. The morphologic aspects of the basal cell suggest that it could play a role in adhesion of the columnar epithelium to the basement membrane. Basal cells form attachments with columnar cells (desmosomes) and with the basement membrane (hemidesmosomes). Columnar cells do not form hemidesmosome attachments with the basement membrane. Basal cells could strengthen the adhesion of columnar cells to the basement membrane by forming hemidesmosome attachments to the basement membrane and desmosome attachments with adjacent columnar cells. Incidental evidence from 2 existing publications concerning airway microanatomy support this concept. As columnar cells grow taller, the proportion of the cell surface in contact with the basement membrane becomes progressively smaller, and thus the cell surface area related to adhesion also becomes smaller. It was found that the number of basal cells per millimeter of basement membrane was closely related to the height of the columnar cell epithelium (r = 0.98), but not to the number of columnar cells (r = 0.42). The consistency of the relationship between increased columnar cell height (and thus decreased surface area for adhesion) and the number of basal cells present (r = 0.98) supports the concept that the basal cell plays a role in adhesion of columnar cells to the basement membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

CD15s/CD62E Interaction Mediates the Adhesion of Non-Small Cell Lung Cancer Cells on Brain Endothelial Cells: Implications for Cerebral Metastasis.

PubMed

Jassam, Samah A; Maherally, Zaynah; Smith, James R; Ashkan, Keyoumars; Roncaroli, Federico; Fillmore, Helen L; Pilkington, Geoffrey J

2017-07-10

Expression of the cell adhesion molecule (CAM), Sialyl Lewis X (CD15s) correlates with cancer metastasis, while expression of E-selectin (CD62E) is stimulated by TNF-α. CD15s/CD62E interaction plays a key role in the homing process of circulating leukocytes. We investigated the heterophilic interaction of CD15s and CD62E in brain metastasis-related cancer cell adhesion. CD15s and CD62E were characterised in human brain endothelium (hCMEC/D3), primary non-small cell lung cancer (NSCLC) (COR-L105 and A549) and metastatic NSCLC (SEBTA-001 and NCI-H1299) using immunocytochemistry, Western blotting, flow cytometry and immunohistochemistry in human brain tissue sections. TNF-α (25 pg/mL) stimulated extracellular expression of CD62E while adhesion assays, under both static and physiological flow live-cell conditions, explored the effect of CD15s-mAb immunoblocking on adhesion of cancer cell-brain endothelium. CD15s was faintly expressed on hCMEC/D3, while high levels were observed on primary NSCLC cells with expression highest on metastatic NSCLC cells ( p < 0.001). CD62E was highly expressed on hCMEC/D3 cells activated with TNF-α, with lower levels on primary and metastatic NSCLC cells. CD15s and CD62E were expressed on lung metastatic brain biopsies. CD15s/CD62E interaction was localised at adhesion sites of cancer cell-brain endothelium. CD15s immunoblocking significantly decreased cancer cell adhesion to brain endothelium under static and shear stress conditions ( p < 0.001), highlighting the role of CD15s-CD62E interaction in brain metastasis.

Amphiphilic cationic peptides mediate cell adhesion to plastic surfaces.

PubMed

Rideout, D C; Lambert, M; Kendall, D A; Moe, G R; Osterman, D G; Tao, H P; Weinstein, I B; Kaiser, E T

1985-09-01

Four amphiphilic peptides, each with net charges of +2 or more at neutrality and molecular weights under 4 kilodaltons, were found to mediate the adhesion of normal rat kidney fibroblasts to polystyrene surfaces. Two of these peptides, a model for calcitonin (peptide 1, MCT) and melittin (peptide 2, MEL), form amphiphilic alpha-helical structures at aqueous/nonpolar interfaces. The other two, a luteinizing hormone-releasing hormone model (peptide 3, LHM) and a platelet factor model (peptide 4, MPF) form beta-strand structures in amphiphilic environments. Although it contains only 10 residues, LHM mediated adhesion to surfaces coated with solutions containing as little as 10 pmoles/ml of peptide. All four of these peptides were capable of forming monolayers at air-buffer interfaces with collapse pressures greater than 20 dynes/cm. None of these four peptides contains the tetrapeptide sequence Arg-Gly-Asp-Ser, which has been associated with fibronectin-mediated cell adhesion. Ten polypeptides that also lacked the sequence Arg-Gly-Asp-Ser but were nonamphiphilic and/or had net charges less than +2 at neutrality were all incapable of mediating cell adhesion (Pierschbacher and Ruoslahti, 1984). The morphologies of NRK cells spread on polystyrene coated with peptide LHM resemble the morphologies on fibronectin-coated surfaces, whereas cells spread on surfaces coated with MCT or MEL exhibit strikingly different morphologies. The adhesiveness of MCT, MEL, LHM, and MPF implies that many amphiphilic cationic peptides could prove useful as well defined adhesive substrata for cell culture and for studies of the mechanism of cell adhesion.

Requirement of the actin cytoskeleton for the association of nectins with other cell adhesion molecules at adherens and tight junctions in MDCK cells.

PubMed

Yamada, Akio; Irie, Kenji; Fukuhara, Atsunori; Ooshio, Takako; Takai, Yoshimi

2004-09-01

Nectins, Ca(2+)-independent immunoglobulin-like cell adhesion molecules (CAMs), first form cell-cell adhesion where cadherins are recruited, forming adherens junctions (AJs) in epithelial cells and fibroblasts. In addition, nectins recruit claudins, occludin, and junctional adhesion molecules (JAMs) to the apical side of AJs, forming tight junctions (TJs) in epithelial cells. Nectins are associated with these CAMs through peripheral membrane proteins (PMPs), many of which are actin filament-binding proteins. We examined here the roles of the actin cytoskeleton in the association of nectins with other CAMs in MDCK cells stably expressing exogenous nectin-1. The nectin-1-based cell-cell adhesion was formed and maintained irrespective of the presence and absence of the actin filament-disrupting agents, such as cytochalasin D and latrunculin A. In the presence of these agents, only afadin remained at the nectin-1-based cell-cell adhesion sites, whereas E-cadherin and other PMPs at AJs, alpha-catenin, beta-catenin, vinculin, alpha-actinin, ADIP, and LMO7, were not concentrated there. The CAMs at TJs, claudin-1, occludin and JAM-1, or the PMPs at TJs, ZO-1 and MAGI-1, were not concentrated there, either. These results indicate that the actin cytoskeleton is required for the association of the nectin-afadin unit with other CAMs and PMPs at AJs and TJs.

Light-triggered in vivo activation of adhesive peptides regulates cell adhesion, inflammation and vascularization of biomaterials.

PubMed

Lee, Ted T; García, José R; Paez, Julieta I; Singh, Ankur; Phelps, Edward A; Weis, Simone; Shafiq, Zahid; Shekaran, Asha; Del Campo, Aránzazu; García, Andrés J

2015-03-01

Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control. Although materials with temporally regulated presentation of bioadhesive ligands using external triggers, such as light and electric fields, have recently been realized for cells in culture, the impact of in vivo temporal ligand presentation on cell-material responses is unknown. Here, we present a general strategy to temporally and spatially control the in vivo presentation of bioligands using cell-adhesive peptides with a protecting group that can be easily removed via transdermal light exposure to render the peptide fully active. We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material. This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

Light-triggered in vivo activation of adhesive peptides regulates cell adhesion, inflammation and vascularization of biomaterials

NASA Astrophysics Data System (ADS)

Lee, Ted T.; García, José R.; Paez, Julieta I.; Singh, Ankur; Phelps, Edward A.; Weis, Simone; Shafiq, Zahid; Shekaran, Asha; Del Campo, Aránzazu; García, Andrés J.

2015-03-01

Materials engineered to elicit targeted cellular responses in regenerative medicine must display bioligands with precise spatial and temporal control. Although materials with temporally regulated presentation of bioadhesive ligands using external triggers, such as light and electric fields, have recently been realized for cells in culture, the impact of in vivo temporal ligand presentation on cell-material responses is unknown. Here, we present a general strategy to temporally and spatially control the in vivo presentation of bioligands using cell-adhesive peptides with a protecting group that can be easily removed via transdermal light exposure to render the peptide fully active. We demonstrate that non-invasive, transdermal time-regulated activation of cell-adhesive RGD peptide on implanted biomaterials regulates in vivo cell adhesion, inflammation, fibrous encapsulation, and vascularization of the material. This work shows that triggered in vivo presentation of bioligands can be harnessed to direct tissue reparative responses associated with implanted biomaterials.

Surface nanoporosity has a greater influence on osteogenic and bacterial cell adhesion than crystallinity and wettability

NASA Astrophysics Data System (ADS)

Rodriguez-Contreras, Alejandra; Guadarrama Bello, Dainelys; Nanci, Antonio

2018-07-01

There has been much emphasis on the influence of crystallinity and wettability for modulating cell activity, particularly for bone biomaterials. In this context, we have generated titanium oxide layers with similar mesoporous topography and surface roughness but with amorphous or crystalline oxide layers and differential wettability. We then investigated their influence on the behavior of MC3T3 osteoblastic and bacterial cells. There was no difference in cell adhesion, spreading and growth on amorphous and crystalline surfaces. The number of focal adhesions was similar, however, cells on the amorphous surface exhibited a higher frequency of mature adhesions. The crystallinity of the surface layers also had no bearing on bacterial adhesion. While it cannot be excluded that surface crystallinity, roughness and wettability contribute to some degree to determining cell behavior, our data suggest that physical characteristics of surfaces represent the major determinant.

Preparation and regulating cell adhesion of anion-exchangeable layered double hydroxide micropatterned arrays.

PubMed

Yao, Feng; Hu, Hao; Xu, Sailong; Huo, Ruijie; Zhao, Zhiping; Zhang, Fazhi; Xu, Fujian

2015-02-25

We describe a reliable preparation of MgAl-layered double hydroxide (MgAl-LDH) micropatterned arrays on gold substrate by combining SO3(-)-terminated self-assembly monolayer and photolithography. The synthesis route is readily extended to prepare LDH arrays on the SO3(-)-terminated polymer-bonded glass substrate amenable for cell imaging. The anion-exchangeable MgAl-LDH micropattern can act both as bioadhesive region for selective cell adhesion and as nanocarrier for drug molecules to regulate cell behaviors. Quantitative analysis of cell adhesion shows that selective HepG2 cell adhesion and spreading are promoted by the micropatterned MgAl-LDH, and also suppressed by methotrexate drug released from the LDH interlayer galleries.

Cytotoxic effects of denture adhesives on primary human oral keratinocytes, fibroblasts and permanent L929 cell lines.

PubMed

Chen, Fengying; Wu, Tianfu; Cheng, Xiangrong

2014-03-01

To date, there have been very little data on the cytotoxic responses of different cell lines to denture adhesives. To determine the cytotoxicity of three denture adhesives on primary human oral keratinocytes (HOKs), fibroblasts (HOFs) and permanent mouse fibroblasts cell lines (L929). Three commercial denture adhesives (two creams and one powder) were prepared for indirect contact using the agar diffusion test, as well as extracts in MTT assay. The results of the MTT assay were statistically analysed by one-way anova and Tukey's test (p < 0.05). All of the tested denture adhesives showed mild to moderate cytotoxicity to primary HOKs (p < 0.001), whereas none of three was toxic to L929 cells (p > 0.05) in both assays. For primary HOFs cultures, slight cytotoxicity was observed for one of the products from the agar diffusion test and undiluted eluates of all tested adhesives with MTT assay (p < 0.01). Denture adhesives are toxic to the primary HOKs and HOFs cultures, whereas non-toxic to L929 cells. The results suggest that primary human oral mucosal cells may provide more valuable information in toxicity screening of denture adhesives. © 2012 John Wiley & Sons A/S and The Gerodontology Association. Published by John Wiley & Sons Ltd.

CHRONIC HYPERTENSION ENHANCES PRE-SYNAPTIC INHIBITION BY BACLOFEN IN THE NUCLEUS OF THE SOLITARY TRACT

PubMed Central

Zhang, Weirong; Mifflin, Steve

2010-01-01

The selective γ-aminobutyric acid B-subtype receptor agonist baclofen activates both pre- and post-synaptic receptors in the brain. Microinjection of baclofen into the nucleus of the solitary tract increases arterial pressure, heart rate and sympathetic nerve discharge consistent with inhibition of the arterial baroreflex. The magnitude of these responses is enhanced in hypertension and is associated with increased post-synaptic GABAB receptor function. We tested whether a pre-synaptic mechanism contributes to the enhanced baclofen inhibition in hypertension. Whole-cell recordings of second-order baroreceptor neurons, identified by 4-(4-(dihexadecylamino)styryl)-N-methylpyridinium iodide labeling of aortic nerve, were obtained in brainstem slices from normotensive control and renal-wrap hypertensive rats. After 4 weeks, arterial blood pressure was 162±9 mmHg in hypertensive (n=6) and 107±3 mmHg in control rats (n=6/11, p<0.001). Baclofen reduced the amplitude of excitatory post-synaptic currents evoked by solitary tract stimulation and the EC50 of this inhibition was greater in control (1.5±0.5 µmol/L, n=6) than hypertensive cells (0.6±0.1 µmol/L, n=9, p<0.05). Baclofen (1 µmol/L) elicited greater inhibition on evoked response in hypertensive (58±6%, n=9) than control cells (40±6%, n=8, p<0.05). Another index of pre-synaptic inhibition, the paired-pulse ratio (ratio of second to first evoked response amplitudes at stimulus intervals of 40 ms), was greater in hypertensive (0.60±0.08, n=8) than control cells (0.48±0.06. n=5, p<0.05). The results suggest that in renal-wrap hypertensive rats, baclofen causes an enhanced pre-synaptic inhibition of glutamate release from baroreceptor afferent terminals to second-order neurons in the nucleus of the solitary tract. This enhanced pre-synaptic inhibition could contribute to altered baroreflex function in hypertension. PMID:20038748

Hyperpolarization-activated current (I(h)) in vestibular calyx terminals: characterization and role in shaping postsynaptic events.

PubMed

Meredith, Frances L; Benke, Tim A; Rennie, Katherine J

2012-12-01

Calyx afferent terminals engulf the basolateral region of type I vestibular hair cells, and synaptic transmission across the vestibular type I hair cell/calyx is not well understood. Calyces express several ionic conductances, which may shape postsynaptic potentials. These include previously described tetrodotoxin-sensitive inward Na(+) currents, voltage-dependent outward K(+) currents and a K(Ca) current. Here, we characterize an inwardly rectifying conductance in gerbil semicircular canal calyx terminals (postnatal days 3-45), sensitive to voltage and to cyclic nucleotides. Using whole-cell patch clamp, we recorded from isolated calyx terminals still attached to their type I hair cells. A slowly activating, noninactivating current (I(h)) was seen with hyperpolarizing voltage steps negative to the resting potential. External Cs(+) (1-5 mM) and ZD7288 (100 μM) blocked the inward current by 97 and 83 %, respectively, confirming that I(h) was carried by hyperpolarization-activated, cyclic nucleotide gated channels. Mean half-activation voltage of I(h) was -123 mV, which shifted to -114 mV in the presence of cAMP. Activation of I(h) was well described with a third order exponential fit to the current (mean time constant of activation, τ, was 190 ms at -139 mV). Activation speeded up significantly (τ=136 and 127 ms, respectively) when intracellular cAMP and cGMP were present, suggesting that in vivo I(h) could be subject to efferent modulation via cyclic nucleotide-dependent mechanisms. In current clamp, hyperpolarizing current steps produced a time-dependent depolarizing sag followed by either a rebound afterdepolarization or an action potential. Spontaneous excitatory postsynaptic potentials (EPSPs) became larger and wider when I(h) was blocked with ZD7288. In a three-dimensional mathematical model of the calyx terminal based on Hodgkin-Huxley type ionic conductances, removal of I(h) similarly increased the EPSP, whereas cAMP slightly decreased simulated EPSP size and width.

Holding Tight: Cell Junctions and Cancer Spread.

PubMed

Knights, Alexander J; Funnell, Alister P W; Crossley, Merlin; Pearson, Richard C M

2012-01-01

Cell junctions are sites of intercellular adhesion that maintain the integrity of epithelial tissue and regulate signalling between cells. These adhesive junctions are comprised of protein complexes that serve to establish an intercellular cytoskeletal network for anchoring cells, in addition to regulating cell polarity, molecular transport and communication. The expression of cell adhesion molecules is tightly controlled and their downregulation is essential for epithelial-mesenchymal transition (EMT), a process that facilitates the generation of morphologically and functionally diverse cell types during embryogenesis. The characteristics of EMT are a loss of cell adhesion and increased cellular mobility. Hence, in addition to its normal role in development, dysregulated EMT has been linked to cancer progression and metastasis, the process whereby primary tumors migrate to invasive secondary sites in the body. This paper will review the current understanding of cell junctions and their role in cancer, with reference to the abnormal regulation of junction protein genes. The potential use of cell junction molecules as diagnostic and prognostic markers will also be discussed, as well as possible therapies for adhesive dysregulation.

Real-time, label-free monitoring of cell viability based on cell adhesion measurements with an atomic force microscope.

PubMed

Yang, Fang; Riedel, René; Del Pino, Pablo; Pelaz, Beatriz; Said, Alaa Hassan; Soliman, Mahmoud; Pinnapireddy, Shashank R; Feliu, Neus; Parak, Wolfgang J; Bakowsky, Udo; Hampp, Norbert

2017-03-22

The adhesion of cells to an oscillating cantilever sensitively influences the oscillation amplitude at a given frequency. Even early stages of cytotoxicity cause a change in the viscosity of the cell membrane and morphology, both affecting their adhesion to the cantilever. We present a generally applicable method for real-time, label free monitoring and fast-screening technique to assess early stages of cytotoxicity recorded in terms of loss of cell adhesion. We present data taken from gold nanoparticles of different sizes and surface coatings as well as some reference substances like ethanol, cadmium chloride, and staurosporine. Measurements were recorded with two different cell lines, HeLa and MCF7 cells. The results obtained from gold nanoparticles confirm earlier findings and attest the easiness and effectiveness of the method. The reported method allows to easily adapt virtually every AFM to screen and assess toxicity of compounds in terms of cell adhesion with little modifications as long as a flow cell is available. The sensitivity of the method is good enough indicating that even single cell analysis seems possible.

E-Cadherin-Dependent Stimulation of Traction Force at Focal Adhesions via the Src and PI3K Signaling Pathways

PubMed Central

Jasaitis, Audrius; Estevez, Maruxa; Heysch, Julie; Ladoux, Benoit; Dufour, Sylvie

2012-01-01

The interplay between cadherin- and integrin-dependent signals controls cell behavior, but the precise mechanisms that regulate the strength of adhesion to the extracellular matrix remains poorly understood. We deposited cells expressing a defined repertoire of cadherins and integrins on fibronectin (FN)-coated polyacrylamide gels (FN-PAG) and on FN-coated pillars used as a micro-force sensor array (μFSA), and analyzed the functional relationship between these adhesion receptors to determine how it regulates cell traction force. We found that cadherin-mediated adhesion stimulated cell spreading on FN-PAG, and this was modulated by the substrate stiffness. We compared S180 cells with cells stably expressing different cadherins on μFSA and found that traction forces were stronger in cells expressing cadherins than in parental cells. E-cadherin-mediated contact and mechanical coupling between cells are required for this increase in cell-FN traction force, which was not observed in isolated cells, and required Src and PI3K activities. Traction forces were stronger in cells expressing type I cadherins than in cells expressing type II cadherins, which correlates with our previous observation of a higher intercellular adhesion strength developed by type I compared with type II cadherins. Our results reveal one of the mechanisms whereby molecular cross talk between cadherins and integrins upregulates traction forces at cell-FN adhesion sites, and thus provide additional insight into the molecular control of cell behavior. PMID:22853894

Capping of the N-terminus of PSD-95 by calmodulin triggers its postsynaptic release

PubMed Central

Zhang, Yonghong; Matt, Lucas; Patriarchi, Tommaso; Malik, Zulfiqar A; Chowdhury, Dhrubajyoti; Park, Deborah K; Renieri, Alessandra; Ames, James B; Hell, Johannes W

2014-01-01

Postsynaptic density protein-95 (PSD-95) is a central element of the postsynaptic architecture of glutamatergic synapses. PSD-95 mediates postsynaptic localization of AMPA receptors and NMDA receptors and plays an important role in synaptic plasticity. PSD-95 is released from postsynaptic membranes in response to Ca2+ influx via NMDA receptors. Here, we show that Ca2+/calmodulin (CaM) binds at the N-terminus of PSD-95. Our NMR structure reveals that both lobes of CaM collapse onto a helical structure of PSD-95 formed at its N-terminus (residues 1–16). This N-terminal capping of PSD-95 by CaM blocks palmitoylation of C3 and C5, which is required for postsynaptic PSD-95 targeting and the binding of CDKL5, a kinase important for synapse stability. CaM forms extensive hydrophobic contacts with Y12 of PSD-95. The PSD-95 mutant Y12E strongly impairs binding to CaM and Ca2+-induced release of PSD-95 from the postsynaptic membrane in dendritic spines. Our data indicate that CaM binding to PSD-95 serves to block palmitoylation of PSD-95, which in turn promotes Ca2+-induced dissociation of PSD-95 from the postsynaptic membrane. PMID:24705785

Capping of the N-terminus of PSD-95 by calmodulin triggers its postsynaptic release.

PubMed

Zhang, Yonghong; Matt, Lucas; Patriarchi, Tommaso; Malik, Zulfiqar A; Chowdhury, Dhrubajyoti; Park, Deborah K; Renieri, Alessandra; Ames, James B; Hell, Johannes W

2014-06-17

Postsynaptic density protein-95 (PSD-95) is a central element of the postsynaptic architecture of glutamatergic synapses. PSD-95 mediates postsynaptic localization of AMPA receptors and NMDA receptors and plays an important role in synaptic plasticity. PSD-95 is released from postsynaptic membranes in response to Ca(2+) influx via NMDA receptors. Here, we show that Ca(2+)/calmodulin (CaM) binds at the N-terminus of PSD-95. Our NMR structure reveals that both lobes of CaM collapse onto a helical structure of PSD-95 formed at its N-terminus (residues 1-16). This N-terminal capping of PSD-95 by CaM blocks palmitoylation of C3 and C5, which is required for postsynaptic PSD-95 targeting and the binding of CDKL5, a kinase important for synapse stability. CaM forms extensive hydrophobic contacts with Y12 of PSD-95. The PSD-95 mutant Y12E strongly impairs binding to CaM and Ca(2+)-induced release of PSD-95 from the postsynaptic membrane in dendritic spines. Our data indicate that CaM binding to PSD-95 serves to block palmitoylation of PSD-95, which in turn promotes Ca(2+)-induced dissociation of PSD-95 from the postsynaptic membrane. © 2014 The Authors.

Treating the Synapse in Major Psychiatric Disorders: The Role of Postsynaptic Density Network in Dopamine-Glutamate Interplay and Psychopharmacologic Drugs Molecular Actions

PubMed Central

Tomasetti, Carmine; Iasevoli, Felice; Buonaguro, Elisabetta Filomena; De Berardis, Domenico; Fornaro, Michele; Fiengo, Annastasia Lucia Carmela; Martinotti, Giovanni; Orsolini, Laura; Valchera, Alessandro; Di Giannantonio, Massimo; de Bartolomeis, Andrea

2017-01-01

Dopamine-glutamate interplay dysfunctions have been suggested as pathophysiological key determinants of major psychotic disorders, above all schizophrenia and mood disorders. For the most part, synaptic interactions between dopamine and glutamate signaling pathways take part in the postsynaptic density, a specialized ultrastructure localized under the membrane of glutamatergic excitatory synapses. Multiple proteins, with the role of adaptors, regulators, effectors, and scaffolds compose the postsynaptic density network. They form structural and functional crossroads where multiple signals, starting at membrane receptors, are received, elaborated, integrated, and routed to appropriate nuclear targets. Moreover, transductional pathways belonging to different receptors may be functionally interconnected through postsynaptic density molecules. Several studies have demonstrated that psychopharmacologic drugs may differentially affect the expression and function of postsynaptic genes and proteins, depending upon the peculiar receptor profile of each compound. Thus, through postsynaptic network modulation, these drugs may induce dopamine-glutamate synaptic remodeling, which is at the basis of their long-term physiologic effects. In this review, we will discuss the role of postsynaptic proteins in dopamine-glutamate signals integration, as well as the peculiar impact of different psychotropic drugs used in clinical practice on postsynaptic remodeling, thereby trying to point out the possible future molecular targets of “synapse-based” psychiatric therapeutic strategies. PMID:28085108

Nephrin phosphorylation regulates podocyte adhesion through the PINCH-1-ILK-α-parvin complex

PubMed Central

Zha, Dongqing; Chen, Cheng; Liang, Wei; Chen, Xinghua; Ma, Tean; Yang, Hongxia; van Goor, Harry; Ding, Guohua

2013-01-01

Nephrin, a structural molecule, is also a signaling molecule after phosphorylation. Inhibition of nephrin phosphorylation is correlated with podocyte injury. The PINCH-1-ILK-α-parvin (PIP) complex plays a crucial role in cell adhesion and cytoskeleton formation. We hypothesized that nephrin phosphorylation influenced cytoskeleton and cell adhesion in podocytes by regulating the PIP complex. The nephrin phosphorylation, PIP complex formation, and F-actin in Wistar rats intraperitoneally injected with puromycin aminonucleoside were gradually decreased but increased with time, coinciding with the recovery from glomerular/podocyte injury and proteinuria. In cultured podocytes, PIP complex knockdown resulted in cytoskeleton reorganization and decreased cell adhesion and spreading. Nephrin and its phosphorylation were unaffected after PIP complex knockdown. Furthermore, inhibition of nephrin phosphorylation suppressed PIP complex expression, disorganized podocyte cytoskeleton, and decreased cell adhesion and spreading. These findings indicate that alterations in nephrin phosphorylation disorganize podocyte cytoskeleton and decrease cell adhesion through a PIP complex-dependent mechanism. [BMB Reports 2013; 46(4): 230-235] PMID:23615266

Heme oxygenase-1 protects INF-gamma primed endothelial cells from Jurkat T-cell adhesion.

PubMed

Du, D; Chang, S; Chen, B; Zhou, H; Chen, Z K

2007-12-01

The heme oxygenase-1 (HO-1) system is associated with the rate-limiting step of conversion of heme, one of the most critical roles in cytoprotective mechanisms. Our study investigated its potential role in protection of endothelial cells from T cells. The recombinant plasmid pcDNA3-HO-1 was transfected into endothelial cells. Indirect fluorescent staining was used to examine the expression of HO-1 protein. Then endothelial cells primed by INF-gamma were mixed in culture with Jurkat T cells labeled with carboxyfluorescein diacetate succinimidyl ester (CFSE). The number of adhesive Jurkat T cells was determined using FACS to evaluate the adhesion effect. After being cultured with endothelial cells, the cell cycle of Jurkat T cells was detected using FACS. Expression of HO-1 on endothelial cells conferred significant protection against Jurkat T-cell-mediated adhesion. The rate of Jurkat T-cell adhesions was reduced to 19.06%, in contrast with 31.42% in the control group (P<.05). After using ZnPP, an inhibitor of HO-1, the rate of Jurkat T-cell adhesion recovered to 29.08%. The binding activities between endothelial cells and Jurkat T cells was blocked by HO-1 expression. The proliferation of Jurkat T cells was inhibited after culture with endothelial cells, which had been transfected with HO-1, which blocked cell cycle entry of T cells. More than 60% of Jurkat T cells remained in G0/G1 compared with 40% among the control group. HO-1 directly protected endothelial cells primed by INF-gamma from Jurkat T cells and down-regulated the expression of HLA-DR on the surface of endothelial cells. These results indicated that transgenic expression of HO-1 may be useful to prevent lymphocytes from responding to endothelial cells.

Cytotoxicity, oxidative stress and expression of adhesion molecules in human umbilical vein endothelial cells exposed to dust from paints with or without nanoparticles.

PubMed

Mikkelsen, Lone; Jensen, Keld A; Koponen, Ismo K; Saber, Anne T; Wallin, Håkan; Loft, Steffen; Vogel, Ulla; Møller, Peter

2013-03-01

Nanoparticles in primary form and nanoproducts might elicit different toxicological responses. We compared paint-related nanoparticles with respect to effects on endothelial oxidative stress, cytotoxicity and cell adhesion molecule expression. Primary human umbilical vein endothelial cells were exposed to primary nanoparticles (fine, photocatalytic or nanosized TiO(2), aluminium silicate, carbon black, nano-silicasol or axilate) and dust from sanding reference- or nanoparticle-containing paints. Most of the samples increased cell surface expressions of vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1), but paint sanding dust samples generally generated less response than primary particles of TiO(2) and carbon black. We found no relationship between the expression of adhesion molecules, cytotoxicity and production of reactive oxygen species. In conclusion, sanding dust from nanoparticle-containing paint did not generate more oxidative stress or expression of cell adhesion molecules than sanding dust from paint without nanoparticles, whereas the primary particles had the largest effect on mass basis.

CRYOPRESERVATION EFFECTS ON RECOMBINANT MYOBLASTS ENCAPSULATED IN ADHESIVE ALGINATE HYDROGELS

PubMed Central

Ahmad, Hajira F.; Sambanis, Athanassios

2013-01-01

Cell encapsulation in hydrogels is widely used in tissue engineering applications, including encapsulation of islets or other insulin-secreting cells in pancreatic substitutes. Use of adhesive, bio-functionalized hydrogels is receiving increasing attention, as cell-matrix interactions in 3-D can be important for various cell processes. With pancreatic substitutes, studies have indicated benefits of 3-D adhesion on the viability and/or function of insulin-secreting cells. As long-term storage of microencapsulated cells is critical for their clinical translation, cryopreservation of cells in hydrogels is actively being investigated. Previous studies have examined the cryopreservation response of cells encapsulated in non-adhesive hydrogels using conventional freezing and/or vitrification (ice-free cryopreservation), however, none have systematically compared the two cryopreservation methods with cells encapsulated within an adhesive 3-D environment. The latter would be significant, as evidence suggests adhesion influences cellular response to cryopreservation. Thus, the objective of this study was to determine the response to conventional freezing and vitrification of insulin-secreting cells encapsulated in an adhesive biomimetic hydrogel. Recombinant insulin-secreting C2C12 myoblasts were encapsulated in oxidized RGD-alginate and cultured 1 or 4 days post-encapsulation, cryopreserved, and assessed up to 3 days post-warming for metabolic activity and insulin secretion, and one day post-warming for cell morphology. Besides certain transient differences of the vitrified group relative to the Fresh control, both conventional freezing and vitrification maintained metabolism, secretion and morphology of the recombinant C2C12 cells. Thus, due to a simpler procedure and slightly superior results, conventional freezing is recommended over vitrification for the cryopreservation of C2C12 cells in oxidized RGD-modified alginate. PMID:23499987

Adhesive interactions of human multiple myeloma cell lines with different extracellular matrix molecules.

PubMed

Kibler, C; Schermutzki, F; Waller, H D; Timpl, R; Müller, C A; Klein, G

1998-06-01

Multiple myeloma represents a human B cell malignancy which is characterized by a predominant localization of the malignant cell clone within the bone marrow. With the exception of the terminal stage of the disease the myeloma tumor cells do not circulate in the peripheral blood. The bone marrow microenvironment is believed to play an important role in homing, proliferation and terminal differentiation of myeloma cells. Here we have studied the expression of several extracellular matrix (ECM) molecules in the bone marrow of multiple myeloma patients and analyzed their adhesive capacities with four different human myeloma-derived cell lines. All ECM molecules analyzed (tenascin, laminin, fibronectin, collagen types I, III, V and VI) could be detected in bone marrow cryostat sections of multiple myeloma patients. Adhesion assays showed that only laminin, the microfibrillar collagen type VI and fibronectin were strong adhesive components for the myeloma cell lines U266, IM-9, OPM-2 and NCI-H929. Tenascin and collagen type I were only weak adhesive substrates for these myeloma cells. Adhesion to laminin and fibronectin was beta 1-integrin-mediated since addition of anti-beta 1-integrin antibodies could inhibit the binding of the four different cell types to both matrix molecules. In contrast, integrins do not seem to be involved in binding of the myeloma cells to collagen type VI. Instead, inhibition of binding by heparin suggested that membrane-bound heparan sulfate proteoglycans are responsible ligands for binding to collagen type VI. Adhesion assays with several B-cell lines resembling earlier differentiation stages revealed only weak interactions with tenascin and no interactions with collagen type VI, laminin or fibronectin. In summary, the interactions of human myeloma cells with the extracellular matrix may explain the specific retention of the plasma cells within the bone marrow.

FAK is required for tension-dependent organization of collective cell movements in Xenopus mesendoderm

PubMed Central

Bjerke, Maureen A.; Dzamba, Bette; Wang, Chong; DeSimone, Douglas W.

2014-01-01

Collective cell movements are integral to biological processes such as embryonic development and wound healing and also have a prominent role in some metastatic cancers. In migrating Xenopus mesendoderm, traction forces are generated by cells through integrin-based adhesions and tension transmitted across cadherin adhesions. This is accompanied by assembly of a mechanoresponsive cadherin adhesion complex containing keratin intermediate filaments and the catenin-family member plakoglobin. We demonstrate that focal adhesion kinase (FAK), a major component of integrin adhesion complexes, is required for normal morphogenesis at gastrulation, closure of the anterior neural tube, axial elongation and somitogenesis. Depletion of zygotically expressed FAK results in disruption of mesendoderm tissue polarity similar to that observed when expression of keratin or plakoglobin is inhibited. Both individual and collective migrations of mesendoderm cells from FAK depleted embryos are slowed, cell protrusions are disordered, and cell spreading and traction forces are decreased. Additionally, keratin filaments fail to organize at the rear of cells in the tissue and association of plakoglobin with cadherin is diminished. These findings suggest that FAK is required for the tension-dependent assembly of the cadherin adhesion complex that guides collective mesendoderm migration, perhaps by modulating the dynamic balance of substrate traction forces and cell cohesion needed to establish cell polarity. PMID:25127991

Characterization of adhesive molecule with affinity to Caco-2 cells in Lactobacillus acidophilus by proteome analysis.

PubMed

Ashida, Nobuhisa; Yanagihara, Sae; Shinoda, Tadashi; Yamamoto, Naoyuki

2011-10-01

The adhesive activities of eight Lactobacillus acidophilus strains toward intestinal epithelial Caco-2 cells were studied to understand the probiotic characteristics of the L. acidophilus L-92 strain. Most of the strains, including L-92, showed high adhesive activity; CP23 showed the lowest adhesive activity. CP23 was selected for comparative analysis of cell wall-associated proteins versus the L-92 strain. Cell wall-associated proteins extracted from L-92 and CP23 were subjected to two-dimensional electrophoresis, and major spots observed in the former were compared to the corresponding spots in the latter. To understand the effects of key components of L-92 on its adhesion to Caco-2 cells, 18 spots with stronger signals in L-92 than those in CP23 were identified by a MALDI-TOF/TOF of Ultraflex analysis. Among the identified proteins of L-92, surface-layer protein A (SlpA) was considered strongly involved in adhesion in the eight L. acidophilus strains. To study the importance of SlpA in the adhesion of L. acidophilus, the amounts of SlpA proteins in LiCl extracts of the eight strains were compared by SDSpolyacrylamide gel electrophoresis. As a result, the adhesive abilities of L. acidophilus strains to Caco-2 cells correlated closely to the amount of SlpA in the cells and the productivity of IL-12, an inflammatory cytokine, in all eight strains. These results strongly suggested that SlpA in L. acidophilus might play a key role in its attachment to Caco-2 cells and in the release of IL-12 from dendritic cells.

Sickle red cell adhesion: many issues and some answers.

PubMed

Kaul, D K

2008-01-01

Among multiple pathologies associated with sickle cell disease, sickle red cell-endothelial interaction has been implicated as a potential initiating mechanism in vaso-occlusive events that characterize this disease. Vast literature exists on various aspects of sickle red cell adhesion, but many issues remain unresolved, especially pertaining to the role of sickle red cell heterogeneity, the relative role of multiple adhesion mechanisms and targets of antiadhesive therapy. This review briefly analyzes these issues.

Rac1 and Rac3 have opposing functions in cell adhesion and differentiation of neuronal cells.

PubMed

Hajdo-Milasinović, Amra; Ellenbroek, Saskia I J; van Es, Saskia; van der Vaart, Babet; Collard, John G

2007-02-15

Rac1 and Rac3 are highly homologous members of the Rho small GTPase family. Rac1 is ubiquitously expressed and regulates cell adhesion, migration and differentiation in various cell types. Rac3 is primarily expressed in brain and may therefore have a specific function in neuronal cells. We found that depletion of Rac1 by short interference RNA leads to decreased cell-matrix adhesions and cell rounding in neuronal N1E-115 cells. By contrast, depletion of Rac3 induces stronger cell adhesions and dramatically increases the outgrowth of neurite-like protrusions, suggesting opposite functions for Rac1 and Rac3 in neuronal cells. Consistent with this, overexpression of Rac1 induces cell spreading, whereas overexpression of Rac3 results in a contractile round morphology. Rac1 is mainly found at the plasma membrane, whereas Rac3 is predominantly localized in the perinuclear region. Residues 185-187, present in the variable polybasic rich region at the carboxyl terminus are responsible for the difference in phenotype induced by Rac1 and Rac3 as well as for their different intracellular localization. The Rac1-opposing function of Rac3 is not mediated by or dependent on components of the RhoA signaling pathway. It rather seems that Rac3 exerts its function through negatively affecting integrin-mediated cell-matrix adhesions. Together, our data reveal that Rac3 opposes Rac1 in the regulation of cell adhesion and differentiation of neuronal cells.

Overexpression of Selenoprotein SelK in BGC-823 Cells Inhibits Cell Adhesion and Migration.

PubMed

Ben, S B; Peng, B; Wang, G C; Li, C; Gu, H F; Jiang, H; Meng, X L; Lee, B J; Chen, C L

2015-10-01

Effects of human selenoprotein SelK on the adhesion and migration ability of human gastric cancer BGC-823 cells using Matrigel adhesion and transwell migration assays, respectively, were investigated in this study. The Matrigel adhesion ability of BGC-823 cells that overexpressed SelK declined extremely significantly (p < 0.01) compared with that of the cells not expressing the protein. The migration ability of BGC-823 cells that overexpressed SelK also declined extremely significantly (p < 0.01). On the other hand, the Matrigel adhesion ability and migration ability of the cells that overexpressed C-terminally truncated SelK did not decline significantly. The Matrigel adhesion ability and migration ability of human embryonic kidney HEK-293 cells that overexpressed SelK did not show significant change (p > 0.05) with the cells that overexpressed the C-terminally truncated protein. In addition to the effect on Matrigel adhesion and migration, the overexpression of SelK also caused a loss in cell viability (as measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide (MTT) colorimetric assay) and induced apoptosis as shown by confocal microscopy and flow cytometry. The cytosolic free Ca2+ level of these cells was significantly increased as detected by flow cytometry. But the overexpression of SelK in HEK-293 cells caused neither significant loss in cell viability nor apoptosis induction. Only the elevation of cytosolic free Ca2+ level in these cells was significant. Taken together, the results suggest that the overexpression of SelK can inhibit human cancer cell Matrigel adhesion and migration and cause both the loss in cell viability and induction of apoptosis. The release of intracellular Ca2+ from the endoplasmic reticulum might be a mechanism whereby the protein exerted its impact. Furthermore, only the full-length protein, but not C-terminally truncated form, was capable of producing such impact. The embryonic cells were not influenced by the elevation of free Ca2+ level in cytosol, probably due to their much greater tolerance to the variation.

Integrin β4 Signaling Promotes Mammary Tumor Cell Adhesion to Brain Microvascular Endothelium by Inducing ErbB2-mediated Secretion of VEGF

PubMed Central

Fan, Jie; Cai, Bin; Zeng, Min; Hao, Yanyan

2015-01-01

Prior studies have indicated that the β4 integrin promotes mammary tumor invasion and metastasis by combining with ErbB2 and amplifying its signaling capacity. However, the effector pathways and cellular functions by which the β4 integrin exerts these effects are incompletely understood. To examine if β4 signaling plays a role during mammary tumor cell adhesion to microvascular endothelium, we have examined ErbB2-transformed mammary tumor cells expressing either a wild-type (WT) or a signaling-defective form of β4 (1355T). We report that WT cells adhere to brain microvascular endothelium in vitro to a significantly larger extent as compared to 1355T cells. Interestingly, integrin β4 signaling does not exert a direct effect on adhesion to the endothelium or the underlying basement membrane. Rather, it enhances ErbB2-dependent expression of VEGF by tumor cells. VEGF in turn disrupts the tight and adherens junctions of endothelial monolayers, enabling the exposure of underlying basement membrane and increasing the adhesion of tumor cells to the intercellular junctions of endothelium. Inhibition of ErbB2 on tumor cells or the VEGFR-2 on endothelial cells suppresses mammary tumor cell adhesion to microvascular endothelium. Our results indicate that β4 signaling regulates VEGF expression by the mammary tumor cells thereby enhancing their adhesion to microvascular endothelium. PMID:21556948

Cell-substrate impedance fluctuations of single amoeboid cells encode cell-shape and adhesion dynamics

NASA Astrophysics Data System (ADS)

Leonhardt, Helmar; Gerhardt, Matthias; Höppner, Nadine; Krüger, Kirsten; Tarantola, Marco; Beta, Carsten

2016-01-01

We show systematic electrical impedance measurements of single motile cells on microelectrodes. Wild-type cells and mutant strains were studied that differ in their cell-substrate adhesion strength. We recorded the projected cell area by time-lapse microscopy and observed irregular oscillations of the cell shape. These oscillations were correlated with long-term variations in the impedance signal. Superposed to these long-term trends, we observed fluctuations in the impedance signal. Their magnitude clearly correlated with the adhesion strength, suggesting that strongly adherent cells display more dynamic cell-substrate interactions.

A viscoelastic-stochastic model of the effects of cytoskeleton remodelling on cell adhesion.

PubMed

Li, Long; Zhang, Wenyan; Wang, Jizeng

2016-10-01

Cells can adapt their mechanical properties through cytoskeleton remodelling in response to external stimuli when the cells adhere to the extracellular matrix (ECM). Many studies have investigated the effects of cell and ECM elasticity on cell adhesion. However, experiments determined that cells are viscoelastic and exhibiting stress relaxation, and the mechanism behind the effect of cellular viscoelasticity on the cell adhesion behaviour remains unclear. Therefore, we propose a theoretical model of a cluster of ligand-receptor bonds between two dissimilar viscoelastic media subjected to an applied tensile load. In this model, the distribution of interfacial traction is assumed to follow classical continuum viscoelastic equations, whereas the rupture and rebinding of individual molecular bonds are governed by stochastic equations. On the basis of this model, we determined that viscosity can significantly increase the lifetime, stability and dynamic strength of the adhesion cluster of molecular bonds, because deformation relaxation attributed to the viscoelastic property can increase the rebinding probability of each open bond and reduce the stress concentration in the adhesion area.

Epithelial cell morphology and adhesion on diamond films deposited and chemically modified by plasma processes.

PubMed

Rezek, Bohuslav; Ukraintsev, Egor; Krátká, Marie; Taylor, Andrew; Fendrych, Frantisek; Mandys, Vaclav

2014-09-01

The authors show that nanocrystalline diamond (NCD) thin films prepared by microwave plasma enhanced chemical vapor deposition apparatus with a linear antenna delivery system are well compatible with epithelial cells (5637 human bladder carcinoma) and significantly improve the cell adhesion compared to reference glass substrates. This is attributed to better adhesion of adsorbed layers to diamond as observed by atomic force microscopy (AFM) beneath the cells. Moreover, the cell morphology can be adjusted by appropriate surface treatment of diamond by using hydrogen and oxygen plasma. Cell bodies, cytoplasmic rims, and filopodia were characterized by Peakforce AFM. Oxidized NCD films perform better than other substrates under all conditions (96% of cells adhered well). A thin adsorbed layer formed from culture medium and supplemented with fetal bovine serum (FBS) covered the diamond surface and played an important role in the cell adhesion. Nevertheless, 50-100 nm large aggregates formed from the RPMI medium without FBS facilitated cell adhesion also on hydrophobic hydrogenated NCD (increase from 23% to 61%). The authors discuss applicability for biomedical uses.

Glutamate receptor activation in the kindled dentate gyrus.

PubMed

Behr, J; Heinemann, U; Mody, I

2000-01-01

The contribution of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), N-methyl-D-aspartate (NMDA), and kainate receptor activation to the enhanced seizure susceptibility of the dentate gyrus was investigated in an experimental model of temporal lobe epilepsy. Using the specific NMDA and AMPA receptor antagonists D-APV and SYM 2206, we examined alterations in glutamate receptor-dependent synaptic currents 48 hours and 28 days after kindling in field-potential and voltage-clamp recordings. Forty-eight hours after kindling, the fractions of AMPA and NMDA receptor-mediated excitatory postsynaptic current components shifted dramatically in favor of the NMDA receptor-mediated response. Four weeks after kindling, however, AMPA and NMDA receptor-mediated excitatory postsynaptic currents reverted to control-like values. Neither single nor repetitive perforant path stimuli evoked kainate receptor-mediated excitatory postsynaptic currents in dentate gyrus granule cells of control or kindled rats. The enhanced excitability of the kindled dentate gyrus 48 hours after the last seizure most likely results from transiently enhanced NMDA receptor activation. The NMDA receptor seems to play a critical role in the induction of the kindled state rather than in the persistence of the enhanced seizure susceptibility.

Direct projections from hypothalamic orexin neurons to brainstem cardiac vagal neurons.

PubMed

Dergacheva, Olga; Yamanaka, Akihiro; Schwartz, Alan R; Polotsky, Vsevolod Y; Mendelowitz, David

2016-12-17

Orexin neurons are known to augment the sympathetic control of cardiovascular function, however the role of orexin neurons in parasympathetic cardiac regulation remains unclear. To test the hypothesis that orexin neurons contribute to parasympathetic control we selectively expressed channelrhodopsin-2 (ChR2) in orexin neurons in orexin-Cre transgenic rats and examined postsynaptic currents in cardiac vagal neurons (CVNs) in the dorsal motor nucleus of the vagus (DMV). Simultaneous photostimulation and recording in ChR2-expressing orexin neurons in the lateral hypothalamus resulted in reliable action potential firing as well as large whole-cell currents suggesting a strong expression of ChR2 and reliable optogenetic excitation. Photostimulation of ChR2-expressing fibers in the DMV elicited short-latency (ranging from 3.2ms to 8.5ms) postsynaptic currents in 16 out of 44 CVNs tested. These responses were heterogeneous and included excitatory glutamatergic (63%) and inhibitory GABAergic (37%) postsynaptic currents. The results from this study suggest different sub-population of orexin neurons may exert diverse influences on brainstem CVNs and therefore may play distinct functional roles in parasympathetic control of the heart. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Inhibitory effects of clotrimazole on TNF-alpha-induced adhesion molecule expression and angiogenesis.

PubMed

Thapa, Dinesh; Lee, Jong Suk; Park, Min-A; Cho, Mi-Yeon; Park, Young-Joon; Choi, Han Gon; Jeong, Tae Cheon; Kim, Jung-Ae

2009-04-01

Cell adhesion molecules play a pivotal role in chronic inflammation and pathological angiogenesis. In the present study, we investigated the inhibitory effects of clotrimazole (CLT) on tumor necrosis factor (TNF)-alpha-induced changes in adhesion molecule expression. CLT dose-dependently inhibited monocyte chemoattractant protein-1 (MCP-1), intercellular cell adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) expressions in TNF-alpha-stimulated HT29 colonic epithelial cells. This inhibitory action of CLT correlated with a significant reduction in TNF-alpha-induced adhesion of monocytes to HT29 cells, which was comparable to the inhibitory effects of anti-ICAM-1 and VCAM-1 monoclonal antibodies on monocyte-epithelial adhesion. These inhibitory actions of CLT were, at least in part, attributable to the inhibition of redox sensitive NF-kappaB activation, as CLT inhibited TNF-alpha-induced ROS generation as well as NF-kappaB nuclear translocation and activation in HT29 cells. Furthermore, the inhibition of TNF-alpha-induced monocyte adhesion was also mimicked by the specific NF-kappaB inhibitor, pyrrolidine dithiocarbamate (PDTC). Inflammatory mediators including TNF-alpha have known to promote angiogenesis, which in turn further contributes to inflammatory pathology. Therefore, we additionally evaluated whether CLT modulates TNF-alpha-induced angiogenesis using in vivo chick chorioallantoic membrane (CAM) assay. The CAM assay showed that CLT dose-dependently attenuated TNF-alpha-induced angiogenesis, and the effect was correlated with decreased inflammation of the CAM tissue. In conclusion, our results suggest that CLT can inhibit TNF-alpha-triggered expression of adhesion molecules, ICAM-1 and VCAM-1, and angiogenesis during inflammation.

Cell-contact-dependent activation of CD4+ T cells by adhesion molecules on synovial fibroblasts.

PubMed

Mori, Masato; Hashimoto, Motomu; Matsuo, Takashi; Fujii, Takao; Furu, Moritoshi; Ito, Hiromu; Yoshitomi, Hiroyuki; Hirose, Jun; Ito, Yoshinaga; Akizuki, Shuji; Nakashima, Ran; Imura, Yoshitaka; Yukawa, Naoichiro; Yoshifuji, Hajime; Ohmura, Koichiro; Mimori, Tsuneyo

2017-05-01

To determine how cell-cell contact with synovial fibroblasts (SF) influence on the proliferation and cytokine production of CD4 +  T cells. Naïve CD4 +  T cells were cultured with SF from rheumatoid arthritis patients, stimulated by anti-CD3/28 antibody, and CD4 +  T cell proliferation and IFN-γ/IL-17 production were analyzed. To study the role of adhesion molecules, cell contact was blocked by transwell plate or anti-intracellular adhesion molecule-1 (ICAM-1)/vascular cell adhesion molecule-1(VCAM-1) antibody. To study the direct role of adhesion molecules for CD4 +  T cells, CD161 +  or CD161 - naïve CD4 +  T cells were stimulated on plastic plates coated by recombinant ICAM-1 or VCAM-1, and the source of IFN-γ/IL-17 were analyzed. SF enhanced naïve CD4 +  T cell proliferation and IFN-γ/IL-17 production in cell-contact and in part ICAM-1-/VCAM-1-dependent manner. Plate-coated ICAM-1 and VCAM-1 enhanced naïve CD4 +  T cell proliferation and IFN-γ production, while VCAM-1 efficiently promoting IL-17 production. CD161 +  naïve T cells upregulating LFA-1 and VLA-4 were the major source of IFN-γ/IL-17 upon interaction with ICAM-1/VCAM-1. CD4 +  T cells rapidly expand and secrete IFN-γ/IL-17 upon cell-contact with SF via adhesion molecules. Interfering with ICAM-1-/VCAM-1 may be beneficial for inhibiting RA synovitis.

Adhesion-Dependent Redistribution of MAP Kinase and MEK Promotes Muscarinic Receptor-Mediated Signaling to the Nucleus

PubMed Central

Slack, Barbara E.; Siniaia, Marina S.

2008-01-01

The mitogen-activated protein kinases (MAPKs) are activated by extracellular signals, and translocate to the nucleus where they modulate transcription. Integrin-mediated cell adhesion to extracellular matrix (ECM) proteins is required for efficient transmission of MAPK-based signals initiated by growth factors. However, the modulation of G protein-coupled receptor (GPCR) signaling by adhesion is less well understood. In the present study we assessed the impact of cell adhesion on MAPK activation by muscarinic M3 receptors. The muscarinic agonist carbachol more efficiently promoted stress fiber formation and tyrosine phosphorylation of focal adhesion-associated proteins in M3 receptor-expressing cells adherent to fibronectin or collagen type I, as compared to polylysine. Overall MAPK activation was robust in cells adherent to all three substrata. However, total levels of MAPK and mitogen-activated protein kinase kinase (MEK) in the nucleus were significantly greater in cells adherent to ECM proteins for 2.5 hours, and levels of activated MAPK and MEK in the nuclei of these cells were higher following carbachol stimulation, relative to levels in cells adherent to polylysine. MEK inhibitors did not prevent adhesion-dependent translocation of MAPK and MEK to the nucleus, and increased nuclear phospho-MEK levels in carbachol-stimulated cells. The results suggest that adhesion of cells to ECM triggers the redistribution of MAPK and MEK to the nucleus, possibly as a result of the cytoskeletal rearrangements that accompany cell spreading. This may represent a mechanism for priming the nucleus with MEK and MAPK, leading to more rapid and pronounced increases in intranuclear phospho-MAPK upon GPCR stimulation. PMID:15779001

CD15s/CD62E Interaction Mediates the Adhesion of Non-Small Cell Lung Cancer Cells on Brain Endothelial Cells: Implications for Cerebral Metastasis

PubMed Central

Jassam, Samah A.; Maherally, Zaynah; Ashkan, Keyoumars; Roncaroli, Federico; Fillmore, Helen L.; Pilkington, Geoffrey J.

2017-01-01

Expression of the cell adhesion molecule (CAM), Sialyl Lewis X (CD15s) correlates with cancer metastasis, while expression of E-selectin (CD62E) is stimulated by TNF-α. CD15s/CD62E interaction plays a key role in the homing process of circulating leukocytes. We investigated the heterophilic interaction of CD15s and CD62E in brain metastasis-related cancer cell adhesion. CD15s and CD62E were characterised in human brain endothelium (hCMEC/D3), primary non-small cell lung cancer (NSCLC) (COR-L105 and A549) and metastatic NSCLC (SEBTA-001 and NCI-H1299) using immunocytochemistry, Western blotting, flow cytometry and immunohistochemistry in human brain tissue sections. TNF-α (25 pg/mL) stimulated extracellular expression of CD62E while adhesion assays, under both static and physiological flow live-cell conditions, explored the effect of CD15s-mAb immunoblocking on adhesion of cancer cell–brain endothelium. CD15s was faintly expressed on hCMEC/D3, while high levels were observed on primary NSCLC cells with expression highest on metastatic NSCLC cells (p < 0.001). CD62E was highly expressed on hCMEC/D3 cells activated with TNF-α, with lower levels on primary and metastatic NSCLC cells. CD15s and CD62E were expressed on lung metastatic brain biopsies. CD15s/CD62E interaction was localised at adhesion sites of cancer cell–brain endothelium. CD15s immunoblocking significantly decreased cancer cell adhesion to brain endothelium under static and shear stress conditions (p < 0.001), highlighting the role of CD15s–CD62E interaction in brain metastasis. PMID:28698503

Dock mediates Scar- and WASp-dependent actin polymerization through interaction with cell adhesion molecules in founder cells and fusion-competent myoblasts.

PubMed

Kaipa, Balasankara Reddy; Shao, Huanjie; Schäfer, Gritt; Trinkewitz, Tatjana; Groth, Verena; Liu, Jianqi; Beck, Lothar; Bogdan, Sven; Abmayr, Susan M; Önel, Susanne-Filiz

2013-01-01

The formation of the larval body wall musculature of Drosophila depends on the asymmetric fusion of two myoblast types, founder cells (FCs) and fusion-competent myoblasts (FCMs). Recent studies have established an essential function of Arp2/3-based actin polymerization during myoblast fusion, formation of a dense actin focus at the site of fusion in FCMs, and a thin sheath of actin in FCs and/or growing muscles. The formation of these actin structures depends on recognition and adhesion of myoblasts that is mediated by cell surface receptors of the immunoglobulin superfamily. However, the connection of the cell surface receptors with Arp2/3-based actin polymerization is poorly understood. To date only the SH2-SH3 adaptor protein Crk has been suggested to link cell adhesion with Arp2/3-based actin polymerization in FCMs. Here, we propose that the SH2-SH3 adaptor protein Dock, like Crk, links cell adhesion with actin polymerization. We show that Dock is expressed in FCs and FCMs and colocalizes with the cell adhesion proteins Sns and Duf at cell-cell contact points. Biochemical data in this study indicate that different domains of Dock are involved in binding the cell adhesion molecules Duf, Rst, Sns and Hbs. We emphasize the importance of these interactions by quantifying the enhanced myoblast fusion defects in duf dock, sns dock and hbs dock double mutants. Additionally, we show that Dock interacts biochemically and genetically with Drosophila Scar, Vrp1 and WASp. Based on these data, we propose that Dock links cell adhesion in FCs and FCMs with either Scar- or Vrp1-WASp-dependent Arp2/3 activation.

Ethanol does not inhibit the adhesive activity of Drosophila neuroglian or human L1 in Drosophila S2 tissue culture cells.

PubMed

Vallejo, Y; Hortsch, M; Dubreuil, R R

1997-05-02

Members of the L1 family of homophilic neural cell adhesion molecules are thought to play an important role in nervous system development and function. It is also suggested that L1 is a direct target of ethanol in fetal alcohol syndrome, since ethanol inhibits the aggregation of cultured cells expressing L1 (Ramanathan, R., Wilkemeyer, M. F., Mittel, B., Perides, G., and Charness, M. E. (1996) J. Cell Biol. 133, 381-390). If ethanol acts directly on the homophilic adhesive function of the L1 molecule, then inhibition of aggregation by ethanol should be observed in any cell type that expresses L1. Here we examined the effect of physiologically relevant concentrations of ethanol on the aggregation of Drosophila S2 cells that expressed either neuroglian (the Drosophila homolog of L1) or human L1. The aggregation of these S2 cells is known to be solely dependent on the homophilic interactions between L1 or neuroglian molecules. Neither cell adhesion molecule was affected when cell aggregation assays were carried out in the presence of >/=38 mM ethanol. The recruitment of membrane skeleton assembly at sites of cell-cell contact (a transmembrane signaling function of human L1) was also unaffected by the presence of ethanol. Thus the previously described inhibition of cell adhesion by ethanol in L1-expressing cells cannot be explained by a simple direct effect on the adhesive activity of L1 family members.

Dynamic pattern of endothelial cell adhesion molecule expression in muscle and perineural vessels from patients with classic polyarteritis nodosa.

PubMed

Coll-Vinent, B; Cebrián, M; Cid, M C; Font, C; Esparza, J; Juan, M; Yagüe, J; Urbano-Márquez, A; Grau, J M

1998-03-01

To investigate endothelial cell adhesion molecule expression in vessels from patients with classic polyarteritis nodosa (PAN). Frozen sections of 21 muscle and 16 nerve samples from 30 patients with biopsy-proven PAN and 12 histologically normal muscle and 2 histologically normal nerve samples from 12 controls were studied immunohistochemically, using specific monoclonal antibodies (MAb) that recognize adhesion molecules. Adhesion molecules identified were intercellular adhesion molecule 1 (ICAM-1), ICAM-2, ICAM-3, vascular cell adhesion molecule 1 (VCAM-1), platelet endothelial cell adhesion molecule 1 (PECAM-1), E-selectin, P-selectin, L-selectin, lymphocyte function-associated antigen 1 (LFA-1), and very late activation antigen 4 (VLA-4). Neutrophils were identified with a MAb recognizing neutrophil elastase. Endothelial cells were identified with the lectin ulex europaeus. In early lesions, expression of PECAM-1, ICAM-1, ICAM-2, and P-selectin was similar to that in control samples, and VCAM-1 and E-selectin were induced in vascular endothelium. In advanced lesions, immunostaining for adhesion molecules diminished or disappeared in luminal endothelium, whereas these molecules were clearly expressed in microvessels within and surrounding inflamed vessels. Staining in endothelia from vessels in a healing stage tended to be negative. A high proportion of infiltrating leukocytes expressed LFA-1 and VLA-4, and only a minority expressed L-selectin. No relationship between the expression pattern of adhesion molecules and clinical features, disease duration, or previous corticosteroid treatment was observed. Endothelial adhesion molecule expression in PAN is a dynamic process that varies according to the histopathologic stage of the vascular lesions. The preferential expression of constitutive and inducible adhesion molecules in microvessels suggests that angiogenesis contributes to the persistence of inflammatory infiltration in PAN.

αV-class integrins exert dual roles on α5β1 integrins to strengthen adhesion to fibronectin

PubMed Central

Bharadwaj, Mitasha; Strohmeyer, Nico; Colo, Georgina P.; Helenius, Jonne; Beerenwinkel, Niko; Schiller, Herbert B.; Fässler, Reinhard; Müller, Daniel J.

2017-01-01

Upon binding to the extracellular matrix protein, fibronectin, αV-class and α5β1 integrins trigger the recruitment of large protein assemblies and strengthen cell adhesion. Both integrin classes have been functionally specified, however their specific roles in immediate phases of cell attachment remain uncharacterized. Here, we quantify the adhesion of αV-class and/or α5β1 integrins expressing fibroblasts initiating attachment to fibronectin (≤120 s) by single-cell force spectroscopy. Our data reveals that αV-class integrins outcompete α5β1 integrins. Once engaged, αV-class integrins signal to α5β1 integrins to establish additional adhesion sites to fibronectin, away from those formed by αV-class integrins. This crosstalk, which strengthens cell adhesion, induces α5β1 integrin clustering by RhoA/ROCK/myosin-II and Arp2/3-mediated signalling, whereas overall cell adhesion depends on formins. The dual role of both fibronectin-binding integrin classes commencing with an initial competition followed by a cooperative crosstalk appears to be a basic cellular mechanism in assembling focal adhesions to the extracellular matrix. PMID:28128308

Fluorescence Fluctuation Approaches to the Study of Adhesion and Signaling

PubMed Central

Bachir, Alexia I.; Kubow, Kristopher E.; Horwitz, Alan R.

2013-01-01

Cell–matrix adhesions are large, multimolecular complexes through which cells sense and respond to their environment. They also mediate migration by serving as traction points and signaling centers and allow the cell to modify the surroucnding tissue. Due to their fundamental role in cell behavior, adhesions are germane to nearly all major human health pathologies. However, adhesions are extremely complex and dynamic structures that include over 100 known interacting proteins and operate over multiple space (nm–µm) and time (ms–min) regimes. Fluorescence fluctuation techniques are well suited for studying adhesions. These methods are sensitive over a large spatiotemporal range and provide a wealth of information including molecular transport dynamics, interactions, and stoichiometry from a single time series. Earlier chapters in this volume have provided the theoretical background, instrumentation, and analysis algorithms for these techniques. In this chapter, we discuss their implementation in living cells to study adhesions in migrating cells. Although each technique and application has its own unique instrumentation and analysis requirements, we provide general guidelines for sample preparation, selection of imaging instrumentation, and optimization of data acquisition and analysis parameters. Finally, we review several recent studies that implement these techniques in the study of adhesions. PMID:23280111

Radil controls neutrophil adhesion and motility through β2-integrin activation

PubMed Central

Liu, Lunhua; Aerbajinai, Wulin; Ahmed, Syed M.; Rodgers, Griffin P.; Angers, Stephane; Parent, Carole A.

2012-01-01

Integrin activation is required to facilitate multiple adhesion-dependent functions of neutrophils, such as chemotaxis, which is critical for inflammatory responses to injury and pathogens. However, little is known about the mechanisms that mediate integrin activation in neutrophils. We show that Radil, a novel Rap1 effector, regulates β1- and β2-integrin activation and controls neutrophil chemotaxis. On activation and chemotactic migration of neutrophils, Radil quickly translocates from the cytoplasm to the plasma membrane in a Rap1a-GTP–dependent manner. Cells overexpressing Radil show a substantial increase in cell adhesion, as well as in integrin/focal adhesion kinase (FAK) activation, and exhibit an elongated morphology, with severe tail retraction defects. This phenotype is effectively rescued by treatment with either β2-integrin inhibitory antibodies or FAK inhibitors. Conversely, knockdown of Radil causes severe inhibition of cell adhesion, β2-integrin activation, and chemotaxis. Furthermore, we found that inhibition of Rap activity by RapGAP coexpression inhibits Radil-mediated integrin and FAK activation, decreases cell adhesion, and abrogates the long-tail phenotype of Radil cells. Overall, these studies establish that Radil regulates neutrophil adhesion and motility by linking Rap1 to β2-integrin activation. PMID:23097489

Radil controls neutrophil adhesion and motility through β2-integrin activation.

PubMed

Liu, Lunhua; Aerbajinai, Wulin; Ahmed, Syed M; Rodgers, Griffin P; Angers, Stephane; Parent, Carole A

2012-12-01

Integrin activation is required to facilitate multiple adhesion-dependent functions of neutrophils, such as chemotaxis, which is critical for inflammatory responses to injury and pathogens. However, little is known about the mechanisms that mediate integrin activation in neutrophils. We show that Radil, a novel Rap1 effector, regulates β1- and β2-integrin activation and controls neutrophil chemotaxis. On activation and chemotactic migration of neutrophils, Radil quickly translocates from the cytoplasm to the plasma membrane in a Rap1a-GTP-dependent manner. Cells overexpressing Radil show a substantial increase in cell adhesion, as well as in integrin/focal adhesion kinase (FAK) activation, and exhibit an elongated morphology, with severe tail retraction defects. This phenotype is effectively rescued by treatment with either β2-integrin inhibitory antibodies or FAK inhibitors. Conversely, knockdown of Radil causes severe inhibition of cell adhesion, β2-integrin activation, and chemotaxis. Furthermore, we found that inhibition of Rap activity by RapGAP coexpression inhibits Radil-mediated integrin and FAK activation, decreases cell adhesion, and abrogates the long-tail phenotype of Radil cells. Overall, these studies establish that Radil regulates neutrophil adhesion and motility by linking Rap1 to β2-integrin activation.

Anti-adhesive effects of a newly developed two-layered gelatin sheet in dogs.

PubMed

Torii, Hiroko; Takagi, Toshitaka; Urabe, Mamoru; Tsujimoto, Hiroyuki; Ozamoto, Yuki; Miyamoto, Hiroe; Ikada, Yoshihito; Hagiwara, Akeo

2017-08-01

Adhesion after pelvic surgery causes infertility, ectopic pregnancy, and ileus or abdominal pain. The materials currently available for clinical use are insufficient. The purpose of this study was to develop an anti-adhesive material that overcomes the limitations of conventional anti-adhesive agents. The adhesion prevention effects of three methods - a two-layered sheet composed of gelatin film and gelatin sponge, Seprafilm and INTERCEED - were evaluated in 37 dogs. Anti-adhesive effects were investigated macroscopically and microscopically in a cauterized uterus adhesion model. Cell growth on the materials in vitro using human peritoneal mesothelial cells, fibroblasts and uterine smooth muscle cells were also evaluated. The two-layered gelatin sheet had significantly superior anti-adhesive effects compared to the conventional materials (Seprafilm and INTERCEED). A single-cell layer of mature mesothelium formed three weeks after surgery in the gelatin group. Peritoneum regeneration in the Seprafilm and INTERCEED groups was delayed and incomplete in the early phase. Little inflammation around the materials occurred and cell growth was significantly proliferated with the gelatin sheet. The anti-adhesive effects of a two-layered gelatin sheet were superior to conventional agents in a cauterized canine uterus model, demonstrating early regeneration of the peritoneum, little inflammation and material endurance. The newly developed two-layered gelatin sheet is a useful option as an anti-adhesive agent for deeply injured and hemorrhagic sites. © 2017 The Authors. Journal of Obstetrics and Gynaecology Research published by John Wiley & Sons Australia, Ltd on behalf of Japan Society of Obstetrics and Gynecology.

[Effect of LPXN Overexpression on the Proliferation, Adhesion and Invasion of THP-1 Cells and Its Mechamisms].

PubMed

Dai, Hai-Ping; Zhu, Guo-Hua; Wu, Li-Li; Wang, Qian; Yao, Hong; Wang, Qin-Rong; Wen, Li-Jun; Qiu, Hui-Ying; Shen, Qun; Chen, Su-Ning; Wu, De-Pei

2017-06-01

To explore the effect of LPXN overexpression on the proliferation, adhesion and invasion of THP-1 cells and its possible mechanism. A THP-1 cell line with stable overexpression of LPXN was constucted by using a lentivirus method, CCK-8 was used to detect the proliferation of cells, adhesion test was used to evaluate adhesion ablity of cells to Fn. Transwell assay was used to detect the change of invasion capability. Western blot was used to detect expression of LPXN, ERK, pERK and integrin α4, α5, β1, the Gelatin zymography was applied to detect activity of MMP2/MMP9 secreted by the THP-1 cells. Successful establishment of THP-1 cells with LPXN overexpression (THP-1 LPXN) was confirmed with Western blot. THP-1 LPXN cells were shown to proliferate faster than the control THP-1 vector cells. Adhesion to Fn and expression of ERK, integrin α4, α5 and β1 in the THP-1 LPXN cells were higher than that in the control cells. Invasion across matrigel and enhanced activity of MMP2 could be detected both in the THP-1 LPXN cells as compared with the control cells. Ectopically ovexpression of LPXN may promote proliferation of THP-1 cells through up-regulation of ERK; promote adhesion of THP-1 cells through up-regulating the integrin α4/β1 as well as integrin α5/β1 complex; promote invasion of THP-1 cells through activating MMP2.

Measurement of cell adhesion force by vertical forcible detachment using an arrowhead nanoneedle and atomic force microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ryu, Seunghwan; Hashizume, Yui; Mishima, Mari

Graphical abstract: - Highlights: • We developed a method to measure cell adhesion force by detaching cell using an arrowhead nanoneedle and AFM. • A nanofilm consisting of fibronectin and gelatin was formed on cell surface to reinforce the cell cortex. • By the nanofilm lamination, detachment efficiencies of strongly adherent cell lines were improved markedly. - Abstract: The properties of substrates and extracellular matrices (ECM) are important factors governing the functions and fates of mammalian adherent cells. For example, substrate stiffness often affects cell differentiation. At focal adhesions, clustered–integrin bindings link cells mechanically to the ECM. In order tomore » quantitate the affinity between cell and substrate, the cell adhesion force must be measured for single cells. In this study, forcible detachment of a single cell in the vertical direction using AFM was carried out, allowing breakage of the integrin–substrate bindings. An AFM tip was fabricated into an arrowhead shape to detach the cell from the substrate. Peak force observed in the recorded force curve during probe retraction was defined as the adhesion force, and was analyzed for various types of cells. Some of the cell types adhered so strongly that they could not be picked up because of plasma membrane breakage by the arrowhead probe. To address this problem, a technique to reinforce the cellular membrane with layer-by-layer nanofilms composed of fibronectin and gelatin helped to improve insertion efficiency and to prevent cell membrane rupture during the detachment process, allowing successful detachment of the cells. This method for detaching cells, involving cellular membrane reinforcement, may be beneficial for evaluating true cell adhesion forces in various cell types.« less

Phosphorylation of tyrosine 285 of PAK1 facilitates βPIX/GIT1 binding and adhesion turnover

PubMed Central

Hammer, Alan; Oladimeji, Peter; De Las Casas, Luis E.; Diakonova, Maria

2015-01-01

The p21-activated serine-threonine kinase (PAK1) regulates cell motility and adhesion. We have previously shown that the prolactin (PRL)-activated tyrosine kinase JAK2 phosphorylates PAK1 in vivo and in vitro and identified tyrosines 153, 201, and 285 in PAK1 as sites of JAK2 tyrosyl phosphorylation. Here, we further investigate the role of the tyrosyl phosphorylated PAK1 (pTyr-PAK1) in regulation of cell adhesion. We use human breast cancer T47D cell lines that stably overexpress PAK1 wild type or PAK1 Y3F mutant in which these 3 JAK2 phosphorylation sites were mutated to phenylalanine. We demonstrate that PRL/JAK2-dependent phosphorylation of these tyrosines promotes a motile phenotype in the cells upon adhesion, participates in regulation of cell adhesion on collagen IV, and is required for maximal PAK1 kinase activity. Down-regulation of PAK1 abolishes the effect of PAK1 on cell adhesion. We show that the tyrosyl phosphorylation of PAK1 promotes PAK1 binding to β-PAK1-interacting guanine-nucleotide exchange factor (βPIX) and G protein-coupled receptor kinase-interacting target 1 (GIT1), phosphorylation of paxillin on Ser273, and formation and distribution of adhesion complexes. Using phosphospecific antibodies (Abs) directed to single phosphorylated tyrosines on PAK1, we identified Tyr285 as a site of PRL-dependent phosphorylation of PAK1 by JAK2. Furthermore, using PAK1 Y285F mutant, we provide evidence for a role of pTyr285 in cell adhesion, enhanced βPIX/GIT1 binding, and adhesion turnover. Our immunohistochemistry analysis demonstrates that pTyr285- PAK1 may modulate PAK1 signaling during tumor progression.—Hammer, A., Oladimeji, P., De La Casas, L. E., Diakonova, M. Phosphorylation of tyrosine 285 of PAK1 facilitates bPIX/GIT1 binding and adhesion turnover. PMID:25466889

Expression of Inflammation-related Intercellular Adhesion Molecules in Cardiomyocytes In Vitro and Modulation by Pro-inflammatory Agents.

PubMed

El-Battrawy, Ibrahim; Tülümen, Erol; Lang, Siegfried; Akin, Ibrahim; Behnes, Michael; Zhou, Xiabo; Mavany, Martin; Bugert, Peter; Bieback, Karen; Borggrefe, Martin; Elmas, Elif

2016-01-01

Cell-surface adhesion molecules regulate multiple intercellular and intracellular processes and play important roles in inflammation by facilitating leukocyte endothelial transmigration. Whether cardiomyocytes express surface-adhesion molecules related to inflammation and the effect of pro-inflammatory mediators remain unknown. In the present study, the expression of different cell-adhesion molecules (CD11a, CD11b, CD31, CD62P, CD162, F11 receptor and mucosal vascular addressin cell adhesion molecule 1 (MADCAM1)) and the effect of pro-inflammatory mediators were investigated in an in vitro model of human cardiomyocytes. Cells were supplied as a primary culture of cardiac alpha actin-positive cells from human heart tissue. The cells were incubated for 24 h with 1 U/ml thrombin or 700 ng/ml lipopolysaccharide (LPS) or with a combination of both. The expression of the cell adhesion molecules was measured by flow cytometry. In cultured human cardiomyocytes, 22.8% of cells expressed CD31, 7.1% MADCAM1 and 2.6% F11R. CD11a, CD11b, CD62P and CD162 were expressed by fewer than 2% of the cells at baseline. CD31 expression increased on incubation of cardiomyocytes with thrombin by 26% (p<0.05) and with LPS by 26% (p=0.06). The combination of thrombin and LPS did not result in increased levels of CD31 (p>0.10). The pro-inflammatory agents LPS and thrombin had no effect on the expression of MADCAM1 and F11R. Inflammation-related cell-adhesion molecules CD31, MADCAM1 and F11R were shown to be expressed on the surface of human cardiomyocytes in an in vitro model. Incubation with LPS or thrombin resulted in increased expression of CD31, however, it did not modify the expression of the cell adhesion molecules MADCAM1 and F11R. Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Single cell adhesion force measurement for cell viability identification using an AFM cantilever-based micro putter

NASA Astrophysics Data System (ADS)

Shen, Yajing; Nakajima, Masahiro; Kojima, Seiji; Homma, Michio; Kojima, Masaru; Fukuda, Toshio

2011-11-01

Fast and sensitive cell viability identification is a key point for single cell analysis. To address this issue, this paper reports a novel single cell viability identification method based on the measurement of single cell shear adhesion force using an atomic force microscopy (AFM) cantilever-based micro putter. Viable and nonviable yeast cells are prepared and put onto three kinds of substrate surfaces, i.e. tungsten probe, gold and ITO substrate surfaces. A micro putter is fabricated from the AFM cantilever by focused ion beam etching technique. The spring constant of the micro putter is calibrated using the nanomanipulation approach. The shear adhesion force between the single viable or nonviable cell and each substrate is measured using the micro putter based on the nanorobotic manipulation system inside an environmental scanning electron microscope. The adhesion force is calculated based on the deflection of the micro putter beam. The results show that the adhesion force of the viable cell to the substrate is much larger than that of the nonviable cell. This identification method is label free, fast, sensitive and can give quantitative results at the single cell level.

Direct Force Measurements of Receptor-Ligand Interactions on Living Cells

NASA Astrophysics Data System (ADS)

Eibl, Robert H.

The characterization of cell adhesion between two living cells at the level of single receptor-ligand bonds is an experimental challenge. This chapter describes how the extremely sensitive method of atomic force microscopy (AFM) based force spectroscopy can be applied to living cells in order to probe for cell-to-cell or cell-to-substrate interactions mediated by single pairs of adhesion receptors. In addition, it is outlined how single-molecule AFM force spectroscopy can be used to detect physiologic changes of an adhesion receptor in a living cell. This force spectroscopy allows us to detect in living cells rapidly changing, chemokine SDF-1 triggered activation states of single VLA-4 receptors. This recently developed AFM application will allow for the detailed investigation of the integrin-chemokine crosstalk of integrin activation mechanisms and on how other adhesion receptors are modulated in health and disease. As adhesion molecules, living cells and even bacteria can be studied by single-molecule AFM force spectroscopy, this method is set to become a powerful tool that can not only be used in biophysics, but in cell biology as well as in immunology and cancer research.

TAp73 is essential for germ cell adhesion and maturation in testis

PubMed Central

Holembowski, Lena; Kramer, Daniela; Riedel, Dietmar; Sordella, Raffaella; Nemajerova, Alice; Dobbelstein, Matthias

2014-01-01

A core evolutionary function of the p53 family is to protect the genomic integrity of gametes. However, the role of p73 in the male germ line is unknown. Here, we reveal that TAp73 unexpectedly functions as an adhesion and maturation factor of the seminiferous epithelium orchestrating spermiogenesis. TAp73 knockout (TAp73KO) and p73KO mice, but not ΔNp73KO mice, display a “near-empty seminiferous tubule” phenotype due to massive premature loss of immature germ cells. The cellular basis of this phenotype is defective cell–cell adhesions of developing germ cells to Sertoli nurse cells, with likely secondary degeneration of Sertoli cells, including the blood–testis barrier, which leads to disruption of the adhesive integrity and maturation of the germ epithelium. At the molecular level, TAp73, which is produced in germ cells, controls a coordinated transcriptional program of adhesion- and migration-related proteins including peptidase inhibitors, proteases, receptors, and integrins required for germ–Sertoli cell adhesion and dynamic junctional restructuring. Thus, we propose the testis as a unique organ with strict division of labor among all family members: p63 and p53 safeguard germ line fidelity, whereas TAp73 ensures fertility by enabling sperm maturation. PMID:24662569

Monitoring in real-time focal adhesion protein dynamics in response to a discrete mechanical stimulus

NASA Astrophysics Data System (ADS)

von Bilderling, Catalina; Caldarola, Martín; Masip, Martín E.; Bragas, Andrea V.; Pietrasanta, Lía I.

2017-01-01

The adhesion of cells to the extracellular matrix is a hierarchical, force-dependent, multistage process that evolves at several temporal scales. An understanding of this complex process requires a precise measurement of forces and its correlation with protein responses in living cells. We present a method to quantitatively assess live cell responses to a local and specific mechanical stimulus. Our approach combines atomic force microscopy with fluorescence imaging. Using this approach, we evaluated the recruitment of adhesion proteins such as vinculin, focal adhesion kinase, paxillin, and zyxin triggered by applying forces in the nN regime to live cells. We observed in real time the development of nascent adhesion sites, evident from the accumulation of early adhesion proteins at the position where the force was applied. We show that the method can be used to quantify the recruitment characteristic times for adhesion proteins in the formation of focal complexes. We also found a spatial remodeling of the mature focal adhesion protein zyxin as a function of the applied force. Our approach allows the study of a variety of complex biological processes involved in cellular mechanotransduction.

Monitoring in real-time focal adhesion protein dynamics in response to a discrete mechanical stimulus.

PubMed

von Bilderling, Catalina; Caldarola, Martín; Masip, Martín E; Bragas, Andrea V; Pietrasanta, Lía I

2017-01-01

The adhesion of cells to the extracellular matrix is a hierarchical, force-dependent, multistage process that evolves at several temporal scales. An understanding of this complex process requires a precise measurement of forces and its correlation with protein responses in living cells. We present a method to quantitatively assess live cell responses to a local and specific mechanical stimulus. Our approach combines atomic force microscopy with fluorescence imaging. Using this approach, we evaluated the recruitment of adhesion proteins such as vinculin, focal adhesion kinase, paxillin, and zyxin triggered by applying forces in the nN regime to live cells. We observed in real time the development of nascent adhesion sites, evident from the accumulation of early adhesion proteins at the position where the force was applied. We show that the method can be used to quantify the recruitment characteristic times for adhesion proteins in the formation of focal complexes. We also found a spatial remodeling of the mature focal adhesion protein zyxin as a function of the applied force. Our approach allows the study of a variety of complex biological processes involved in cellular mechanotransduction.

Limitation of Cell Adhesion by the Elasticity of the Extracellular Matrix

PubMed Central

Nicolas, Alice; Safran, Samuel. A.

2006-01-01

Cell/matrix adhesions are modulated by cytoskeletal or external stresses and adapt to the mechanical properties of the extracellular matrix. We propose that this mechanosensitivity arises from the activation of a mechanosensor located within the adhesion itself. We show that this mechanism accounts for the observed directional growth of focal adhesions and the reduction or even cessation of their growth when cells adhere to a soft extracellular matrix. We predict quantitatively that both the elasticity and the thickness of the matrix play a key role in the dynamics of focal adhesions. Two different types of dynamics are expected depending on whether the thickness of the matrix is of order of or much larger than the adhesion size. In the latter situation, we predict that the adhesion region reaches a saturation size that can be tuned by the mechanical properties of the matrix. PMID:16581840

Heparan sulfate proteoglycans regulate autophagy in Drosophila.

PubMed

Reynolds-Peterson, Claire E; Zhao, Na; Xu, Jie; Serman, Taryn M; Xu, Jielin; Selleck, Scott B

2017-08-03

Heparan sulfate-modified proteoglycans (HSPGs) are important regulators of signaling and molecular recognition at the cell surface and in the extracellular space. Disruption of HSPG core proteins, HS-synthesis, or HS-degradation can have profound effects on growth, patterning, and cell survival. The Drosophila neuromuscular junction provides a tractable model for understanding the activities of HSPGs at a synapse that displays developmental and activity-dependent plasticity. Muscle cell-specific knockdown of HS biosynthesis disrupted the organization of a specialized postsynaptic membrane, the subsynaptic reticulum (SSR), and affected the number and morphology of mitochondria. We provide evidence that these changes result from a dysregulation of macroautophagy (hereafter referred to as autophagy). Cellular and molecular markers of autophagy are all consistent with an increase in the levels of autophagy in the absence of normal HS-chain biosynthesis and modification. HS production is also required for normal levels of autophagy in the fat body, the central energy storage and nutritional sensing organ in Drosophila. Genetic mosaic analysis indicates that HS-dependent regulation of autophagy occurs non-cell autonomously, consistent with HSPGs influencing this cellular process via signaling in the extracellular space. These findings demonstrate that HS biosynthesis has important regulatory effects on autophagy and that autophagy is critical for normal assembly of postsynaptic membrane specializations.

Glucosamine Treatment-mediated O-GlcNAc Modification of Paxillin Depends on Adhesion State of Rat Insulinoma INS-1 Cells*

PubMed Central

Kwak, Tae Kyoung; Kim, Hyeonjung; Jung, Oisun; Lee, Sin-Ae; Kang, Minkyung; Kim, Hyun Jeong; Park, Ji-Min; Kim, Sung-Hoon; Lee, Jung Weon

2010-01-01

Protein-protein interactions and/or signaling activities at focal adhesions, where integrin-mediated adhesion to extracellular matrix occurs, are critical for the regulation of adhesion-dependent cellular functions. Although the phosphorylation and activities of focal adhesion molecules have been intensively studied, the effects of the O-GlcNAc modification of their Ser/Thr residues on cellular functions have been largely unexplored. We investigated the effects of O-GlcNAc modification on actin reorganization and morphology of rat insulinoma INS-1 cells after glucosamine (GlcN) treatment. We found that paxillin, a key adaptor molecule in focal adhesions, could be modified by O-GlcNAc in INS-1 cells treated with GlcN and in pancreatic islets from mice treated with streptozotocin. Ser-84/85 in human paxillin appeared to be modified by O-GlcNAc, which was inversely correlated to Ser-85 phosphorylation (Ser-83 in rat paxillin). Integrin-mediated adhesion signaling inhibited the GlcN treatment-enhanced O-GlcNAc modification of paxillin. Adherent INS-1 cells treated with GlcN showed restricted protrusions, whereas untreated cells showed active protrusions for multiple-elongated morphologies. Upon GlcN treatment, expression of a triple mutation (S83A/S84A/S85A) resulted in no further restriction of protrusions. Together these observations suggest that murine pancreatic β cells may have restricted actin organization upon GlcN treatment by virtue of the O-GlcNAc modification of paxillin, which can be antagonized by a persistent cell adhesion process. PMID:20829364

The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells.

PubMed

Leckel, K; Beecken, W-D; Jonas, D; Oppermann, E; Coman, M C; Beck, K-F; Cinatl, J; Hailer, N P; Auth, M K H; Bechstein, W O; Shipkova, M; Blaheta, R A

2003-11-01

Immunosuppression correlates with the development and recurrence of cancer. Mycophenolate mofetil (MMF) has been shown to reduce adhesion molecule expression and leucocyte recruitment into the donor organ. We have hypothesized that MMF might also prevent receptor-dependent tumour dissemination. Therefore, we have investigated the effects of MMF on tumour cell adhesion to human umbilical vein endothelial cells (HUVEC) and compared them with the effects on T cell-endothelial cell interactions. Influence of MMF on cellular adhesion to HUVEC was analysed using isolated CD4+ and CD8+ T cells, or WiDr colon adenocarcinoma cells as the model tumour. HUVEC receptors ICAM-1, VCAM-1, E-selectin and P-selectin were detected by flow cytometry, Western blot or Northern blot analysis. Binding activity of T cells or WiDr cells in the presence of MMF were measured using immobilized receptor globulin chimeras. MMF potently blocked both T cell and WiDr cell binding to endothelium by 80%. Surface expression of the endothelial cell receptors was reduced by MMF in a dose-dependent manner. E-selectin mRNA was concurrently reduced with a maximum effect at 1 microm. Interestingly, MMF acted differently on T cells and WiDr cells. Maximum efficacy of MMF was reached at 10 and 1 microm, respectively. Furthermore, MMF specifically suppressed T cell attachment to ICAM-1, VCAM-1 and P-selectin. In contrast, MMF prevented WiDr cell attachment to E-selectin. In conclusion, our data reveal distinct effects of MMF on both T cell adhesion and tumour cell adhesion to endothelial cells. This suggests that MMF not only interferes with the invasion of alloactivated T cells, but might also be of value in managing post-transplantation malignancy.

μ-Opioid Receptors Selectively Regulate Basal Inhibitory Transmission in the Central Amygdala: Lack of Ethanol Interactions

PubMed Central

Kang-Park, Maeng-Hee; Kieffer, Brigitte L.; Roberts, Amanda J.; Roberto, Marisa; Madamba, Samuel G.; Siggins, George Robert; Moore, Scott D.

2009-01-01

Endogenous opioid systems are implicated in the actions of ethanol. For example, μ-opioid receptor (MOR) knockout (KO) mice self-administer less alcohol than the genetically intact counterpart wild-type (WT) mice (Roberts et al., 2000). MOR KO mice also exhibit less anxiety-like behavior than WT mice (Filliol et al., 2000). To investigate the neurobiological mechanisms underlying these behaviors, we examined the effect of ethanol in brain slices from MOR KO and WT mice using sharp-electrode and whole-cell patch recording techniques. We focused our study in the central nucleus of the amygdala (CeA) because it is implicated in alcohol drinking behavior and stress behavior. We found that the amplitudes of evoked inhibitory postsynaptic currents (IPSCs) or inhibitory postsynaptic potentials (IPSPs) were significantly greater in MOR KO mice than WT mice. In addition, the baseline frequencies of spontaneous and miniature GABAA receptor-mediated inhibitory postsynaptic currents were significantly greater in CeA neurons from MOR KO than WT mice. However, ethanol enhancements of evoked IPSP and IPSC amplitudes and the frequency of miniature IPSCs were comparable between WT and MOR KO mice. Baseline spontaneous and miniature excitatory postsynaptic currents (EPSCs) and ethanol effects on EPSCs were not significantly different between MOR KO and WT mice. Based on knowledge of CeA circuitry and projections, we hypothesize that the role of MOR- and GABA receptor-mediated mechanisms in CeA underlying reinforcing effects of ethanol operate independently, possibly through pathway-specific responses within CeA. PMID:18854491

In vitro effects of ATG-Fresenius on immune cell adhesion.

PubMed

Kanzler, I; Seitz-Merwald, I; Schleger, S; Kaczmarek, I; Kur, F; Beiras-Fernandez, A

2013-06-01

ATG-Fresenius, a purified rabbit polyclonal anti-human T-lymphocyte immunoglobulin is used for induction immunosuppression as well as prevention and treatment of acute rejection episodes among patients receiving solid organ transplants. The aim of this study was to investigate the in vitro activity of ATG-Fresenius upon immune cell adhesion, which may explain its activity to mitigate ischemia-reperfusion injury. Human vascular endothelial cells (HUVEC) and peripheral blood mononuclear cells (PBMCs) isolated from umbilical vein or peripheral blood were incubated 20 to 24 hours before analysis. HUVEC were incubated with 10 and 100 μg/mL ATG-Fresenius or reference polyclonal rabbit immunoglobulin G. Analysis of immune cell adhesion to endothelial cells was studied in cocultures of PBMCs and adherent HUVEC. Endothelial cell expression of adhesion molecules CD62E and CD54 was determined by flow cytometry. The numbers of T-, B- and natural killer cells attached to HUVEC were also determined by flow cytometry. Groups were compared using one-way analysis of variance. We showed that ATG-Fresenius binds to endothelial cells particularly activated ones expressing increased levels of E-selectin and ICAM-1. The increased binding of ATG-Fresenius to activated endothelial cells was consistent with its known binding to Intercellular Adhesion Molecule 1 (ICAM-1) and selectins. We also showed that ATG-Fresenius inhibited adhesion of prestimulated immune cells to activated endothelium. We demonstrated dose-dependent binding of ATG-Fresenius to activated endothelial cells. Copyright © 2013 Elsevier Inc. All rights reserved.

[Effects of selenium compounds on proliferation, migration and adhesion of HeLa cells].

PubMed

Sun, Licui; Lu, Jiaxi; Wang, Qin; Liu, Yiqun; Han, Feng; Yang, Yanhua; Zhang, Hongkun; Huang, Zhenwu

2015-03-01

To explore the effects of methylseleninic acid (MeSeA), selenomethionine (SeMet) and methylselenocysteine (MeSeCys) on proliferation, migration and adhesion of HeLa cells. HeLa cells were cultured and treated with MeSeA, SeMet and MeSeCys for 12 - 72 h respectively. MTT assay, healing assay and in vitro cell Matrigel adhesion assay were used to detect the proliferation, migration and adhesion of HeLa cells. Compared to the control group, the proliferation of HeLa cells was remarkably inhibited by MeSeA (P <0. 01). The migration of HeLa cells in MeSeA group was inhibited by 34% (P < 0. 05) and 26% (P < 0. 05) in 4 h and 8 h, respectively. However, the migration of HeLa cells with inhibitions of 18% and 13% was in SeMet group in 4 h and 8 h. The inhibitions of HeLa cell migration in MeSeCys group was 28% (P < 0.05) and 5% in 4 h and 8 h, respectively. In addition, the adhesive function of HeLa cells in the MeSeA group, the SeMet group as well as the MeSeCys group were inhibited by 36% (P < 0. 01), 25% and 49% (P < 0. 01). The proliferation and migration of HeLa cell were effectively inhibited by MeSeA, while the adhesive function of HeLa cell was remarkably inhibited by MeSeCys.

Flavonoids inhibit cytokine-induced endothelial cell adhesion protein gene expression.

PubMed Central

Gerritsen, M. E.; Carley, W. W.; Ranges, G. E.; Shen, C. P.; Phan, S. A.; Ligon, G. F.; Perry, C. A.

1995-01-01

Treatment of human endothelial cells with cytokines such as interleukin-1, tumor necrosis factor-alpha (TNF-alpha) or interferon-gamma induces the expression of specific leukocyte adhesion molecules on the endothelial cell surface. Interfering with either leukocyte adhesion or adhesion protein upregulation is an important therapeutic target as evidenced by the potent anti-inflammatory actions of neutralizing antibodies to these ligands in various animal models and in patients. In the present study we report that cotreatment of human endothelial cells with certain hydroxyflavones and flavanols blocks cytokine-induced ICAM-1, VCAM-1, and E-selectin expression on human endothelial cells. One of the most potent flavones, apigenin, exhibited a dose- and time-dependent, reversible effect on adhesion protein expression as well as inhibiting adhesion protein upregulation at the transcriptional level. Apigenin also inhibited IL-1 alpha-induced prostaglandin synthesis and TNF-alpha-induced IL-6 and IL-8 production, suggesting that the hydroxyflavones may act as general inhibitors of cytokine-induced gene expression. Although apigenin did not inhibit TNF-alpha-induced nuclear translocation of NF-kappa B(p50(NFKB1)/p65(RelA)) we found this flavonoid did inhibit TNF-alpha induced beta-galactosidase activity in SW480 cells stably transfected with a beta-galactosidase reporter construct driven by four NF-kappa B elements, suggesting an action on NF-kappa B transcriptional activation. Adhesion of leukocytes to cytokine-treated endothelial cells was blocked in endothelial cells cotreated with apigenin. Finally, apigenin demonstrated potent anti-inflammatory activity in carrageenan induced rat paw edema and delayed type hypersensitivity in the mouse. We conclude that flavonoids offer important therapeutic potential for the treatment of a variety of inflammatory diseases involving an increase in leukocyte adhesion and trafficking. Images Figure 7 Figure 8 Figure 11 PMID:7543732

Endothelial cell SHP-2 negatively regulates neutrophil adhesion and promotes transmigration by enhancing ICAM-1-VE-cadherin interaction.

PubMed

Yan, Meiping; Zhang, Xinhua; Chen, Ao; Gu, Wei; Liu, Jie; Ren, Xiaojiao; Zhang, Jianping; Wu, Xiaoxiong; Place, Aaron T; Minshall, Richard D; Liu, Guoquan

2017-11-01

Intercellular adhesion molecule-1 (ICAM-1) mediates the firm adhesion of leukocytes to endothelial cells and initiates subsequent signaling that promotes their transendothelial migration (TEM). Vascular endothelial (VE)-cadherin plays a critical role in endothelial cell-cell adhesion, thereby controlling endothelial permeability and leukocyte transmigration. This study aimed to determine the molecular signaling events that originate from the ICAM-1-mediated firm adhesion of neutrophils that regulate VE-cadherin's role as a negative regulator of leukocyte transmigration. We observed that ICAM-1 interacts with Src homology domain 2-containing phosphatase-2 (SHP-2), and SHP-2 down-regulation via silencing of small interfering RNA in endothelial cells enhanced neutrophil adhesion to endothelial cells but inhibited neutrophil transmigration. We also found that VE-cadherin associated with the ICAM-1-SHP-2 complex. Moreover, whereas the activation of ICAM-1 leads to VE-cadherin dissociation from ICAM-1 and VE-cadherin association with actin, SHP-2 down-regulation prevented ICAM-1-VE-cadherin association and promoted VE-cadherin-actin association. Furthermore, SHP-2 down-regulation in vivo promoted LPS-induced neutrophil recruitment in mouse lung but delayed neutrophil extravasation. These results suggest that SHP-2- via association with ICAM-1-mediates ICAM-1-induced Src activation and modulates VE-cadherin switching association with ICAM-1 or actin, thereby negatively regulating neutrophil adhesion to endothelial cells and enhancing their TEM.-Yan, M., Zhang, X., Chen, A., Gu, W., Liu, J., Ren, X., Zhang, J., Wu, X., Place, A. T., Minshall, R. D., Liu, G. Endothelial cell SHP-2 negatively regulates neutrophil adhesion and promotes transmigration by enhancing ICAM-1-VE-cadherin interaction. © FASEB.

Surface wettability of plasma SiOx:H nanocoating-induced endothelial cells' migration and the associated FAK-Rho GTPases signalling pathways

PubMed Central

Shen, Yang; Wang, Guixue; Huang, Xianliang; Zhang, Qin; Wu, Jiang; Tang, Chaojun; Yu, Qingsong; Liu, Xiaoheng

2012-01-01

Vascular endothelial cell (EC) adhesion and migration are essential processes in re-endothelialization of implanted biomaterials. There is no clear relationship and mechanism between EC adhesion and migration behaviour on surfaces with varying wettabilities. As model substrates, plasma SiOx:H nanocoatings with well-controlled surface wettability (with water contact angles in the range of 98.5 ± 2.3° to 26.3 ± 4.0°) were used in this study to investigate the effects of surface wettability on cell adhesion/migration and associated protein expressions in FAK-Rho GTPases signalling pathways. It was found that EC adhesion/migration showed opposite behaviour on the hydrophilic and hydrophobic surfaces (i.e. hydrophobic surfaces promoted EC migration but were anti-adhesions). The number of adherent ECs showed a maximum on hydrophilic surfaces, while cells adhered to hydrophobic surfaces exhibited a tendency for cell migration. The focal adhesion kinase (FAK) inhibitor targeting the Y-397 site of FAK could significantly inhibit cell adhesion/migration, suggesting that EC adhesion and migration on surfaces with different wettabilities involve (p)FAK and its downstream signalling pathways. Western blot results suggested that the FAK-Rho GTPases signalling pathways were correlative to EC migration on hydrophobic plasma SiOx:H surfaces, but uncertain to hydrophilic surfaces. This work demonstrated that surface wettability could induce cellular behaviours that were associated with different cellular signalling events. PMID:21715399

Isoform-specific function of calpains in cell adhesion disruption: studies in postlactational mammary gland and breast cancer.

PubMed

Rodríguez-Fernández, Lucía; Ferrer-Vicens, Iván; García, Concha; Oltra, Sara S; Zaragozá, Rosa; Viña, Juan R; García-Trevijano, Elena R

2016-09-15

Cleavage of adhesion proteins is the first step for physiological clearance of undesired cells during postlactational regression of the mammary gland, but also for cell migration in pathological states such as breast cancer. The intracellular Ca(2+)-dependent proteases, calpains (CAPNs), are known to cleave adhesion proteins. The isoform-specific function of CAPN1 and CAPN2 was explored and compared in two models of cell adhesion disruption: mice mammary gland during weaning-induced involution and breast cancer cell lines according to tumor subtype classification. In both models, E-cadherin, β-catenin, p-120, and talin-1 were cleaved as assessed by western blot analysis. Both CAPNs were able to cleave adhesion proteins from lactating mammary gland in vitro Nevertheless, CAPN2 was the only isoform found to co-localize with E-cadherin in cell junctions at the peak of lactation. CAPN2/E-cadherin in vivo interaction, analyzed by proximity ligation assay, was dramatically increased during involution. Calpain inhibitor administration prevented the cytosolic accumulation of truncated E-cadherin cleaved by CAPN2. Conversely, in breast cancer cells, CAPN2 was restricted to the nuclear compartment. The isoform-specific expression of CAPNs and CAPN activity was dependent on the breast cancer subtype. However, CAPN1 and CAPN2 knockdown cells showed that cleavage of adhesion proteins and cell migration was mediated by CAPN1, independently of the breast cancer cell line used. Data presented here suggest that the subcellular distribution of CAPN1 and CAPN2 is a major issue in target-substrate recognition; therefore, it determines the isoform-specific role of CAPNs during disruption of cell adhesion in either a physiological or a pathological context. © 2016 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

Focal adhesion kinase is involved in mechanosensing during fibroblast migration

NASA Technical Reports Server (NTRS)

Wang, H. B.; Dembo, M.; Hanks, S. K.; Wang, Y.

2001-01-01

Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase localized at focal adhesions and is believed to mediate adhesion-stimulated effects. Although ablation of FAK impairs cell movement, it is not clear whether FAK might be involved in the guidance of cell migration, a role consistent with its putative regulatory function. We have transfected FAK-null fibroblasts with FAK gene under the control of the tetracycline repression system. Cells were cultured on flexible polyacrylamide substrates for the detection of traction forces and the application of mechanical stimulation. Compared with control cells expressing wild-type FAK, FAK-null cells showed a decrease in migration speed and directional persistence. In addition, whereas FAK-expressing cells responded to exerted forces by reorienting their movements and forming prominent focal adhesions, FAK-null cells failed to show such responses. Furthermore, FAK-null cells showed impaired responses to decreases in substrate flexibility, which causes control cells to generate weaker traction forces and migrate away from soft substrates. Cells expressing Y397F FAK, which cannot be phosphorylated at a key tyrosine site, showed similar defects in migration pattern and force-induced reorientation as did FAK-null cells. However, other aspects of F397-FAK cells, including the responses to substrate flexibility and the amplification of focal adhesions upon mechanical stimulation, were similar to that of control cells. Our results suggest that FAK plays an important role in the response of migrating cells to mechanical input. In addition, phosphorylation at Tyr-397 is required for some, but not all, of the functions of FAK in cell migration.

Synaptic excitation mediated by AMPA receptors in rat cerebellar slices is selectively enhanced by aniracetam and cyclothiazide.

PubMed

Boxall, A R; Garthwaite, J

1995-05-01

AMPA receptors mediate fast, glutamatergic synaptic transmission in the central nervous system. The time-course of the associated postsynaptic current has been suggested to be determined principally by the kinetics of glutamate binding and receptor desensitization. Aniracetam and cyclothiazide are drugs capable of selectively preventing desensitization of the AMPA receptor. To investigate the relevance of desensitization to fast synaptic transmission in the cerebellum we have tested these compounds against AMPA-induced depolarizations and postsynaptic potentials using the grease-gap recording technique. Aniracetam (1 microM-5 mM) and cyclothiazide (1 microM-500 microM) both enhanced the depolarising action of AMPA (1 microM) on Purkinje cells in a concentration-dependent manner. At the highest concentrations tested, the increases over controls were approximately 600% and 800% respectively. Aniracetam also increased, in a concentration-dependent manner, the amplitude of the evoked synaptic potentials of both parallel fibre-Purkinje cell and mossy fibre-granule cell pathways, with the highest concentrations tested enhancing the potentials by approximately 60% and 75% respectively. These data suggest that, at two different synapses in the cerebellum, AMPA receptor desensitization occurs physiologically and is likely to contribute to the shape of fast synaptic currents.

[The effect of atropine on the ultrastructural postsynaptic plasticity of the associative type in the rat neocortex].

PubMed

Khludova, G G; Gusev, P A

1998-01-01

The effect of muscarinic antagonist atropine on thickness of postsynaptic density of axodendritic synapses was studied in the sensorimotor region of the brain cortex of rats during paired repeated microapplication of glutamate and acetylcholine. In the applied conditioning paradigm atropine significantly decreased morphological dimensions of the postsynaptic density, however, the control values were not reached. This finding testifies to participation of both muscarinic and nicotinic cholinoreceptors in associative postsynaptic plasticity.

Cold plasma selectivity in the interaction with various types of the cells

NASA Astrophysics Data System (ADS)

Volotskova, Olga; Stepp, Mary Ann; Keidar, Michael

2011-10-01

Present research in the area of cold atmospheric plasma (CAP) demonstrates great potential in various areas including medicine and biology. Depending on their configuration they can be used for wound healing, sterilization, targeted cell/tissue removal, and cancer treatments. Here we explore potential mechanisms by which CAP alters cell migration and influences cell adhesion. The migration studies are focused on the CAP interaction with fibroblasts and corneal epithelial cells. Data show that various types of cells have different thresholds (treatment times) required to achieve maximum inhibition of cell migration which is around ~30-40%. Studies to assess the impact of CAP treatment on the activation state of integrins and focal adhesion size by immunofluorescence showed more active b1 integrin on the cell surface and large focal adhesions after CAP treatment. Based on these data, a thermodynamic model is presented to explain how CAP leads to integrin activation and focal adhesion assembly. Also responses of the various types of the cells to the cold plasma treatment on the example of the epithelial keratinocytes, papilloma and carcinoma cells are studied. Cell cycle, migration and cell vitality analysis were performed. The goal of this study is to understand the mechanism by which the CAP jet alters cell migration, influences adhesion and cell survival.

PI3K{gamma} activation by CXCL12 regulates tumor cell adhesion and invasion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Monterrubio, Maria; Mellado, Mario; Carrera, Ana C.

Tumor dissemination is a complex process, in which certain steps resemble those in leukocyte homing. Specific chemokine/chemokine receptor pairs have important roles in both processes. CXCL12/CXCR4 is the most commonly expressed chemokine/chemokine receptor pair in human cancers, in which it regulates cell adhesion, extravasation, metastatic colonization, angiogenesis, and proliferation. All of these processes require activation of signaling pathways that include G proteins, phosphatidylinositol-3 kinase (PI3K), JAK kinases, Rho GTPases, and focal adhesion-associated proteins. We analyzed these pathways in a human melanoma cell line in response to CXCL12 stimulation, and found that PI3K{gamma} regulates tumor cell adhesion through mechanisms different frommore » those involved in cell invasion. Our data indicate that, following CXCR4 activation after CXCL12 binding, the invasion and adhesion processes are regulated differently by distinct downstream events in these signaling cascades.« less

Association of Cell-adhesion Activities with Virulence in Shiga toxin-producing Escherichia coli O103：H2.

PubMed

Kobayashi, Naoki; Maeda, Eriko; Saito, Shioko; Furukawa, Ichiro; Ohnishi, Takahiro; Watanabe, Maiko; Terajima, Jun; Hara-Kudo, Yukiko

2016-01-01

The characteristics of 11 strains of Stx1-producing and Stx2-non-producing STEC O103：H2 were analyzed to investigate the differences in virulence in a single serotype of Shiga toxin (Stx) -producing Escherichia coli (STEC). Differences in the cell-adhesion activity to Caco-2 cells were observed among the strains. The activity of the one strain, isolated from a patient with hemolytic uremic syndrome was 4-20-fold higher than those of the other strains. Although the strains with high cell-adhesion activity showed high expressions of eae, espB, espD, and tir in the locus of enterocyte effacement related with cell-adhesion, those were not specific for this strain. In addition, the Stx1 production level of the strain was not particularly high. It was indicated that the high adhesion activity might be a potential factor to associate serious symptom.

The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells

PubMed Central

LECKEL, K; BEECKEN, W-D; JONAS, D; OPPERMANN, E; COMAN, M C; BECK, K-F; CINATL, J; HAILER, N P; AUTH, M K H; BECHSTEIN, W O; SHIPKOVA, M; BLAHETA, R A

2003-01-01

Immunosuppression correlates with the development and recurrence of cancer. Mycophenolate mofetil (MMF) has been shown to reduce adhesion molecule expression and leucocyte recruitment into the donor organ. We have hypothesized that MMF might also prevent receptor-dependent tumour dissemination. Therefore, we have investigated the effects of MMF on tumour cell adhesion to human umbilical vein endothelial cells (HUVEC) and compared them with the effects on T cell–endothelial cell interactions. Influence of MMF on cellular adhesion to HUVEC was analysed using isolated CD4+ and CD8+ T cells, or WiDr colon adenocarcinoma cells as the model tumour. HUVEC receptors ICAM-1, VCAM-1, E-selectin and P-selectin were detected by flow cytometry, Western blot or Northern blot analysis. Binding activity of T cells or WiDr cells in the presence of MMF were measured using immobilized receptor globulin chimeras. MMF potently blocked both T cell and WiDr cell binding to endothelium by 80%. Surface expression of the endothelial cell receptors was reduced by MMF in a dose-dependent manner. E-selectin mRNA was concurrently reduced with a maximum effect at 1 µm. Interestingly, MMF acted differently on T cells and WiDr cells. Maximum efficacy of MMF was reached at 10 and 1 µm, respectively. Furthermore, MMF specifically suppressed T cell attachment to ICAM-1, VCAM-1 and P-selectin. In contrast, MMF prevented WiDr cell attachment to E-selectin. In conclusion, our data reveal distinct effects of MMF on both T cell adhesion and tumour cell adhesion to endothelial cells. This suggests that MMF not only interferes with the invasion of alloactivated T cells, but might also be of value in managing post-transplantation malignancy. PMID:14616783

Single and collective cell migration: the mechanics of adhesions

PubMed Central

De Pascalis, Chiara; Etienne-Manneville, Sandrine

2017-01-01

Chemical and physical properties of the environment control cell proliferation, differentiation, or apoptosis in the long term. However, to be able to move and migrate through a complex three-dimensional environment, cells must quickly adapt in the short term to the physical properties of their surroundings. Interactions with the extracellular matrix (ECM) occur through focal adhesions or hemidesmosomes via the engagement of integrins with fibrillar ECM proteins. Cells also interact with their neighbors, and this involves various types of intercellular adhesive structures such as tight junctions, cadherin-based adherens junctions, and desmosomes. Mechanobiology studies have shown that cell–ECM and cell–cell adhesions participate in mechanosensing to transduce mechanical cues into biochemical signals and conversely are responsible for the transmission of intracellular forces to the extracellular environment. As they migrate, cells use these adhesive structures to probe their surroundings, adapt their mechanical properties, and exert the appropriate forces required for their movements. The focus of this review is to give an overview of recent developments showing the bidirectional relationship between the physical properties of the environment and the cell mechanical responses during single and collective cell migration. PMID:28684609

The conveyor belt hypothesis for thymocyte migration: participation of adhesion and de-adhesion molecules.

PubMed

Villa-Verde, D M; Calado, T C; Ocampo, J S; Silva-Monteiro, E; Savino, W

1999-05-01

Thymocyte differentiation is the process by which bone marrow-derived precursors enter the thymus, proliferate, rearrange the genes and express the corresponding T cell receptors, and undergo positive and/or negative selection, ultimately yielding mature T cells that will represent the so-called T cell repertoire. This process occurs in the context of cell migration, whose cellular and molecular basis is still poorly understood. Kinetic studies favor the idea that these cells leave the organ in an ordered pattern, as if they were moving on a conveyor belt. We have recently proposed that extracellular matrix glycoproteins, such as fibronectin, laminin and type IV collagen, among others, produced by non-lymphoid cells both in the cortex and in the medulla, would constitute a macromolecular arrangement allowing differentiating thymocytes to migrate. Here we discuss the participation of both molecules with adhesive and de-adhesive properties in the intrathymic T cell migration. Functional experiments demonstrated that galectin-3, a soluble beta-galactoside-binding lectin secreted by thymic microenvironmental cells, is a likely candidate for de-adhesion proteins by decreasing thymocyte interaction with the thymic microenvironment.

Adhesion molecules and receptors

USDA-ARS?s Scientific Manuscript database

Adhesion molecules are necessary for leukocyte trafficking and differentiation. They serve to initiate cell-cell interactions under conditions of shear, and they sustain the cell-cell and cell-matrix interactions needed for cellular locomotion. They also can serve directly as signaling molecules act...

The state diagram for cell adhesion under flow: leukocyte rolling and firm adhesion.

PubMed

Chang, K C; Tees, D F; Hammer, D A

2000-10-10

Leukocyte adhesion under flow in the microvasculature is mediated by binding between cell surface receptors and complementary ligands expressed on the surface of the endothelium. Leukocytes adhere to endothelium in a two-step mechanism: rolling (primarily mediated by selectins) followed by firm adhesion (primarily mediated by integrins). Using a computational method called "Adhesive Dynamics," we have simulated the adhesion of a cell to a surface in flow, and elucidated the relationship between receptor-ligand functional properties and the dynamics of adhesion. We express this relationship in a state diagram, a one-to-one map between the biophysical properties of adhesion molecules and various adhesive behaviors. Behaviors that are observed in simulations include firm adhesion, transient adhesion (rolling), and no adhesion. We varied the dissociative properties, association rate, bond elasticity, and shear rate and found that the unstressed dissociation rate, k(r)(o), and the bond interaction length, gamma, are the most important molecular properties controlling the dynamics of adhesion. Experimental k(r)(o) and gamma values from the literature for molecules that are known to mediate rolling adhesion fall within the rolling region of the state diagram. We explain why L-selectin-mediated rolling, which has faster k(r)(o) than other selectins, is accompanied by a smaller value for gamma. We also show how changes in association rate, shear rate, and bond elasticity alter the dynamics of adhesion. The state diagram (which must be mapped for each receptor-ligand system) presents a concise and comprehensive means of understanding the relationship between bond functional properties and the dynamics of adhesion mediated by receptor-ligand bonds.

A model study of factors involved in adhesion of Pseudomonas fluorescens to meat.

PubMed Central

Piette, J P; Idziak, E S

1992-01-01

A study was undertaken to investigate the factors involved in the adhesion of Pseudomonas fluorescens to model meat surfaces (tendon slices). Adhesion was fast (less than 2.5 min) and was not suppressed by killing the cells with UV, gamma rays, or heat, indicating that physiological activity was not required. In various salt solutions (NaCl, KCl, CaCl2, MgCl2), adhesion increased with increasing ionic strength up to 10 to 100 mM, suggesting that, at low ionic strengths, electrostatic interactions were involved in the adhesion process. At higher ionic strengths (greater than 10 to 100 mM) or in the presence of Al3+ ions, adhesion was sharply reduced. Selectively blocking of carboxyl or amino groups at the cell surface by chemical means did not affect adhesion. These groups are therefore not directly involved in an adhesive bond with tendon. Given a sufficient cell concentration (10(10) CFU.ml-1) in the adhesion medium, the surface of tendon was almost entirely covered with adherent bacteria. This suggests that if the adhesion is specific, the attachment sites on the tendon surface must be located within collagen or proteoglycan molecules. Images PMID:1444387

Dynamin2 controls Rap1 activation and integrin clustering in human T lymphocyte adhesion

PubMed Central

Eppler, Felix J.

2017-01-01

Leukocyte trafficking is crucial to facilitate efficient immune responses. Here, we report that the large GTPase dynamin2, which is generally considered to have a key role in endocytosis and membrane remodeling, is an essential regulator of integrin-dependent human T lymphocyte adhesion and migration. Chemical inhibition or knockdown of dynamin2 expression significantly reduced integrin-dependent T cell adhesion in vitro. This phenotype was not observed when T cells were treated with various chemical inhibitors which abrogate endocytosis or actin polymerization. We furthermore detected dynamin2 in signaling complexes and propose that it controls T cell adhesion via FAK/Pyk2- and RapGEF1-mediated Rap1 activation. In addition, the dynamin2 inhibitor-induced reduction of lymphocyte adhesion can be rescued by Rap1a overexpression. We demonstrate that the dynamin2 effect on T cell adhesion does not involve integrin affinity regulation but instead relies on its ability to modulate integrin valency. Taken together, we suggest a previously unidentified role of dynamin2 in the regulation of integrin-mediated lymphocyte adhesion via a Rap1 signaling pathway. PMID:28273099

[A new hypothesis for the treatment of amblyopia: the flicker stimulator].

PubMed

Parrozzani, A; Fedriga, P; Ferrari, E; De Vincentiis, L

1984-01-01

A variety of cells are involved in the pathogenesis of amblyopia : ON, OFF, ON-OFF cells, postsynaptic cells, neurons of striate cortex and the select interest of the macula. The need for stimulation of these cells in treating amblyopia forms the theoretical basis of the Flicker stimulator with red monochromatic light (LED, 655 nm). The authors present a clinical investigation on 35 subjects with anisometropic or strabismic amblyopia, before extensive treatment with classic anti-amblyopic techniques without satisfactory improvement obtaining significant statistical results (p less than 0,001).

X-rays effects on cytoskeleton mechanics of healthy and tumor cells.

PubMed

Panzetta, Valeria; De Menna, Marta; Musella, Ida; Pugliese, Mariagabriella; Quarto, Maria; Netti, Paolo A; Fusco, Sabato

2017-01-01

Alterations in the cytoskeleton structure are frequently found in several diseases and particularly in cancer cells. It is also through the alterations of the cytoskeleton structure that cancer cells acquire most of their common features such as uncontrolled cell proliferation, cell death evasion, and the gaining of migratory and invasive characteristics. Although radiation therapies currently represent one of the most effective treatments for patients, the effects of X-irradiation on the cytoskeleton architecture are still poorly understood. In this case we investigated the effects, over time of two different doses of X-ray irradiation, on cell cytoskeletons of BALB/c3T3 and Sv40-transformed BALB/c 3T3 cells (SVT2). Biophysical parameters - focal adhesion size, actin bundles organization, and cell mechanical properties - were measured before and after irradiations (1 and 2 Gy) at 24 and 72 h, comparing the cytoskeleton properties of normal and transformed cells. The differences, before and after X-irradiation, were revealed in terms of cell morphology and deformability. Finally, such parameters were correlated to the alterations of cytoskeleton dynamics by evaluating cell adhesion at the level of focal adhesion and cytoskeleton mechanics. X-irradiation modifies the structure and the activity of cell cytoskeleton in a dose-dependent manner. For transformed cells, radiation sensitively increased cell adhesion, as indicated by paxillin-rich focal adhesion, flat morphology, a well-organized actin cytoskeleton, and intracellular mechanics. On the other hand, for normal fibroblasts IR had negligible effects on cytoskeletal and adhesive protein organization. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Alpha lipoic acid selectively inhibits proliferation and adhesion to fibronectin of v-H-ras-transformed 3Y1 cells.

PubMed

Yamasaki, Masao; Iwase, Masahiro; Kawano, Kazuo; Sakakibara, Yoichi; Suiko, Masahito; Nishiyama, Kazuo

2012-05-01

Here, we focused on the effects of racemic α-lipoic acid on proliferation and adhesion properties of 3Y1 rat fibroblasts and the v-H-ras-transformed derivative, HR-3Y1-2 cells. Racemic α-lipoic acid inhibited proliferation of HR-3Y1-2 but not 3Y1 cells at 0.3 and 1.0 mM. R-(+)-α-lipoic acid also inhibited proliferation of HR-3Y1-2 cells equivalent to that of racemic α-lipoic acid. In addition, racemic α-lipoic acid decreased intracellular reactive oxygen species levels in HR-3Y1 cells but not 3Y1 cells. Next, we evaluated the effects of racemic α-lipoic acid on cell adhesion to fibronectin. The results indicated that racemic α-lipoic acid decreased adhesive ability of HR-3Y1-2 cells to fibronectin-coated plates. As blocking antibody experiment revealed that β1-integrin plays a key role in cell adhesion in this experimental system, the effects of racemic α-lipoic acid on the expression of β1-integrin were examined. The results indicated that racemic α-lipoic acid selectively downregulated the expression of cell surface β1-integrin expression in HR-3Y1-2 cells. Intriguingly, exogenous hydrogen peroxide upregulated cell surface β1-integrin expression in 3Y1 cells. Taken together, these data suggest that reduction of intracellular reactive oxygen species levels by α-lipoic acid could be an effective means of ameliorating abnormal growth and adhesive properties in v-H-ras transformed cells.

State diagram for adhesion dynamics of deformable capsules under shear flow.

PubMed

Luo, Zheng Yuan; Bai, Bo Feng

2016-08-17

Due to the significance of understanding the underlying mechanisms of cell adhesion in biological processes and cell capture in biomedical applications, we numerically investigate the adhesion dynamics of deformable capsules under shear flow by using a three-dimensional computational fluid dynamic model. This model is based on the coupling of the front tracking-finite element method for elastic mechanics of the capsule membrane and the adhesion kinetics simulation for adhesive interactions between capsules and functionalized surfaces. Using this model, three distinct adhesion dynamic states are predicted, such as detachment, rolling and firm-adhesion. Specifically, the effects of capsule deformability quantified by the capillary number on the transitions of these three dynamic states are investigated by developing an adhesion dynamic state diagram for the first time. At low capillary numbers (e.g. Ca < 0.0075), whole-capsule deformation confers the capsule a flattened bottom in contact with the functionalized surface, which hence promotes the rolling-to-firm-adhesion transition. It is consistent with the observations from previous studies that cell deformation promotes the adhesion of cells lying in the rolling regime. However, it is surprising to find that, at relatively high capillary numbers (e.g. 0.0075 < Ca < 0.0175), the effect of capsule deformability on its adhesion dynamics is far more complex than just promoting adhesion. High deformability of capsules makes their bottom take a concave shape with no adhesion bond formation in the middle. The appearance of this specific capsule shape inhibits the transitions of both rolling-to-firm-adhesion and detachment-to-rolling, and it means that capsule deformation no longer promotes the capsule adhesion. Besides, it is interesting to note that, when the capillary number exceeds a critical value (e.g. Ca = 0.0175), the rolling state no longer appears, since capsules exhibit large deviation from the spherical shape.

[Study on the effect of phloretin on inhibiting malignant pheotype of BEL-7402 cells].

PubMed

Luo, Hui; Wang, Ya-Jun; Chen, Jie; Liu, Jiang-Qin

2008-07-01

To investigate the effect of phloretin on inhibiting BEL-7402 cells' growth, invasive, migration and adhesion ability and the rate of colony formation. BEL-7402 cells' growth, invasive, migration and adhesion ability and the rate of colony formation were examined with MIT method and Costar Transwell. Phloretin inhibited the growth, invasive, migration and adhesion ability of BEL-7402 cells and reduced the rate of colony formation in dose-dependent. Phloretin can inhibit BEL-7402 cells' malignant pheotype.

Calcium influx through the TRPV1 channel of endothelial cells (ECs) correlates with a stronger adhesion between monocytes and ECs.

PubMed

Himi, N; Hamaguchi, A; Hashimoto, K; Koga, T; Narita, K; Miyamoto, O

2012-01-01

Atherosclerosis is thought to be initiated by the transendothelial migration of monocytes. In the early stage of this process, the adhesion of monocytes to endothelial cells is supported by an increase in the intracellular concentration of calcium ion ([Ca(2+)]i) in endothelial cells. However, the main source of Ca(2+) has been unclear. In this study, the changes in ionic transmittance and [Ca(2+)]i due to the adhesion of monocytes were continuously measured by an electrophysiological technique and fluorescent imaging. Especially, we focused on transient receptor potential vanilloid channel 1 (TRPV1) as a Ca(2+) channel that could influence the adhesion of monocytes. Whole-cell current was continuously recorded in human umbilical vein endothelial cells (HUVECs) by a patch electrode. The adhesion of monocytes (THP-1) induced a transient inward current in HUVECs, as well as an elevation of [Ca(2+)]i. This inward element was abolished by the application of 100 nM SB366,791, a selective antagonist of TRPV1 channel. Furthermore, SB366,791 significantly decreased the number of THP-1 cells that adhered to HUVECs (control: 231 ± 38, SB366,791: 96 ± 16 cells/mm2). These results suggest that an inward calcium current via the TRPV1 channels of endothelial cells correlates with a stronger adhesion between monocytes and endothelial cells.