generation-scalable-metallic-high-aspect-ratio-nanocomposites

Generation of Scalable, Metallic High-Aspect Ratio Nanocomposites in a Biological Liquid Medium

The goal of this protocol is to describe the synthesis of two novel biocomposites with high-aspect ratio structures. The biocomposites consist of copper ...

Louisiana Tech University

Secreted phospholipases A2 (sPLA2s) modulate neuronal survival and neurotransmitter release. Here we show that sPLA2 (group III) synergistically increases glutamate-induced cell death and intracellular calcium ([Ca2+]i) in cultured primary cortical and hippocampal neurons. Whereas 1 microM glutamate elicited transient [Ca2+]i increases in all neurons that recovered 66% to baseline, 25 ng/ml sPLA2 pretreatment resulted in sustained [Ca2+]i increases, with only 5% recovery. At 250 nM glutamate, 25% of neurons failed to respond, and the average recovery time was 101 +/- 12 sec; sPLA2 increased recovery time to 158 +/- 6 sec, and only 2% of cells failed to respond. Both the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 and the calcium-channel blocker cobalt inhibited this effect. Experiments with the glutamate uptake inhibitor L-trans-pyrollidine-2,4-dicarboxylic acid (2.5 microM) indicated that glutamate uptake sites are not a likely modulation point by sPLA2, whereas arachidonic acid (AA) potentiated calcium responses to glutamate. Thus the enhancement of glutamate-induced [Ca2+]i increases by sPLA2 may be due to modulation at NMDA receptors and/or calcium channels by AA. These results indicate that sPLA2 affects neuronal responses to both nontoxic (0.1-10 microM) and toxic (=25 microM) concentrations of glutamate, implicating this enzyme in neuronal functions in pathology.

Secreted phospholipase A2 potentiates glutamate-induced calcium increase and cell death in primary neuronal cultures.

The lipid mediators generated by phospholipases A(2) (PLA(2)), free arachidonic acid (AA), eicosanoids, and platelet-activating factor, modulate neuronal activity; when overproduced, some of them become potent neurotoxins. We have shown, using primary cortical neuron cultures, that glutamate and secretory PLA(2) (sPLA(2)) from bee venom (bv sPLA(2)) and Taipan snake venom (OS2) elicit synergy in inducing neuronal cell death. Low concentrations of sPLA(2) are selective ligands of cell-surface sPLA(2) receptors. We investigated which neuronal arachidonoyl phospholipids are targeted by glutamate-activated cytosolic calcium-dependent PLA(2) (cPLA(2)) and by sPLA(2). Treatment of (3)H-AA-labeled cortical neurons with mildly toxic concentrations of sPLA(2) (25 ng/ml, 1.78 nM) for 45 min resulted in a two- to threefold higher loss of (3)H-AA from phosphatidylcholine (PC) than from phosphatidylethanolamine (PE) and in minor changes in other phospholipids. A similar profile, although of greater magnitude, was observed 20 hr posttreatment. Glutamate (80 microM) induced much less mobilization of (3)H-AA than did sPLA(2) and resulted in a threefold greater degradation of (3)H-AA PE than of (3)H-AA PC by 20 hr posttreatment. Combining sPLA(2) and glutamate resulted in a greater degradation of PC and PE, and the N-methyl-D-aspartate receptor antagonist MK-801 only blocked glutamate effects. Thus, activation of the arachidonate cascade induced by glutamate and sPLA(2) under experimental conditions that lead to neuronal cell death involves the hydrolysis of different (perhaps partially overlapping) cellular phospholipid pools.

Glutamate signalling and secretory phospholipase A2 modulate the release of arachidonic acid from neuronal membranes.

Secreted phospholipase A(2) (sPLA(2)) has been shown to cause both necrotic and apoptotic neuronal injury in the central nervous system. The acute effects of group III sPLA(2) (sPLA(2)-III) include an ability to potentiate glutamate-induced necrosis and calcium influx in primary rat neuronal cultures. Thus, a brief exposure to glutamate and sPLA(2)-III results in an immediate potentiated calcium influx and a potentiated necrosis the following day after treatment, compared to addition of either agonist alone. In contrast to these acute effects, sPLA(2) from both groups IB and IIA have been shown to induce neuronal cell death via apoptosis. Here, we offer the first evidence that group III sPLA(2) potently causes extensive neuronal apoptosis in rat cortical cultures, as evidenced by DNA fragmentation. Distinct from this effect, even high concentrations of glutamate did not cause neuronal damage under the same conditions, and there was no evidence of sPLA(2) damage to cortical astrocytes.

Group III secreted phospholipase A2 causes apoptosis in rat primary cortical neuronal cultures.

Platelet-activating factor (PAF) is a potent phospholipid messenger in the nervous system that participates in synaptic plasticity and in pathologic processes, including neurodegeneration. Oxidative stress plays important roles in neuronal cell death. To define the interaction between PAF and oxidative radicals in neuronal death, we studied the effects of PAF in the presence of oxidative radicals in primary neurons in culture. Exogenous PAF (50 microM) caused PAF receptor-independent injury to neurons. A nonneurotoxic PAF concentration (500 nM) potentiated neuronal death caused by hydrogen peroxide as determined by lactate dehydrogenase (LDH) assay, Hoechst staining, and TUNEL analysis, but it did not potentiate neuronal death caused by menadione, a superoxide donor, or by the nitric oxide donors 3-morpholino-sydnonimine (SIN-1) and sodium nitroprusside (SNP). This potentiation of the hydrogen peroxide effect was selectively blocked by a PAF membrane-receptor antagonist, BN52021 (5 microM). The neurotoxic effect of PAF and hydrogen peroxide was also completely blocked by ebselen and partially decreased by pretreatment with (S)-3,5-dihydroxyphenylglycine (DHPG), a group I metabotropic glutamate receptor (mGluR) agonist. This study suggests that PAF-receptor antagonists may be useful for neuroprotection. A similar effect might also be obtained with group I mGluR agonists, probably by way of a different underlying mechanism.

Interplay among platelet-activating factor, oxidative stress, and group I metabotropic glutamate receptors modulates neuronal survival.

The terminal deoxynucleotidyl transferase (TdT) dUTP nick end labeling (TUNEL) stain is in wide use for measuring apoptosis in neurons, as well as in other cell types. TUNEL may give false positive results due to variations in labeling technique as well as staining of cells that have undergone non-apoptotic DNA strand breaks. Therefore, in isolation, TUNEL is not a certain indicator of apoptosis. Recently, we have demonstrated the potent apoptotic effect of secreted phospholipase A2 from group III (sPLA2-III) on primary cortical neurons from rat. Here we describe a computer-assisted method for quantifying TUNEL-positive neurons after sPLA2-III induced apoptosis. Extent of TUNEL is normalized to total nuclear content using 4',6-diamidino-2-phenylindole (DAPI) staining. Furthermore, DAPI counterstaining allows for determination of a nuclear morphology indicator, based on nuclear size and roundness, which we call the nuclear area factor. We found that the nuclear area factor is an early indicator of cell death (significant after 4 h post treatment), while TUNEL staining is significant at later times (26 h). Thus, the independent staining techniques using TUNEL and DAPI complement each other, and with commercially available image analysis software, may be used to indicate early as well as delayed cell injury processes.

Quantification of sPLA2-induced early and late apoptosis changes in neuronal cell cultures using combined TUNEL and DAPI staining.

Inflammatory stimuli such as lipopolysaccharide increase nitric oxide and secreted phospholipase A2 release from glial cells. However, the signaling mechanism(s) regulating secreted phospholipase A2 in glial cells is not known. Here, rat brain astrocytes treated with lipopolysaccharide generated nitrite and released secreted phospholipase A2, while microglia generated nitrite without releasing secreted phospholipase A2. Unexpectedly, attenuation of nitrite production by pretreatment with the nitric oxide synthase inhibitor N-Omega-nitro-L-arginine methyl ester greatly enhanced lipopolysaccharide-stimulated secreted phospholipase A2 release from astrocytes; postreatment with N-Omega-nitro-L-arginine methyl ester did not potentiate secreted phospholipase A2 release, and addition of an nitric oxide donor attenuated the secreted phospholipase A2 release. The regulation of secreted phospholipase A2 may act via the transcription factor nuclear factor-kappaB, as a nuclear factor-kappaB inhibitor attenuated lipopolysaccharide-stimulated secreted phospholipase A2 release. These results demonstrate the role of basal nitric oxide levels as a regulator of inflammatory secreted phospholipase A2 release from glial cells of the brain.

Role of nitric oxide in regulating secreted phospholipase A2 release from astrocytes.

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Micropatterns of different biomaterials with micro- and nanoscale features and defined spatial arrangement on a single substrate are useful tools for studying cellular-level interactions, and recent reports have highlighted the strong influence of scaffold compliance in determining cell behavior. In this paper, a simple yet versatile and precise patterning technique for the fabrication of interdigitated micropatterns of nanocomposite multilayer coatings on a single substrate is demonstrated through a combination of lithography and layer-by-layer (LbL) assembly processes, termed polymer surface micromachining (PSM). The first nanofilm pattern is constructed using lithography, followed by LbL multilayer assembly and lift-off, and the process is repeated with optical alignment to obtain interdigitated patterns on the same substrate. Thus, the method is analogous to surface micromachining, except that the deposition materials are polymers and biological materials that are used to produce multilayer nanocomposite structures. A key feature of the multilayers is the capability to tune properties such as stiffness by appropriate selection of materials, deposition conditions, and postdeposition treatments. Two- and four-component systems on glass coverslips are presented to demonstrate the versatility of the approach to construct precisely defined, homogeneous nanofilm patterns. In addition, an example of a complex system used as a testbed for in vitro cell adhesion and growth is provided: micropatterns of poly(sodium 4-styrenesulfonate)/poly-L-lysine hydrobromide (PSS/PLL) and secreted phospholipase A(2)/poly(ethyleneimine) (sPLA(2)/PEI) multilayers. The interdigitated square nanofilm array patterns were obtained on a single coverslip with poly(diallyldimethylammonium chloride) (PDDA) as a cell-repellent background. Cell culture experiments show that cortical neurons respond and bind specifically to the sPLA(2) micropatterns in competition with PLL micropatterns. The fabrication and the initial biological results on the nanofilm micropatterns support the usefulness of this technique for use in studies aimed at elucidating important biological structure-function relationships, but the applicability of the fabrication method is much broader and may impact electronics, photonics, and chemical microsystems.

Fabrication of interdigitated micropatterns of self-assembled polymer nanofilms containing cell-adhesive materials.

Micromotifs de diff&eacute;rents biomat&eacute;riaux avec micro - et nanom&eacute;trique caract&eacute;ristiques et une disposition spatiale d&eacute;finie sur un m&ecirc;me substrat sont des outils utiles pour l&#39;&eacute;tude des interactions au niveau cellulaire, et des rapports r&eacute;cents ont mis en &eacute;vidence la forte influence de conformit&eacute; &eacute;chafaudage pour d&eacute;terminer le comportement de la cellule. Dans cet article, une technique de mod&eacute;lisation simple mais polyvalent et pr&eacute;cis pour la fabrication des micromotifs interdigit&eacute;es de nanocomposites rev&ecirc;tements multicouches sur un m&ecirc;me substrat est d&eacute;montr&eacute;e gr&acirc;ce &agrave; une combinaison de lithographie et de processus d&#39;assemblage (LbL) couche par couche, appel&eacute;es surface micro-usinage de polym&egrave;re (PSM). Le premier mod&egrave;le nanofilm est construit &agrave; l&#39;aide de la lithographie, suivie de LbL Assembl&eacute;e multicouche et de d&eacute;collage, et le processus est r&eacute;p&eacute;t&eacute; avec un alignement optique pour obtenir des motifs entrelac&eacute;s sur un m&ecirc;me support. Ainsi, la m&eacute;thode est analogue &agrave; la surface micro usinage, sauf que les mat&eacute;riaux de d&eacute;p&ocirc;ts sont des polym&egrave;res et mat&eacute;riaux biologiques qui sont utilis&eacute;s pour produire des structures multicouches nanocomposite. Un &eacute;l&eacute;ment cl&eacute; des multicouches est la capacit&eacute; d&#39;accorder des propri&eacute;t&eacute;s telles que la rigidit&eacute; en s&eacute;lection appropri&eacute;e des mat&eacute;riaux, les conditions de d&eacute;p&ocirc;t et traitements postdeposition. Les syst&egrave;mes bi-composants et quatre-sur couvre-objet en verre sont pr&eacute;sent&eacute;s pour montrer la polyvalence de la d&eacute;marche pour construire des mod&egrave;les pr&eacute;cis&eacute;ment d&eacute;finis, homog&egrave;ne nanofilm. En outre, un exemple d&#39;un syst&egrave;me complexe utilis&eacute; comme un banc d&#39;essai pour la croissance et l&#39;adh&eacute;rence cellulaire in vitro est fourni : micropatterns du bromhydrate de poly(sodium 4-styrenesulfonate)/poly-L-lysine (PSS/PLL) et phospholipase s&eacute;cr&eacute;t&eacute;e A(2)/poly(ethyleneimine) (sPLA(2)/PEI) multicouches. Les mod&egrave;les de tableau interdigit&eacute;es nanofilm carr&eacute;s ont &eacute;t&eacute; obtenus sur un lamelle couvre-objet unique avec poly(diallyldimethylammonium chloride) (PDDA) comme un arri&egrave;re-plan de cellule-produit r&eacute;pulsif. Les exp&eacute;riences de culture cellulaire montrent que les neurones corticaux r&eacute;pondent et se lient sp&eacute;cifiquement &agrave; la micromotifs de sPLA(2) en concurrence avec PLL micropatterns. La fabrication et les premiers r&eacute;sultats biologiques sur les micropatterns nanofilm soutiennent l&#39;utilit&eacute; de cette technique pour une utilisation dans des &eacute;tudes visant &agrave; &eacute;lucider les relations structure-fonction biologique importante, mais l&#39;applicabilit&eacute; de la m&eacute;thode de fabrication est beaucoup plus large et peut-&ecirc;tre avoir un impact &eacute;lectronique, photonique et microsyst&egrave;mes chimique.

Fabrication de micropatterns interdigit&eacute;es de nanofilms auto-assembl&eacute;s polym&egrave;re contenant des mat&eacute;riaux cellulaires-adh&eacute;sif.

Micropatterns von verschiedenen Biomaterialien mit Mikro- und nanoskaligen Eigenschaften und definierten r&auml;umlichen Anordnung auf einem einzigen Substrat sind n&uuml;tzliche Werkzeuge f&uuml;r die zellul&auml;re Ebene Interaktionen zu studieren, und die j&uuml;ngsten Berichte haben den starken Einfluss der Ger&uuml;st-Compliance bei der Bestimmung der Zelle Verhalten hervorgehoben. In diesem Papier wird eine einfache und doch vielseitige und genaue Musterung-Technik zur Herstellung von ineinandergreifenden Micropatterns Nanocomposite mehrlagige Beschichtungen auf einem einzigen Substrat durch eine Kombination von Lithographie und Schicht f&uuml;r Schicht (LbL) Montageprozesse, genannt Polymer Oberfl&auml;che Micromachining (PSM) demonstriert. Das erste ist-Muster ist mit Lithographie, gefolgt von LbL mehrlagige Montage- und Lift-Off, und der Vorgang wiederholt sich mit optischen Ausrichtung ineinandergreifenden Muster auf dem gleichen Substrat zu erhalten. Somit ist die Methode analog zur Oberfl&auml;che Micromachining, au&szlig;er dass die Absetzung-Materialien sind Polymere und biologische Materialien, die verwendet werden, um mehrlagige Nanocomposite Strukturen zu produzieren. Ein wesentliches Merkmal der Multischichten ist die M&ouml;glichkeit, die Eigenschaften wie Steifigkeit durch entsprechende Auswahl der Materialien, Absetzung Bedingungen und postdeposition Behandlungen zu optimieren. Zwei und vier-Komponenten-Systeme auf Glasdeckgl&auml;ser werden vorgestellt, um die Vielseitigkeit der Ansatz zum Erstellen von genau definierten, homogene diesen Mustern zu demonstrieren. Dar&uuml;ber hinaus wird ein Beispiel f&uuml;r ein komplexes System verwendet als Versuchstr&auml;ger f&uuml;r in-vitro-Zelladh&auml;sion und Wachstum bereitgestellt: Micropatterns poly(sodium 4-styrenesulfonate)/Poly-L-Lysin-Hydrobromid (PSS/PLL) und sezernierte Phospholipase A(2)/poly(ethyleneimine) (sPLA(2)/PEI) Multischichten. Die ineinandergreifenden Quadrat ist Array Muster wurden auf einem einzigen Deckglas mit Poly(diallyldimethylammonium chloride) (PDDA) als Zelle-abweisenden Hintergrund erzielt. Zelle Kultur Experimente zeigen, dass die kortikale Neuronen reagieren und speziell auf die sPLA(2)-Micropatterns im Wettbewerb mit PLL-Micropatterns zu binden. Die Fertigung und die ersten biologischen Ergebnisse auf die ist-Micropatterns unterst&uuml;tzen die N&uuml;tzlichkeit dieser Technik f&uuml;r den Einsatz in Studien, die darauf abzielen, elucidating wichtige biologische Struktur-Funktionsbeziehungen, aber die Anwendbarkeit der Fabrikation Methode ist viel breiter und Elektronik, Photonik und chemischen Microsystems auswirken kann.

Herstellung von ineinandergreifenden Micropatterns von monomolekularen Polymer Nanofilms Zelle klebender Materialien enthalten.

Microdisegni diversi biomateriali con caratteristiche micro - e nanometriche e disposizione spaziale definito su un unico substrato sono strumenti utili per lo studio delle interazioni a livello cellulare, e i rapporti recenti hanno messo in evidenza la forte influenza della conformit&agrave; impalcature nel determinare il comportamento delle cellule. In questa carta, una tecnica di modellatura semplice ma versatile e preciso per la fabbricazione di microdisegni cosparse di nanocompositi rivestimenti multistrato su un singolo substrato &egrave; dimostrata attraverso una combinazione di litografia e processi di assemblaggio (LbL) strato dopo strato, chiamati polimero micromachining di superficie (PSM). Il primo modello di intercollegamento &egrave; costruito utilizzando la litografia, seguita dall&#39;Assemblea multistrato LbL e decollo, e il processo viene ripetuto con allineamento ottico per ottenere modelli cosparse sul substrato stesso. Cos&igrave;, il metodo &egrave; analogo alla superficie micromachining, tranne per il fatto che i materiali di deposizione sono polimeri e materiali biologici che vengono utilizzati per produrre strutture multistrato nanocompositi. Una caratteristica fondamentale del multistrato &egrave; la capacit&agrave; di sintonizzare le propriet&agrave; quali la rigidit&agrave; di appropriata selezione dei materiali, condizioni di deposizione e trattamenti postdeposition. Sistemi a due e quattro componenti su lamelle di vetro sono presentati per dimostrare la versatilit&agrave; dell&#39;approccio per costruire modelli di intercollegamento omogeneo, definito precisamente. Inoltre, viene fornito un esempio di un sistema complesso, utilizzato come banco di prova per la crescita e l&#39;adesione delle cellule in vitro: Microfantasie di poly(sodium 4-styrenesulfonate)/poly-L-lisina hydrobromide (PSS/PLL) e fosfolipasi secreta A(2)/poly(ethyleneimine) (sPLA(2)/PEI) multistrato. I modelli di matrice interdigitating intercollegamento quadrati sono stati ottenuti su un unico vetrino coprioggetto con poly(diallyldimethylammonium chloride) (PDDA) come sfondo cella-repellente. Cella cultura esperimenti mostrano che i neuroni corticali rispondono e legano specificamente per il sPLA(2) microdisegni in concorrenza con PLL microdisegni. La fabbricazione e i primi risultati biologici sull&#39;intercollegamento microdisegni supportano l&#39;utilit&agrave; di questa tecnica per l&#39;utilizzo in studi volti a chiarire le relazioni struttura-funzione biologica importante, ma l&#39;applicabilit&agrave; del metodo di fabbricazione &egrave; molto pi&ugrave; ampia e possa influenzare microsystems chimica elettronica e fotonica.

Fabbricazione di microdisegni cosparse di polimero auto-assemblati nanofilms contenenti materiali adesivi cell.

&#x7570;&#x306A;&#x308B;&#x751F;&#x4F53;&#x6750;&#x6599;&#x306E;&#x30DE;&#x30A4;&#x30AF;&#x30ED;&#x304A;&#x3088;&#x3073;&#x30CA;&#x30CE;&#x30B9;&#x30B1;&#x30FC;&#x30EB;&#x306E;&#x6A5F;&#x80FD;&#x3068;&#x5B9A;&#x7FA9;&#x6E08;&#x307F;&#x306E;&#x7A7A;&#x9593;&#x7684;&#x914D;&#x7F6E;&#x5358;&#x4E00;&#x57FA;&#x677F;&#x4E0A;&#x306E;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x7D30;&#x80DE;&#x30EC;&#x30D9;&#x30EB;&#x306E;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x3092;&#x7814;&#x7A76;&#x3059;&#x308B;&#x305F;&#x3081;&#x306E;&#x4FBF;&#x5229;&#x306A;&#x30C4;&#x30FC;&#x30EB;&#x3067;&#x3042;&#x308A;&#x3001;&#x6700;&#x8FD1;&#x306E;&#x30EC;&#x30DD;&#x30FC;&#x30C8; &#x30BB;&#x30EB;&#x306E;&#x52D5;&#x4F5C;&#x3092;&#x6C7A;&#x5B9A;&#x3059;&#x308B;&#x8DB3;&#x5834;&#x30B3;&#x30F3;&#x30D7;&#x30E9;&#x30A4;&#x30A2;&#x30F3;&#x30B9;&#x306E;&#x5F37;&#x3044;&#x5F71;&#x97FF;&#x529B;&#x3092;&#x5F37;&#x8ABF;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x672C;&#x7A3F;&#x3067;&#x306F;&#x3001;&#x5358;&#x7D14;&#x306A;&#x307E;&#x3060;&#x6C4E;&#x7528;&#x6027;&#x3068;&#x6B63;&#x78BA;&#x306A;&#x30D1;&#x30BF;&#x30FC;&#x30CB;&#x30F3;&#x30B0;&#x6280;&#x8853;&#x306E;&#x5358;&#x4E00;&#x57FA;&#x677F;&#x30CA;&#x30CE;&#x30B3;&#x30F3;&#x30DD;&#x30B8;&#x30C3;&#x30C8;&#x591A;&#x5C64;&#x819C;&#x306E;&#x6ADB;&#x578B;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x4F5C;&#x88FD;&#x306E;&#x305F;&#x3081;&#x30EA;&#x30BD;&#x30B0;&#x30E9;&#x30D5;&#x30A3;&#x3068;&#x30DD;&#x30EA;&#x30DE;&#x30FC;&#x8868;&#x9762;&#x30DE;&#x30A4;&#x30AF;&#x30ED;&#x30DE;&#x30B7;&#x30CB;&#x30F3;&#x30B0; (PSM) &#x3068;&#x547C;&#x3070;&#x308C;&#x308B;&#x5C64;&#x306B;&#x3088;&#x3063;&#x3066;&#x5C64; (LbL) &#x30A2;&#x30BB;&#x30F3;&#x30D6;&#x30EA; &#x30D7;&#x30ED;&#x30BB;&#x30B9;&#x306E;&#x7D44;&#x307F;&#x5408;&#x308F;&#x305B;&#x3092;&#x4ECB;&#x3057;&#x3066;&#x793A;&#x3055;&#x308C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;LbL &#x591A;&#x5C64;&#x30A2;&#x30BB;&#x30F3;&#x30D6;&#x30EA;&#x3068;&#x30EA;&#x30D5;&#x30C8;&#x30AA;&#x30D5;&#x304C;&#x7D9A;&#x304D;&#x307E;&#x3059;&#x30EA;&#x30BD;&#x30B0;&#x30E9;&#x30D5;&#x30A3;&#x30FC;&#x306B;&#x3088;&#x308B;&#x6700;&#x521D;&#x306E;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x69CB;&#x7BC9;&#x3057;&#x3001;&#x6ADB;&#x578B;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3068;&#x540C;&#x3058;&#x57FA;&#x677F;&#x4E0A;&#x3092;&#x53D6;&#x5F97;&#x3059;&#x308B;&#x5149;&#x5B66;&#x914D;&#x7F6E;&#x3092;&#x6301;&#x3064;&#x30D7;&#x30ED;&#x30BB;&#x30B9;&#x304C;&#x7E70;&#x308A;&#x8FD4;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x3057;&#x305F;&#x304C;&#x3063;&#x3066;&#x3001;&#x30DD;&#x30EA;&#x30DE;&#x30FC;&#x3068;&#x591A;&#x5C64;&#x30CA;&#x30CE;&#x30B3;&#x30F3;&#x30DD;&#x30B8;&#x30C3;&#x30C8;&#x306E;&#x69CB;&#x9020;&#x3092;&#x751F;&#x6210;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x4F7F;&#x7528;&#x3055;&#x308C;&#x308B;&#x751F;&#x7269;&#x6750;&#x6599;&#x304C;&#x84B8;&#x7740;&#x6750;&#x6599;&#x3092;&#x3042;&#x308B;&#x3053;&#x3068;&#x3092;&#x9664;&#x3051;&#x3070;&#x3001;&#x30E1;&#x30BD;&#x30C3;&#x30C9;&#x306F;&#x8868;&#x9762;&#x30DE;&#x30A4;&#x30AF;&#x30ED;&#x30DE;&#x30B7;&#x30CB;&#x30F3;&#x30B0;&#x306B;&#x4F3C;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;Co/cu &#x306E;&#x4E3B;&#x306A;&#x6A5F;&#x80FD;&#x306F;&#x3001;&#x9069;&#x5207;&#x306A;&#x6750;&#x6599;&#x3001;&#x6210;&#x819C;&#x6761;&#x4EF6;&#x304A;&#x3088;&#x3073; postdeposition &#x306E;&#x6CBB;&#x7642;&#x6CD5;&#x306E;&#x9078;&#x629E;&#x306B;&#x3088;&#x308B;&#x525B;&#x6027;&#x306A;&#x3069;&#x306E;&#x30D7;&#x30ED;&#x30D1;&#x30C6;&#x30A3;&#x3092;&#x8ABF;&#x6574;&#x3059;&#x308B;&#x6A5F;&#x80FD;&#x3067;&#x3059;&#x3002;2 &#x304A;&#x3088;&#x3073; 4 &#x6210;&#x5206;&#x7CFB;&#x30AC;&#x30E9;&#x30B9; coverslips &#x306B;&#x6B63;&#x78BA;&#x306B;&#x5B9A;&#x7FA9;&#x3057;&#x3001;&#x5747;&#x8CEA;&#x306A;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x69CB;&#x7BC9;&#x3078;&#x306E;&#x30A2;&#x30D7;&#x30ED;&#x30FC;&#x30C1;&#x306E;&#x591A;&#x69D8;&#x6027;&#x3092;&#x793A;&#x3059;&#x305F;&#x3081;&#x306B;&#x8868;&#x793A;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x3055;&#x3089;&#x306B;&#x3001;in vitro &#x306B;&#x304A;&#x3051;&#x308B;&#x7D30;&#x80DE;&#x63A5;&#x7740;&#x3068;&#x6210;&#x9577;&#x306E;&#x305F;&#x3081;&#x306E;&#x30C6;&#x30B9;&#x30C8;&#x30D9;&#x30C3;&#x30C9;&#x3068;&#x3057;&#x3066;&#x4F7F;&#x7528;&#x3055;&#x308C;&#x308B;&#x8907;&#x96D1;&#x306A;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x306E;&#x4F8B;&#x304C;&#x63D0;&#x4F9B;&#x3055;&#x308C;&#x307E;&#x3059;&#xFF1A; &#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306E; poly(sodium 4-styrenesulfonate)/&#x30DD;&#x30EA;-L-&#x30EA;&#x30B8;&#x30F3;&#x81ED;&#x5316;&#x6C34;&#x7D20; (PSS/PLL) &#x3068;&#x5206;&#x6CCC;&#x6027;&#x30DB;&#x30B9;&#x30DB;&#x30EA;&#x30D1;&#x30FC;&#x30BC; A(2)/poly(ethyleneimine) (sPLA(2)/PEI) &#x591A;&#x5C64;&#x819C;&#x3002;&#x6ADB;&#x578B;&#x6B63;&#x65B9;&#x5F62;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x306E;&#x914D;&#x5217;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x4E0A;&#x5358;&#x4E00; coverslip poly(diallyldimethylammonium chloride) (PDDA) &#x3068;&#x30BB;&#x30EB;&#x64A5;&#x80CC;&#x666F;&#x3068;&#x3057;&#x3066;&#x5F97;&#x3089;&#x308C;&#x307E;&#x3057;&#x305F;&#x3002;&#x7D30;&#x80DE;&#x57F9;&#x990A;&#x5B9F;&#x9A13;&#x3067;&#x306F;&#x76AE;&#x8CEA;&#x30CB;&#x30E5;&#x30FC;&#x30ED;&#x30F3;&#x5FDC;&#x7B54;&#x3053;&#x3068;&#x3092;&#x793A;&#x3059;&#x5177;&#x4F53;&#x7684;&#x306B;&#x7AF6;&#x4E89;&#x306E;&#x4E2D;&#x3067; sPLA(2) &#x306E;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3; PLL &#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3067;&#x30D0;&#x30A4;&#x30F3;&#x30C9;&#x3057;&#x307E;&#x3059;&#x3002;&#x4F5C;&#x88FD;&#x3068;&#x521D;&#x671F;&#x306E;&#x751F;&#x7269;&#x5B66;&#x7684;&#x7D50;&#x679C;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306B;&#x672C;&#x624B;&#x6CD5;&#x306E;&#x6709;&#x7528;&#x6027;&#x7814;&#x7A76;&#x306E;&#x91CD;&#x8981;&#x306A;&#x751F;&#x7269;&#x5B66;&#x7684;&#x69CB;&#x9020;&#x3068;&#x6A5F;&#x80FD;&#x306E;&#x95A2;&#x4FC2;&#x3092;&#x89E3;&#x660E;&#x3059;&#x308B;&#x76EE;&#x7684;&#x3067;&#x4F7F;&#x7528;&#x3059;&#x308B;&#x305F;&#x3081;&#x30B5;&#x30DD;&#x30FC;&#x30C8;&#x3057;&#x307E;&#x3059;&#x304C;&#x3001;&#x4F5C;&#x88FD;&#x6CD5;&#x306E;&#x9069;&#x7528;&#x6027;&#x304C;&#x306F;&#x308B;&#x304B;&#x306B;&#x5E83;&#x3044;&#x3067;&#x3059;&#x3057;&#x3001;&#x30A8;&#x30EC;&#x30AF;&#x30C8;&#x30ED;&#x30CB;&#x30AF;&#x30B9;&#x3001;&#x30D5;&#x30A9;&#x30C8;&#x30CB;&#x30AF;&#x30B9;&#x3001;&#x304A;&#x3088;&#x3073;&#x30DE;&#x30A4;&#x30AF;&#x30ED;&#x5316;&#x5B66;&#x30B7;&#x30B9;&#x30C6;&#x30E0;&#x306B;&#x5F71;&#x97FF;&#x3092;&#x4E0E;&#x3048;&#x308B;&#x53EF;&#x80FD;&#x6027;&#x304C;&#x3042;&#x308A;&#x307E;&#x3059;&#x3002;

&#x81EA;&#x5DF1;&#x7D44;&#x7E54;&#x5316;&#x9AD8;&#x5206;&#x5B50;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x306E;&#x7D30;&#x80DE;&#x63A5;&#x7740;&#x6750;&#x6599;&#x3092;&#x542B;&#x3080;&#x6ADB;&#x578B;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306E;&#x4F5C;&#x88FD;

Micropatterns &#xB9C8;&#xC774;&#xD06C;&#xB85C; &#xBC0F; &#xB098;&#xB178; &#xAE30;&#xB2A5; &#xBC0F; &#xB2E8;&#xC77C; &#xAE30;&#xD310;&#xC5D0; &#xC815;&#xC758; &#xB41C; &#xACF5;&#xAC04; &#xBC30;&#xCE58;&#xC640; &#xB2E4;&#xB978; &#xC0DD;&#xCCB4;&#xC758; &#xC138;&#xD3EC; &#xC218;&#xC900;&#xC758; &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC744; &#xACF5;&#xBD80; &#xD558;&#xB294; &#xB370; &#xC720;&#xC6A9;&#xD55C; &#xB3C4;&#xAD6C; &#xC774;&#xBA70; &#xCD5C;&#xADFC; &#xBCF4;&#xACE0;&#xC11C; &#xC140; &#xB3D9;&#xC791;&#xC744; &#xACB0;&#xC815;&#xD560; &#xB54C; &#xBE44; &#xACC4; &#xC900;&#xC218;&#xC758; &#xAC15;&#xD55C; &#xC601;&#xD5A5;&#xC744; &#xAC15;&#xC870; &#xD588;&#xB2E4;. &#xC774; &#xB17C;&#xBB38;&#xC5D0;&#xC11C;&#xB294; &#xB2E8;&#xC77C; &#xAE30;&#xD310;&#xC5D0; &#xB098;&#xB178; &#xB2E4;&#xCE35; &#xCF54;&#xD305;&#xC758; &#xC13C;&#xC11C; micropatterns&#xC758; &#xC81C;&#xC870;&#xC5D0; &#xB300; &#xD55C; &#xAC04;&#xB2E8;&#xD55C; &#xC544;&#xC9C1; &#xB2E4;&#xC591; &#xD558; &#xACE0; &#xC815;&#xBC00;&#xD55C; &#xD328;&#xD130; &#xB2DD; &#xAE30;&#xC220; &#xB9AC;&#xC18C; &#xADF8;&#xB798;&#xD53C;&#xC640; &#xD3F4;&#xB9AC;&#xBA38; &#xD45C;&#xBA74; &#xBBF8;&#xC138; (PSM) &#xB418; &#xB098; &#xB808;&#xC774;&#xC5B4;, &#xB808;&#xC774;&#xC5B4; (LbL) &#xC5B4;&#xC148;&#xBE14;&#xB9AC; &#xD504;&#xB85C;&#xC138;&#xC2A4;&#xC758; &#xACB0;&#xD569;&#xC744; &#xD1B5;&#xD574; &#xBCF4;&#xC5EC; &#xC90D;&#xB2C8;&#xB2E4;. LbL &#xB2E4;&#xCE35; &#xC5B4;&#xC148;&#xBE14;&#xB9AC; &#xBC0F; &#xC774;&#xB959;, &#xB9AC;&#xC18C; &#xADF8;&#xB798;&#xD53C;&#xB97C; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; &#xCCAB; &#xBC88;&#xC9F8; nanofilm &#xD328;&#xD134; &#xC0DD;&#xC131; &#xBC0F; &#xB3D9;&#xC77C;&#xD55C; &#xAE30;&#xD310;&#xC5D0; &#xD328;&#xD134;&#xC744; &#xC13C;&#xC11C;&#xB97C; &#xAD11;&#xD559; &#xC815;&#xB82C;&#xC744; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; &#xD504;&#xB85C;&#xC138;&#xC2A4;&#xAC00; &#xBC18;&#xBCF5; &#xB429;&#xB2C8;&#xB2E4;. &#xB530;&#xB77C;&#xC11C;,&#xB294; &#xBA54;&#xC11C;&#xB4DC;&#xB294; &#xC99D; &#xCC29; &#xC7AC;&#xB8CC;&#xB294; &#xACE0;&#xBD84;&#xC790;&#xC640; &#xB2E4;&#xCE35; &#xB098;&#xB178; &#xAD6C;&#xC870;&#xB97C; &#xC0DD;&#xC0B0; &#xD558;&#xB294; &#xC0DD;&#xBB3C; &#xD559;&#xC801; &#xC7AC;&#xB8CC;&#xC5D0; &#xD45C;&#xBA74; &#xBBF8;&#xC138; &#xC720;&#xC0AC; &#xD569;&#xB2C8;&#xB2E4;. &#xB2E4;&#xCE35;&#xC758; &#xC8FC;&#xC694; &#xAE30;&#xB2A5;&#xC740; &#xC7AC;&#xB8CC;, &#xC99D; &#xCC29; &#xC870;&#xAC74; &#xBC0F; postdeposition &#xCE58;&#xB8CC;&#xC758; &#xC801;&#xC808; &#xD55C; &#xC120;&#xD0DD;&#xC5D0; &#xC758;&#xD574; &#xAC15;&#xC131; &#xB4F1;&#xC758; &#xC18D;&#xC131;&#xC744; &#xC870;&#xC815; &#xD558;&#xB294; &#xAE30;&#xB2A5;&#xC785;&#xB2C8;&#xB2E4;. &#xC720;&#xB9AC; coverslips&#xC5D0; 2-4 &#xCEF4;&#xD3EC;&#xB10C;&#xD2B8; &#xC2DC;&#xC2A4;&#xD15C;&#xC744; &#xC815;&#xD655; &#xD558; &#xAC8C; &#xC815;&#xC758; &#xB41C;, &#xADE0;&#xC9C8; nanofilm &#xD328;&#xD134;&#xC744; &#xB9CC;&#xB4DC;&#xB294; &#xBC29;&#xBC95;&#xC758; &#xB2E4;&#xC591;&#xC131;&#xC744; &#xBCF4;&#xC5EC; &#xC8FC;&#xAE30; &#xC704;&#xD574; &#xC81C;&#xACF5; &#xB429;&#xB2C8;&#xB2E4;. &#xB610;&#xD55C;, &#xCCB4; &#xC678; &#xC138;&#xD3EC; &#xC811;&#xCC29; &#xBC0F; &#xC131;&#xC7A5;&#xC5D0; &#xB300; &#xD55C; &#xD14C;&#xC2A4;&#xD2B8; &#xBCA0;&#xB4DC;&#xB85C;&#xB85C; &#xC0AC;&#xC6A9; &#xB418;&#xB294; &#xBCF5;&#xC7A1; &#xD55C; &#xC2DC;&#xC2A4;&#xD15C;&#xC758; &#xC608;&#xB97C; &#xC81C;&#xACF5;: micropatterns poly(sodium 4-styrenesulfonate)/&#xD3F4; &#xB9AC;-L-&#xB9AC; &#xC2E0; hydrobromide (PSS/PLL) &#xBC0F; &#xBE44; phospholipase A(2)/poly(ethyleneimine) (sPLA(2)/PEI) &#xB2E4;&#xCE35;. &#xC13C;&#xC11C; &#xD3C9;&#xBC29; nanofilm &#xBC30;&#xC5F4; &#xD328;&#xD134; &#xC140; &#xAD6C; &#xCDA9; &#xC81C; &#xBC30;&#xACBD;&#xC73C;&#xB85C; poly(diallyldimethylammonium chloride) (PDDA)&#xC640; &#xB2E8;&#xC77C; coverslip&#xC5D0;&#xC11C; &#xC5BB;&#xC740; &#xD588;&#xB2E4;. &#xC138;&#xD3EC; &#xBC30;&#xC591; &#xC2E4;&#xD5D8; &#xB300;&#xB1CC; &#xD53C; &#xC9C8;&#xC758; &#xB274;&#xB7F0; &#xC751;&#xB2F5; PLL micropatterns&#xC640; &#xACBD;&#xC7C1;&#xC5D0;&#xC11C; sPLA(2) micropatterns &#xD2B9;&#xD788; &#xBC14;&#xC778;&#xB529;&#xD560; &#xBCF4;&#xC5EC;&#xC90D;&#xB2C8;&#xB2E4;. &#xC81C;&#xC791;&#xACFC; nanofilm micropatterns&#xC5D0;&#xC11C; &#xCD08;&#xAE30; &#xC0DD;&#xBB3C;&#xD559;&#xC801; &#xACB0;&#xACFC; &#xC911;&#xC694; &#xD55C; &#xC0DD;&#xBB3C; &#xD559;&#xC801; &#xAD6C;&#xC870;-&#xAE30;&#xB2A5; &#xAD00;&#xACC4;&#xB97C; elucidating &#xACA8;&#xB0E5; &#xD558;&#xB294; &#xC5F0;&#xAD6C;&#xC5D0; &#xC0AC;&#xC6A9; &#xD558;&#xAE30; &#xC704;&#xD574;&#xC774; &#xAE30;&#xBC95;&#xC758; &#xC720;&#xC6A9;&#xC131;&#xC744; &#xC9C0;&#xC6D0; &#xD558;&#xC9C0;&#xB9CC; &#xC81C;&#xC870; &#xBC29;&#xBC95;&#xC758; &#xC801;&#xC6A9; &#xAC00;&#xB2A5;&#xC131; &#xD6E8;&#xC52C; &#xAD11;&#xBC94;&#xC704; &#xD558; &#xACE0; &#xC804;&#xC790;, &#xD3EC;&#xD1A0;&#xB2C9;&#xC2A4;, &#xBC0F; &#xD654;&#xD559; &#xB9C8;&#xC774;&#xD06C;&#xB85C;&#xC2DC;&#xC2A4;&#xD15C;&#xC988;&#xC5D0; &#xC601;&#xD5A5;&#xC744; &#xC904; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;.

&#xC870;&#xB9BD;&#xB41C; &#xD3F4;&#xB9AC;&#xBA38; nanofilms &#xD3EC;&#xD568; &#xD558;&#xB294; &#xC138;&#xD3EC; &#xC811;&#xCC29; &#xC7AC;&#xB8CC;&#xC758; &#xC13C;&#xC11C; micropatterns&#xC758; &#xC81C;&#xC870;.

Micropatrones de diferentes biomateriales con caracter&iacute;sticas micro y nanoescala y arreglo espacial definido sobre un mismo sustrato son herramientas &uacute;tiles para el estudio de las interacciones del nivel celular, y los informes recientes han puesto de manifiesto la fuerte influencia del cumplimiento del andamio para determinar el comportamiento de la c&eacute;lula. En este papel, una t&eacute;cnica de dise&ntilde;o simple pero vers&aacute;til y precisa para la fabricaci&oacute;n de micropatrones interdigitados de recubrimientos multicapa de nanocompuestos sobre un mismo sustrato es demostrada a trav&eacute;s de una combinaci&oacute;n de litograf&iacute;a y procesos de ensamblaje (LbL) capa por capa, denominados micromecanizado superficial del pol&iacute;mero (PSM). El primer patr&oacute;n de nanofilm est&aacute; construido con la litograf&iacute;a, seguida por Asamblea multicapa LbL y despegue, y se repite el proceso con una alineaci&oacute;n &oacute;ptica para obtener patrones interdigitados en el mismo sustrato. As&iacute;, el m&eacute;todo es an&aacute;logo al micromecanizado superficial, salvo que los materiales de deposici&oacute;n son pol&iacute;meros y materiales biol&oacute;gicos que se utilizan para producir estructuras multicapa nanocompuestos. Una caracter&iacute;stica clave de las multicapas es la capacidad para ajustar propiedades como rigidez por la selecci&oacute;n adecuada de materiales, condiciones de deposici&oacute;n y tratamientos postdeposition. Y cuatro sistemas bicomponentes en cubreobjetos de vidrio se presentan para demostrar la versatilidad del enfoque para construir precisamente definida y homog&eacute;nea nanofilm patrones. Adem&aacute;s, se proporciona un ejemplo de un sistema complejo que se utiliza como un banco de pruebas para el crecimiento y la adhesi&oacute;n celular in vitro: micropatrones de poly(sodium 4-styrenesulfonate)/Poli-l-lisina bromhidrato (PSS/PLL) y secretada fosfolipasa A(2)/poly(ethyleneimine) (sPLA(2)/PEI) multicapas. Los patrones de la matriz de nanofilm cuadrados interdigitados se obtuvieron en un cubreobjetos solo con poly(diallyldimethylammonium chloride) (PDDA) como un fondo de celda-repelente. C&eacute;lula cultura experimentos muestran que las neuronas corticales responden y se unen espec&iacute;ficamente a la micropatrones sPLA(2) en competencia con PLL micropatrones. La fabricaci&oacute;n y los primeros resultados biol&oacute;gicos en la micropatrones nanofilm apoyan la utilidad de esta t&eacute;cnica para su uso en estudios tendientes a dilucidar las relaciones estructura-funci&oacute;n biol&oacute;gica importante, pero la aplicabilidad del m&eacute;todo de fabricaci&oacute;n es mucho m&aacute;s amplia y puede afectar microsystems qu&iacute;mica, electr&oacute;nica y fot&oacute;nica.

Fabricaci&oacute;n de micropatrones interdigitados de autoensambladas pol&iacute;mero nanofilms que contengan c&eacute;lulas adhesivas.

Metodi per la produzione di modelli di biomateriali con caratteristiche di distribuzione spaziale definito micro - e nano-scala sono importanti per lo studio delle interazioni cellulari-livello, comprese le comunicazioni cellula-materiale e alla cella base. Questo lavoro riporta sulla fabbricazione di substrati per studiare adesione cellulare a microdisegni multicomponenti di film multistrato di accoppiamento fotolitografia convenzionale e tecniche di LbL, conosciute come la tecnica L-LbL. Verso questa estremit&agrave;, substrati con microdisegni intercollegamento di due differenti bio-funzionalit&agrave; sono stati fabbricati per sPLA/sub 2 / e PLL e sono stati utilizzati per studi di coltura cellulare in vitro usando i neuroni, che hanno esibito preferenziale ad alta efficienza e adesione selettiva al sPLA/sub 2 / nanofilms. Questi risultati supportano l&#39;uso immediato di multicomponenti microdisegni come banchi di prova biologica per gli studi di base delle cellule e forniscano una base per ulteriori espansioni dei processi di fabbricazione per la produzione di ponteggi per definizione precisa delle interazioni cellula-materiale e cellula-cellula, tale che i costrutti risultanti imitano il comportamento e l&#39;organizzazione delle cellule in vivo.

Delle cellule di adesione test utilizzando nuovi banchi di prova contenente microdisegni di nanoingegneria complesso film multistrato.

M&eacute;thodes pour produire des mod&egrave;les de biomat&eacute;riau dot&eacute; de micro - et nano-scale distribution spatiale d&eacute;finie sont importants pour l&#39;&eacute;tude des interactions au niveau cellulaire, y compris la base de transmission de mat&eacute;riel-&agrave;-cellule et cellule-cellule. Ce travail rapporte sur la fabrication des substrats utilis&eacute;s pour &eacute;tudier l&#39;adh&eacute;sion cellulaire &agrave; multicomposants micromotifs de films multicouches en couplant la photolithographie classique et techniques LbL, connues comme &eacute;tant la technique de L-LbL. &Agrave; cette fin, des substrats avec nanofilm micromotifs de deux diff&eacute;rents bio-fonctionnalit&eacute;s ont &eacute;t&eacute; fabriqu&eacute;es pour Alps/sub 2 / et PLL et ont &eacute;t&eacute; utilis&eacute;s pour des &eacute;tudes de culture cellulaire in vitro &agrave; l&#39;aide de neurones, qui pr&eacute;sentaient pr&eacute;f&eacute;rentielle et une efficacit&eacute; &eacute;lev&eacute;es et adh&eacute;rence s&eacute;lective &agrave; la sPLA/sup 2 / nanofilms. Ces r&eacute;sultats appuient l&#39;utilisation imm&eacute;diate de plusieurs composants micropatterns comme bancs d&#39;essais biologiques pour des &eacute;tudes de base des cellules et la base d&#39;autres expansions des processus de fabrication pour produire des &eacute;chafaudages pour la d&eacute;finition pr&eacute;cise des interactions cellule-cellule et de cellule-&agrave;-mat&eacute;riel, tels que les constructions qui en r&eacute;sulte imitent le comportement et organisation cellulaire in vivo.

Cellule adh&eacute;rence test &agrave; l&#39;aide de nouveaux bancs d&#39;essai contenant micropatterns du complexe nano films multicouches.

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Methods for producing biomaterial patterns with defined spatial distribution micro- and nano-scale features are important for studying the cellular-level interactions, including basic cell-to-material and cell-to-cell communications. This work reports on the fabrication of substrates to study cell adhesion to multicomponent micropatterns of multilayer films by coupling conventional photolithography and LbL techniques, known as the L-LbL technique. Toward this end, substrates with nanofilm micropatterns of two different bio-functionalities have been fabricated for sPLA/sub 2/ and PLL and were used for in vitro cell-culture studies using neurons, which exhibited preferential and high efficiency and selective adhesion to sPLA/sub 2/ nanofilms. These results support the immediate use of multicomponent micropatterns as biological testbeds for basic studies of cells, and provide a basis for further expansions of the fabrication processes to produce scaffolds for precise definition of cell-to-material and cell-to-cell interactions, such that the resulting constructs mimic in vivo cell organization and behavior.

Cell adhesion testing using novel testbeds containing micropatterns of complex nanoengineered multilayer films.

Methoden zur Herstellung von Biomaterial Muster mit definierten r&auml;umlichen Verteilung Mikro- und Nano-Ma&szlig;stab Merkmale sind wichtig f&uuml;r die zellul&auml;re Ebene Interaktionen, einschlie&szlig;lich grundlegende Zelle-zu-Material und Zell-Zell-Kommunikation zu studieren. Diese Arbeit berichtet &uuml;ber die Herstellung von Substraten, Zelladh&auml;sion, Mehrkomponenten Micropatterns von Mehrschichtfolien zu studieren, durch die Kopplung von konventionellen Fotolithografie und LbL-Techniken, bekannt als die L-LbL-Technik. Dazu haben Substraten mit diesen Micropatterns von zwei verschiedenen Bio-Funktionalit&auml;ten f&uuml;r sPLA/Sub 2 fabriziert worden / und PLL und dienten f&uuml;r in-vitro Zellkultur-Studien mit Neuronen, die bevorzugte und hohe Effizienz und selektive Haftung sPLA/Sub 2 ausgestellt / Nanofilms. Diese Ergebnisse unterst&uuml;tzen von Mehrkomponenten-Micropatterns als biologische Testumgebungen eingesetzt f&uuml;r Grundstudium der Zellen und bilden die Grundlage f&uuml;r weitere Expansionen des der Fabrikation Produktionsprozesse f&uuml;r Ger&uuml;ste f&uuml;r genaue Definition der Zelle-zu-Material und Zell-Zell-Interaktionen, so dass die resultierende Konstrukte in-vivo-Zelle Organisation und Verhalten zu imitieren.

Zelle Adh&auml;sion mit Hilfe neuartiger Testbeds mit Micropatterns von komplexen Nanoengineered Mehrschichtfolien testen.

&#x5B9A;&#x7FA9;&#x6E08;&#x307F;&#x306E;&#x7A7A;&#x9593;&#x5206;&#x5E03;&#x306E;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30FB; &#x30CA;&#x30CE;&#x30B9;&#x30B1;&#x30FC;&#x30EB;&#x306E;&#x6A5F;&#x80FD;&#x3092;&#x6301;&#x3064;&#x751F;&#x4F53;&#x6750;&#x6599;&#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x4F5C;&#x308A;&#x51FA;&#x3059;&#x305F;&#x3081;&#x306E;&#x30E1;&#x30BD;&#x30C3;&#x30C9;&#x306F;&#x57FA;&#x672C;&#x7684;&#x306A;&#x7269;&#x8CEA;-&#x7D30;&#x80DE;&#x3068;&#x7D30;&#x80DE;&#x9593;&#x306E;&#x901A;&#x4FE1;&#x3092;&#x542B;&#x3080;&#x7D30;&#x80DE;&#x30EC;&#x30D9;&#x30EB;&#x306E;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x3092;&#x7814;&#x7A76;&#x3059;&#x308B;&#x305F;&#x3081;&#x91CD;&#x8981;&#x3067;&#x3059;&#x3002;&#x3053;&#x306E;&#x4F5C;&#x696D;&#x306F;&#x3001;&#x30D5;&#x30A9;&#x30C8;&#x30EA;&#x30BD;&#x30B0;&#x30E9;&#x30D5;&#x30A3;&#x30FC;&#x3068; L LbL &#x6280;&#x6CD5;&#x3068;&#x3057;&#x3066;&#x77E5;&#x3089;&#x308C;&#x3066;&#x3044;&#x308B;&#x3001;LbL &#x306E;&#x30C6;&#x30AF;&#x30CB;&#x30C3;&#x30AF;&#x3092;&#x7D50;&#x5408;&#x3059;&#x308B;&#x3053;&#x3068;&#x306B;&#x3088;&#x308A;&#x3001;&#x591A;&#x5C64;&#x819C;&#x306E;&#x591A;&#x6210;&#x5206;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306B;&#x5BFE;&#x3059;&#x308B;&#x7D30;&#x80DE;&#x63A5;&#x7740;&#x6027;&#x3092;&#x7814;&#x7A76;&#x3059;&#x308B;&#x57FA;&#x677F;&#x306E;&#x4F5C;&#x88FD;&#x3092;&#x5831;&#x544A;&#x3057;&#x307E;&#x3059;&#x3002;&#x3053;&#x306E;&#x7D42;&#x308F;&#x308A;&#x306B;&#x5411;&#x304B;&#x3063;&#x3066;&#x304C; sPLA/&#x30B5;&#x30D6; 2 &#x306E; 2 &#x3064;&#x306E;&#x7570;&#x306A;&#x308B;&#x30D0;&#x30A4;&#x30AA;&#x6A5F;&#x80FD;&#x30CA;&#x30CE;&#x8584;&#x819C;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x57FA;&#x677F;&#x3092;&#x4F5C;&#x88FD;&#x3055;&#x308C;&#x3066;/&#x304A;&#x3088;&#x3073; PLL &#x3068;&#x512A;&#x9047;&#x3068;&#x9AD8;&#x52B9;&#x7387;&#x3068; sPLA/&#x30B5;&#x30D6; 2 &#x3078;&#x306E;&#x9078;&#x629E;&#x7684;&#x63A5;&#x7740;&#x3092;&#x51FA;&#x5C55;&#x3057;&#x305F;&#x30CB;&#x30E5;&#x30FC;&#x30ED;&#x30F3;&#x3092;&#x7528;&#x3044;&#x305F; in vitro &#x7D30;&#x80DE;&#x57F9;&#x990A;&#x7814;&#x7A76;&#x306E;&#x305F;&#x3081;&#x4F7F;&#x7528;&#x3055;&#x308C;&#x305F;/&#x30CA;&#x30CE;&#x8584;&#x819C;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x7D50;&#x679C;&#x306F;&#x3001;&#x7D30;&#x80DE;&#x306E;&#x305F;&#x3081;&#x306E;&#x57FA;&#x790E;&#x7814;&#x7A76;&#x751F;&#x7269;&#x30C6;&#x30B9;&#x30C8;&#x30D9;&#x30C3;&#x30C9;&#x3068;&#x3057;&#x3066;&#x591A;&#x6210;&#x5206;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x306E;&#x5373;&#x6642;&#x4F7F;&#x7528;&#x3092;&#x30B5;&#x30DD;&#x30FC;&#x30C8;&#x3001;&#x7D50;&#x679C;&#x30B3;&#x30F3;&#x30B9;&#x30C8;&#x30E9;&#x30AF;&#x30C8; in vivo &#x306E;&#x7D30;&#x80DE;&#x7D44;&#x7E54;&#x3084;&#x884C;&#x52D5;&#x3092;&#x6A21;&#x5023;&#x3059;&#x308B;&#x3088;&#x3046;&#x306B;&#x62E1;&#x5927;&#x7269;&#x8CEA; &#xFF0D; &#x7D30;&#x80DE;&#x3068;&#x7D30;&#x80DE;&#x9593;&#x76F8;&#x4E92;&#x4F5C;&#x7528;&#x306E;&#x6B63;&#x78BA;&#x306A;&#x5B9A;&#x7FA9;&#x306E;&#x305F;&#x3081;&#x306E;&#x8DB3;&#x5834;&#x3092;&#x751F;&#x7523;&#x3059;&#x308B;&#x88FD;&#x9020;&#x30D7;&#x30ED;&#x30BB;&#x30B9;&#x306E;&#x305F;&#x3081;&#x306E;&#x57FA;&#x790E;&#x3092;&#x63D0;&#x4F9B;&#x3057;&#x307E;&#x3059;&#x3002;

&#x8907;&#x96D1;&#x306A;&#x30CA;&#x30CE;&#x30C6;&#x30AF;&#x30CE;&#x30ED;&#x30B8;&#x30FC;&#x6280;&#x8853;&#x306B;&#x3088;&#x308B;&#x591A;&#x5C64;&#x819C;&#x306E;&#x30DE;&#x30A4;&#x30AF;&#x30ED; &#x30D1;&#x30BF;&#x30FC;&#x30F3;&#x3092;&#x542B;&#x3080;&#x65B0;&#x898F;&#x306E;&#x30C6;&#x30B9;&#x30C8;&#x30D9;&#x30C3;&#x30C9;&#x3092;&#x7528;&#x3044;&#x305F;&#x4ED8;&#x7740;&#x6027;&#x8A66;&#x9A13;&#x3092;&#x30BB;&#x30EB;&#x3057;&#x307E;&#x3059;&#x3002;

&#xC815;&#xC758; &#xACF5;&#xAC04; &#xBD84;&#xD3EC; &#xB9C8;&#xC774;&#xD06C;&#xB85C; &#xBC0F; &#xB098;&#xB178; &#xC2A4;&#xCF00;&#xC77C; &#xAE30;&#xB2A5; biomaterial &#xD328;&#xD134; &#xC81C;&#xC791;&#xC744; &#xC704;&#xD55C; &#xBA54;&#xC11C;&#xB4DC;&#xB294; &#xAE30;&#xBCF8; &#xC140; &#xC18C;&#xC7AC; &#xBC0F; &#xC140; &#xD1B5;&#xC2E0;&#xC744; &#xD3EC;&#xD568; &#xD558; &#xC5EC; &#xC138;&#xD3EC; &#xC218;&#xC900;&#xC758; &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC744; &#xACF5;&#xBD80; &#xD558;&#xB294; &#xAC83;&#xC774; &#xC911;&#xC694; &#xD569;&#xB2C8;&#xB2E4;. &#xC774; &#xC791;&#xD488; &#xBCF4;&#xACE0; &#xAE30;&#xD310;&#xC758; &#xC81C;&#xC870; &#xAE30;&#xC874;&#xC758; &#xD3C9;&#xD310;&#xACFC; LbL &#xAE30;&#xBC95;, L LbL &#xAE30;&#xC220;&#xB85C; &#xC54C;&#xB824;&#xC9C4; &#xCEE4;&#xD50C;&#xB9C1; &#xD558; &#xC5EC; &#xB2E4;&#xCE35; &#xD544;&#xB984;&#xC758; &#xAE61;&#xD1B5;&#xC774; micropatterns &#xC138;&#xD3EC; &#xC811;&#xCC29; &#xB825;&#xC744; &#xACF5;&#xBD80; &#xD558;. &#xC774; &#xB05D;&#xC744; &#xD5A5;&#xD574; &#xB450; &#xAC1C;&#xC758; &#xC11C;&#xB85C; &#xB2E4;&#xB978; &#xBC14;&#xC774;&#xC624; &#xAE30;&#xB2A5; nanofilm micropatterns&#xC640; &#xD568;&#xAED8; &#xAE30;&#xD310; sPLA/&#xD558;&#xC704; 2&#xC5D0; &#xB300; &#xD55C; &#xC870;&#xC791; &#xB41C; /&#xC640; PLL &#xCCB4; &#xC678; &#xC138;&#xD3EC; &#xBC30;&#xC591; &#xC5F0;&#xAD6C; &#xC804;&#xC2DC; &#xD558; &#xACE0; &#xC6B0;&#xB300; &#xD558; &#xACE0; &#xB192;&#xC740; &#xD6A8;&#xC728;&#xACFC; sPLA/sub 2 &#xC120;&#xD0DD;&#xC801; &#xC811;&#xCC29; &#xC2E0;&#xACBD; &#xC138;&#xD3EC;&#xB97C; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; &#xC0AC;&#xC6A9; &#xD558; &#xACE0; / nanofilms. &#xC774;&#xB7EC;&#xD55C; &#xACB0;&#xACFC; &#xC140;&#xC758; &#xAE30;&#xBCF8; &#xC5F0;&#xAD6C;&#xC5D0; &#xB300; &#xD55C; &#xC0DD;&#xBB3C; &#xD559;&#xC801; testbeds &#xC989;&#xAC01;&#xC801;&#xC778; &#xD65C;&#xC6A9; &#xAE61;&#xD1B5;&#xC774; micropatterns &#xC9C0;&#xC6D0; &#xD558; &#xACE0; &#xACB0;&#xACFC; &#xAD6C;&#xBB38; &#xBAA8;&#xBC29; in vivo &#xC138;&#xD3EC; &#xC870;&#xC9C1; &#xBC0F; &#xB3D9;&#xC791; &#xB418;&#xB3C4;&#xB85D; &#xC140; &#xC7AC;&#xB8CC;&#xC640; &#xC138;&#xD3EC; &#xC138;&#xD3EC; &#xC0C1;&#xD638; &#xC791;&#xC6A9;&#xC758; &#xC815;&#xD655;&#xD55C; &#xC815;&#xC758; &#xB300; &#xD55C; &#xAC74;&#xC124; &#xAE30;&#xACC4; &#xC784;&#xB300; &#xC0DD;&#xC0B0; &#xC81C;&#xC870; &#xD504;&#xB85C;&#xC138;&#xC2A4;&#xC758; &#xCD94;&#xAC00; &#xD655;&#xC7A5;&#xC744; &#xC704;&#xD55C; &#xAE30;&#xCD08;&#xB97C; &#xC81C;&#xACF5;.

&#xC138;&#xD3EC; &#xC811;&#xCC29; &#xC18C;&#xC124; testbeds &#xD3EC;&#xD568; &#xD558;&#xB294; &#xBCF5;&#xC7A1; &#xD55C; nanoengineered &#xB2E4;&#xCE35; &#xD544;&#xB984;&#xC758; micropatterns&#xB97C; &#xC0AC;&#xC6A9; &#xD558; &#xC5EC; &#xD14C;&#xC2A4;&#xD2B8;.

M&eacute;todos para producir patrones de biomaterial con caracter&iacute;sticas de micro - y nano-escala de distribuci&oacute;n espacial definido son importantes para el estudio de las interacciones de nivel celular, incluyendo comunicaciones b&aacute;sicas-a-material de la c&eacute;lula y c&eacute;lula a c&eacute;lula. Este trabajo se informa sobre la fabricaci&oacute;n de sustratos para estudiar la adhesi&oacute;n celular a micropatrones multicomponente de pel&iacute;culas de m&uacute;ltiples capas por acoplamiento Fotolitograf&iacute;a convencional y t&eacute;cnicas LbL, conocidas como la t&eacute;cnica de L-LbL. Hacia ese fin, sustratos con micropatrones de nanofilm de dos diferentes bio-funcionalidades han sido fabricados para sPLA/sub 2 / y PLL y fueron utilizados para estudios de cultivo de c&eacute;lulas in vitro usando las neuronas, que exhibi&oacute; preferencial y de alta eficiencia y adhesi&oacute;n selectiva al sPLA/sub 2 / nanofilms. Estos resultados apoyan el uso inmediato de micropatrones multicomponente como demostradores biol&oacute;gica para estudios b&aacute;sicos de las c&eacute;lulas y proporcionan una base para otras expansiones de los procesos de fabricaci&oacute;n para producir andamios para definir con precisi&oacute;n las interacciones-a-material de la c&eacute;lula y c&eacute;lula a c&eacute;lula, tal que las construcciones que imitan el comportamiento y organizaci&oacute;n de c&eacute;lulas in vivo.

Pruebas de adherencia utilizando novela demostradores que contiene micropatrones de pel&iacute;culas de m&uacute;ltiples capas del complejo nanoengineered de la c&eacute;lula.

Physiological tissues, including brain and other organs, have three-dimensional (3-D) aspects that need to be supported to model them in vitro. Here we report the use of cellulose microfibers combined with cross-linked gelatin to make biocompatible porous microscaffolds for the sustained growth of brain cell and human mesenchymal stem cells (hMSCs) in 3-D structure. Live imaging using confocal microscopy indicated that 3-D microscaffolds composed of gelatin or cellulose fiber/gelatin both supported brain cell adhesion and growth for 16days in vitro. Cellulose microfiber/gelatin composites containing up to 75% cellulose fibers can withstand a higher mechanical load than gelatin alone, and composites also provided linear pathways along which brain cells could grow compared to more clumped cell growth in gelatin alone. Therefore, the bulk cellulose microfiber provides a novel skeleton in this new scaffold material. Cellulose fiber/gelatin scaffold supported hMSCs growth and extracellular matrix formation. hMSCs osteogenic and adipogenic assays indicated that hMSCs cultured in cellulose fiber/gelatin composite preserved the multilineage differentiation potential. As natural, biocompatible components, the combination of gelatin and cellulose microfibers, fabricated into 3-D matrices, may therefore provide optimal porosity and tensile strength for long-term maintenance and observation of cells.

Porous biocompatible three-dimensional scaffolds of cellulose microfiber/gelatin composites for cell culture.

Our previous studies show that group III secreted phospholipases A(2) (sPLA(2)s III) induces extensive neuronal apoptosis in brain cortical cultures. However, the molecular mechanisms underlying sPLA(2) III-induced neuronal injury/death are still unknown. Also it is not clear whether hypoxic pre-conditioning (HPC) is able to protect neurons from the sPLA(2) III insult. In this report, we demonstrate that sPLA(2) III significantly decreased production of Bcl-xl and the ratio of Bcl-xl/Bax, and increased expression of Bax, cleaved caspase 3, and cleaved alpha-Fodrin in primary neuronal culture. HPC prevented the sPLA(2) III-induced decreases in production of Bcl-xl and the ratio of Bcl-xl/Bax, and increases in expression of Bax, cleaved caspase 3, and alpha-Fodrin. However, the HPC-produced neuronal protection was eliminated or attenuated by AG490, rapamycin, and STAT3 shRNA. Our results suggest that sPLA(2) III-induced neuronal apoptosis is likely because of its alterations in expression and activity of Bcl-xl, Bax, caspase 3, and its target gene fodrin; and that HPC-produced neuroprotection against the sPLA(2) III toxicity is mediated via JAK-STAT signal pathways that regulate the expression of Bcl-xl, Bax, and cleaved caspase 3 in cultured cortical neurons.

Hypoxic preconditioning suppresses group III secreted phospholipase A2-induced apoptosis via JAK2-STAT3 activation in cortical neurons.

The hedgehog signal pathway plays a crucial role in the angiogenesis and vascular remodeling. However, the function of this pathway in the pulmonary vascular smooth cell proliferation in response to hypoxia remains unknown. In this study, we have demonstrated that the main components of the hedgehog pathway, including sonic hedgehog (SHH), patched1 (PTCH1), smoothened (SMO), GLI and hypoxia-inducible factor 1 (HIF1) are expressed in the human pulmonary arterial smooth muscle cells (HPASMCs). Interestingly, hypoxia significantly enhanced the expression of SHH and HIF1, facilitated the translocation of GLI1 into the nuclei, and promoted the proliferation of HPASMCs. Furthermore, direct activation of the SHH pathway through incubation with the purified recombinant human SHH or with purmorphamine and SAG, two Smo agonists, also enhanced the proliferation of HPASMCs. Importantly, the treatment with anti-SHH and anti-HIF1 antibodies or cyclopamine, a specific SMO inhibitor, markedly inhibited the nuclear translocation of GLI1 and cell proliferation in the HPASMCs induced by hypoxia and activation of the SHH pathway. Moreover, the treatment with cyclopamine increased apoptosis in the hypoxic HPASMCs. These data strongly demonstrate for the first time that the SHH signaling plays a crucial role in the regulation of HPASMC growth in response to hypoxia.

Activation of the sonic hedgehog signaling controls human pulmonary arterial smooth muscle cell proliferation in response to hypoxia.

Secreted phospholipase A₂-IIA (sPLA₂-IIA) is an inducible enzyme released under several inflammatory conditions. It has been shown that sPLA₂-IIA is released from rat brain astrocytes after inflammatory stimulus, and lipopolysaccharide (LPS) and nitric oxide (NO) have been implicated in regulation of this release. Here, brain microvascular endothelial cells (BMVECs) were treated with LPS to uncover whether sPLA₂-IIA was released, whether nitric oxide regulated this release, and any related signal mechanisms.

Article	Year
Secreted phospholipase A2 potentiates glutamate-induced calcium increase and cell death in primary neuronal cultures. Journal of neuroscience research\| PubMed ID: 11891776	2002
Glutamate signalling and secretory phospholipase A2 modulate the release of arachidonic acid from neuronal membranes. Journal of neuroscience research\| PubMed ID: 12111845	2002
Group III secreted phospholipase A2 causes apoptosis in rat primary cortical neuronal cultures. Brain research\| PubMed ID: 14519523	2003
Interplay among platelet-activating factor, oxidative stress, and group I metabotropic glutamate receptors modulates neuronal survival. Journal of neuroscience research\| PubMed ID: 15264222	2004
Quantification of sPLA2-induced early and late apoptosis changes in neuronal cell cultures using combined TUNEL and DAPI staining. Brain research. Brain research protocols\| PubMed ID: 15296851	2004
Role of nitric oxide in regulating secreted phospholipase A2 release from astrocytes. Neuroreport\| PubMed ID: 16056137	2005
Fabrication of interdigitated micropatterns of self-assembled polymer nanofilms containing cell-adhesive materials. Langmuir : the ACS journal of surfaces and colloids\| PubMed ID: 16519477	2006
Cell adhesion testing using novel testbeds containing micropatterns of complex nanoengineered multilayer films. Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference\| PubMed ID: 17270826	2004
Porous biocompatible three-dimensional scaffolds of cellulose microfiber/gelatin composites for cell culture. Acta biomaterialia\| PubMed ID: 20035906	2010
Hypoxic preconditioning suppresses group III secreted phospholipase A2-induced apoptosis via JAK2-STAT3 activation in cortical neurons. Journal of neurochemistry\| PubMed ID: 20492356	2010
Activation of the sonic hedgehog signaling controls human pulmonary arterial smooth muscle cell proliferation in response to hypoxia. Biochimica et biophysica acta\| PubMed ID: 20840857	2010
Regulatory effects of the JAK3/STAT1 pathway on the release of secreted phospholipase A₂-IIA in microvascular endothelial cells of the injured brain. Journal of neuroinflammation\| PubMed ID: 22788969	2012

Mark A. DeCoster

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