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1.
Biophys Rev ; 15(4): 497-513, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37681084

RESUMO

Cellular systems must deal with mechanical forces to satisfy their physiological functions. In this context, proteins with mechanosensitive properties play a crucial role in sensing and responding to environmental changes. The discovery of aquaporins (AQPs) marked a significant breakthrough in the study of water transport. Their transport capacity and regulation features make them key players in cellular processes. To date, few AQPs have been reported to be mechanosensitive. Like mechanosensitive ion channels, AQPs respond to tension changes in the same range. However, unlike ion channels, the aquaporin's transport rate decreases as tension increases, and the molecular features of the mechanism are unknown. Nevertheless, some clues from mechanosensitive ion channels shed light on the AQP-membrane interaction. The GxxxG motif may play a critical role in the water permeation process associated with structural features in AQPs. Consequently, a possible gating mechanism triggered by membrane tension changes would involve a conformational change in the cytoplasmic extreme of the single file region of the water pathway, where glycine and histidine residues from loop B play a key role. In view of their transport capacity and their involvement in relevant processes related to mechanical forces, mechanosensitive AQPs are a fundamental piece of the puzzle for understanding cellular responses.

2.
Polymers (Basel) ; 15(3)2023 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-36771837

RESUMO

Biodegradable polymers have been strongly recognized as an alternative to replace traditional petrochemical plastics, which have become a global problem due to their long persistence in the environment. In this work, the effect of the addition of titanium dioxide nanoparticles (TiO2NP) on the morphology, physicochemical properties and biodegradation under industrial composting conditions of cassava starch-based nanocomposites obtained by extrusion at different screw speeds (80 and 120 rpm) were investigated. Films performed at 120 rpm (S120 and S120-TiO2NP) showed completely processed starch and homogeneously distributed nanoparticles, leading to much more flexible nanocomposites than those obtained at 80 rpm. The incorporation of TiO2NP led to an increase in storage modulus of all films and, in the case of S120-TiO2NP, to higher strain at break values. From the Kohlrausch-Williams-Watts theoretical model (KWW), an increase in the relaxation time of the nanocomposites was observed due to a decrease in the number of polymer chains involved in the relaxation process. Additionally, S120-TiO2NP showed effective protection against UV light, greater hydrophobicity and faster biodegradation in compost, resulting in a promising material for food packaging applications.

3.
Biochim Biophys Acta Mol Cell Res ; 1869(4): 119209, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34999138

RESUMO

In Saccharomyces cerevisiae cAMP regulates different cellular processes through PKA. The specificity of the response of the cAMP-PKA pathway is highly regulated. Here we address the mechanism through which the cAMP-PKA pathway mediates its response to heat shock and thermal adaptation in yeast. PKA holoenzyme is composed of a regulatory subunit dimer (Bcy1) and two catalytic subunits (Tpk1, Tpk2, or Tpk3). PKA subunits are differentially expressed under certain growth conditions. Here we demonstrate the increased abundance and half-life of TPK1 mRNA and the assembly of this mRNA in cytoplasmic foci during heat shock at 37 °C. The resistance of the foci to cycloheximide-induced disassembly along with the polysome profiling analysis suggest that TPK1 mRNA is impaired for entry into translation. TPK1 expression was also evaluated during a recurrent heat shock and thermal adaptation. Tpk1 protein level is significantly increased during the recovery periods. The crosstalk of cAMP-PKA pathway and CWI signalling was also studied. Wsc3 sensor and some components of the CWI pathway are necessary for the TPK1 expression upon heat shock. The assembly in foci upon thermal stress depends on Wsc3. Tpk1 expression is lower in a wsc3∆ mutant than in WT strain during thermal adaptation and thus the PKA levels are also lower. An increase in Tpk1 abundance in the PKA holoenzyme in response to heat shock is presented, suggesting that a recurrent stress enhanced the fitness for the coming favourable conditions. Therefore, the regulation of TPK1 expression by thermal stress contributes to the specificity of cAMP-PKA signalling.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/genética , Meia-Vida , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Polirribossomos/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , RNA Mensageiro/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Temperatura
4.
PLoS One ; 16(5): e0251411, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33974655

RESUMO

Cells exert traction forces on the extracellular matrix to which they are adhered through the formation of focal adhesions. Spatial-temporal regulation of traction forces is crucial in cell adhesion, migration, cellular division, and remodeling of the extracellular matrix. By cultivating cells on polyacrylamide hydrogels of different stiffness we were able to investigate the effects of substrate stiffness on the generation of cellular traction forces by Traction Force Microscopy (TFM), and characterize the molecular dynamics of the focal adhesion protein zyxin by Fluorescence Correlation Spectroscopy (FCS) and Fluorescence Recovery After Photobleaching (FRAP). As the rigidity of the substrate increases, we observed an increment of both, cellular traction generation and zyxin residence time at the focal adhesions, while its diffusion would not be altered. Moreover, we found a positive correlation between the traction forces exerted by cells and the residence time of zyxin at the substrate elasticities studied. We found that this correlation persists at the subcellular level, even if there is no variation in substrate stiffness, revealing that focal adhesions that exert greater traction present longer residence time for zyxin, i.e., zyxin protein has less probability to dissociate from the focal adhesion.


Assuntos
Estresse Mecânico , Zixina/química , Citoesqueleto de Actina/efeitos dos fármacos , Amidas/farmacologia , Animais , Bovinos , Adesão Celular , Citocalasina D/farmacologia , Células Endoteliais , Recuperação de Fluorescência Após Fotodegradação , Adesões Focais , Proteínas de Fluorescência Verde , Microscopia Intravital , Cinética , Lasers , Camundongos , Camundongos Endogâmicos BALB C , Piridinas/farmacologia , Proteínas Recombinantes de Fusão/química , Vinculina/química , Quinases Associadas a rho/antagonistas & inibidores
5.
Sci Rep ; 10(1): 20258, 2020 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-33219264

RESUMO

Parkinson's disease (PD) is a neurodegenerative disorder for which only symptomatic treatments are available. Repurposing drugs that target α-synuclein aggregation, considered one of the main drivers of PD progression, could accelerate the development of disease-modifying therapies. In this work, we focused on chemically modified tetracycline 3 (CMT-3), a derivative with reduced antibiotic activity that crosses the blood-brain barrier and is pharmacologically safe. We found that CMT-3 inhibited α-synuclein amyloid aggregation and led to the formation of non-toxic molecular species, unlike minocycline. Furthermore, CMT-3 disassembled preformed α-synuclein amyloid fibrils into smaller fragments that were unable to seed in subsequent aggregation reactions. Most interestingly, disaggregated species were non-toxic and less inflammogenic on brain microglial cells. Finally, we modelled the interactions between CMT-3 and α-synuclein aggregates by molecular simulations. In this way, we propose a mechanism for fibril disassembly. Our results place CMT-3 as a potential disease modifier for PD and possibly other synucleinopathies.


Assuntos
Inflamação/induzido quimicamente , Tetraciclinas/farmacologia , alfa-Sinucleína/toxicidade , Reposicionamento de Medicamentos , Humanos , Doença de Parkinson/tratamento farmacológico , Agregados Proteicos , Tetraciclinas/uso terapêutico , alfa-Sinucleína/metabolismo
6.
FASEB J ; 34(1): 735-753, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31914594

RESUMO

Upon overnutrition, adipocytes activate a homeostatic program to adjust anabolic pressure. An inflammatory response enables adipose tissue (AT) expansion with concomitant enlargement of its capillary network, and reduces energy storage by increasing insulin resistance. Galectin-12 (Gal-12), an endogenous lectin preferentially expressed in AT, plays a key role in adipocyte differentiation, lipolysis, and glucose homeostasis. Here, we reveal biochemical and biophysical determinants of Gal-12 structure, including its preferential recognition of 3-fucosylated structures, a unique feature among members of the galectin family. Furthermore, we identify a previously unanticipated role for this lectin in the regulation of angiogenesis within AT. Gal-12 showed preferential localization within the inner side of lipid droplets, and its expression was upregulated under hypoxic conditions. Through glycosylation-dependent binding to endothelial cells, Gal-12 promoted in vitro angiogenesis. Moreover, analysis of in vivo AT vasculature showed reduced vascular networks in Gal-12-deficient (Lgals12-/-) compared to wild-type mice, supporting a role for this lectin in AT angiogenesis. In conclusion, this study unveils biochemical, topological, and functional features of a hypoxia-regulated galectin in AT, which modulates endothelial cell function through recognition of 3-fucosylated glycans. Thus, glycosylation-dependent programs may control AT homeostasis by modulating endothelial cell biology with critical implications in metabolic disorders and inflammation.


Assuntos
Adipócitos/metabolismo , Células Endoteliais/metabolismo , Galectinas/metabolismo , Neovascularização Patológica/metabolismo , Tecido Adiposo/metabolismo , Animais , Fenômenos Fisiológicos Celulares/fisiologia , Resistência à Insulina/fisiologia , Gotículas Lipídicas/metabolismo , Lipólise/fisiologia , Camundongos Knockout , Polissacarídeos/metabolismo
7.
Phys Chem Chem Phys ; 21(41): 22947-22954, 2019 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-31598616

RESUMO

Layer by layer assembly of polyelectrolytes with proteins is a convenient tool for the development of functional biomaterials. Most of the studies presented in the literature are based on the electrostatic interaction between components of opposite charges, limiting the assembly possibilities. However, this process can be tuned by modifying the environment where the main constituents are dissolved. In this work, the electron transfer behavior between an electroactive polyelectrolyte (polyallylamine derivatized with an osmium complex) and a redox enzyme (glucose oxidase) is studied by assembling them in the presence of phosphate ions at different ionic strengths. Our results show that the environment from which the assembly is constructed has a significant effect on the electrochemical response. Notably, the polyelectrolyte dissolved in the presence of phosphate at high ionic strength presents a globular structure which is preserved after adsorption with substantial effects on the buildup of the multilayer system, improving the electron transfer process through the film.

8.
Polymers (Basel) ; 11(2)2019 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-30960274

RESUMO

The tunable swelling and mechanical properties of nanostructures polymers are crucial parameters for the creation of adaptive devices to be used in diverse fields, such as drug delivery, nanomedicine, and tissue engineering. We present the use of anodic aluminum oxide templates as a nanoreactor to copolymerize butyl methacrylate and 2-hydroxyethyl acrylate under radical conditions. The copolymer obtained under confinement showed significant differences with respect to the same copolymer obtained in bulk conditions. Molecular weights, molecular weight dispersities, Young's modulus, and wetting behaviors were significantly modified. The combination of selected monomers allowed us to obtain nanopillar structures with an interesting softening surface and extraordinary swelling capacity that could be of special interest to surface science and specifically, cell culture.

9.
Soft Matter ; 15(7): 1640-1650, 2019 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-30676599

RESUMO

Supramolecular self-assembly of molecular building blocks represents a powerful "nanoarchitectonic" tool to create new functional materials with molecular-level feature control. Here, we propose a simple method to create tunable phosphate/polyamine-based films on surfaces by successive assembly of poly(allylamine hydrochloride) (PAH)/phosphate anions (Pi) supramolecular networks. The growth of the films showed a great linearity and regularity with the number of steps. The coating thickness can be easily modulated by the bulk concentration of PAH and the deposition cycles. The PAH/Pi networks showed chemical stability between pH 4 and 10. The transport properties of the surface assemblies formed from different deposition cycles were evaluated electrochemically by using different redox probes in aqueous solution. The results revealed that either highly permeable films or efficient anion transport selectivity can be created by simply varying the concentration of PAH. This experimental evidence indicates that this new strategy of supramolecular self-assembly can be useful for the rational construction of single polyelectrolyte nanoarchitectures with multiple functionalities.

10.
Biomater Sci ; 6(8): 2230-2247, 2018 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-29978861

RESUMO

The modulation of cell adhesion via biologically inspired materials plays a key role in the development of realistic platforms to envisage not only mechanistic descriptions of many physiological and pathological processes but also new biointerfacial designs compatible with the requirements of biomedical devices. In this work, we show that the cell adhesion and proliferation of three different cell lines can be easily manipulated by using a novel biologically inspired supramolecular coating generated via dip coating of the working substrates in an aqueous solution of polyallylamine in the presence of phosphate anions-a simple one-step modification procedure. Our results reveal that selective cell adhesion can be controlled by varying the deposition time of the coating. Cell proliferation experiments showed a cell type-dependent quasi-exponential growth demonstrating the nontoxic properties of the supramolecular platform. After reaching a certain surface coverage, the supramolecular films based on phosphate-polyamine networks displayed antiadhesive activity towards cells, irrespective of the cell type. However and most interestingly, these antiadherent substrates developed strong adhesive properties after thermal annealing at 37 °C for 3 days. These results were interpreted based on the changes in the coating hydrophilicity, topography and stiffness, with the latter being assessed by atomic force microscopy imaging and indentation experiments. The reported approach is simple, robust and flexible, and would offer opportunities for the development of tunable, biocompatible interfacial architectures to control cell attachment for various biomedical applications.


Assuntos
Materiais Biocompatíveis/química , Substâncias Macromoleculares/química , Fosfatos/química , Poliaminas/química , Células 3T3 , Absorção Fisiológica , Animais , Materiais Biocompatíveis/síntese química , Adesão Celular , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Células HeLa , Humanos , Cinética , Substâncias Macromoleculares/síntese química , Camundongos , Microscopia de Força Atômica , Tamanho da Partícula , Molhabilidade
11.
Sci Rep ; 8(1): 9788, 2018 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-29955093

RESUMO

Mechanical stimuli play a key role in many cell functions such as proliferation, differentiation and migration. In the mammary gland, mechanical signals such as the distension of mammary epithelial cells due to udder filling are proposed to be directly involved during lactation and involution. However, the evolution of focal adhesions -specialized multiprotein complexes that mechanically connect cells with the extracellular matrix- during the mammary gland development, as well as the influence of the mechanical stimuli involved, remains unclear. Here we present the use of an equibiaxial stretching device for exerting a sustained normal strain to mammary epithelial cells while quantitatively assessing cell responses by fluorescence imaging techniques. Using this approach, we explored changes in focal adhesion dynamics in HC11 mammary cells in response to a mechanical sustained stress, which resembles the physiological stimuli. We studied the relationship between a global stress and focal adhesion assembly/disassembly, observing an enhanced persistency of focal adhesions under strain as well as an increase in their size. At a molecular level, we evaluated the mechanoresponses of vinculin and zyxin, two focal adhesion proteins postulated as mechanosensors, observing an increment in vinculin molecular tension and a slower zyxin dynamics while increasing the applied normal strain.


Assuntos
Células Epiteliais/metabolismo , Adesões Focais/metabolismo , Imageamento Tridimensional , Glândulas Mamárias Animais/citologia , Mecanotransdução Celular , Estresse Mecânico , Animais , Sobrevivência Celular , Feminino , Fluorescência , Recuperação de Fluorescência Após Fotodegradação , Cinética , Camundongos Endogâmicos BALB C , Fibras de Estresse/metabolismo , Vinculina/metabolismo , Zixina/metabolismo
12.
J Colloid Interface Sci ; 518: 92-101, 2018 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-29448230

RESUMO

We present the covalent modification of a Pani-like conducting polymer (polyaminobenzylamine, PABA) by grafting of a polyelectrolyte brush (poly [2-(methacryloyloxy)-ethyl-trimethylammonium chloride], PMETAC). As PABA has extra pendant amino moieties, the grafting procedure does not affect the backbone nitrogen atoms that are implicated in the electronic structure of the conducting polymers. Moreover, perchlorate anions interact very strongly with the quaternary ammonium pendant groups of PMETAC through ion pairing. Therefore, the grafting does not only keep the electroactivity of PABA in aqueous solutions but it adds the ion-actuation properties of the PMETAC brush to the modified electrode as demonstrated by contact angle measurements and electrochemical methods. In this way, the conjugation of the electron transfer properties of the conducting polymer with the anion responsiveness of the integrated brush renders perchlorate actuation of the electrochemical response. These results constitute a rational integration of nanometer-sized polymer building blocks that yields synergism of functionalities and illustrate the potentialities of nanoarchitectonics for pushing the limits of soft material science into the nanoworld.

13.
Soft Matter ; 14(10): 1939-1952, 2018 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-29479625

RESUMO

Supramolecular self-assembly is of paramount importance for the development of novel functional materials with molecular-level feature control. In particular, the interest in creating well-defined stratified multilayers through simple methods using readily available building blocks is motivated by a multitude of research activities in the field of "nanoarchitectonics" as well as evolving technological applications. Herein, we report on the facile preparation and application of highly organized stacked multilayers via layer-by-layer assembly of lipid-like surfactants and polyelectrolytes. Polyelectrolyte multilayers with high degree of stratification of the internal structure were constructed through consecutive assembly of polyallylamine and dodecyl phosphate, a lipid-like surfactant that act as a structure-directing agent. We show that multilayers form well-defined lamellar hydrophilic/hydrophobic domains oriented parallel to the substrate. More important, X-ray reflectivity characterization conclusively revealed the presence of Bragg peaks up to fourth order, evidencing the highly stratified structure of the multilayer. Additionally, hydrophobic lamellar domains were used as hosts for ferrocene in order to create an electrochemically active film displaying spatially-addressed redox units. Stacked multilayers were then assembled integrating redox-tagged polyallylamine and glucose oxidase into the stratified hydrophilic domains. Bioelectrocatalysis and "redox wiring" in the presence of glucose was demonstrated to occur inside the stratified multilayer.

14.
Langmuir ; 34(12): 3711-3719, 2018 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-29480725

RESUMO

Responsive interfacial architectures of practical interest commonly require the combination of conflicting properties in terms of their demand upon material structure. Switchable stiffness, wettability, and permeability, key features for tissue engineering applications, are in fact known to be exclusively interdependent. Here, we present a nanoarchitectonic approach that decouples these divergent properties by the use of thermoresponsive microgels as building blocks for the construction of three-dimensional arrays of interconnected pores. Layer-by-layer assembled poly( N-isopropylacrylamide- co-methacrylic acid) microgel films were found to exhibit an increase in hydrophobicity, stiffness, and adhesion properties upon switching the temperature from below to above the lower critical solution temperature, whereas the permeability of redox probes through the film remained unchanged. Our findings indicate that the switch in hydrophilicity and nanomechanical properties undergone by the microgels does not compromise the porosity of the film, thus allowing the free diffusion of redox probes through the polymer-free volume of the submicrometer pores. This novel approach for decoupling conflicting properties provides a strategic route for creating tailorable scaffolds with unforeseen functionalities.

15.
Nanoscale ; 10(3): 1189-1195, 2018 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-29271438

RESUMO

The possibility of combining more than one stimulus-responsive property into a single material holds interesting potential for the creation of adaptive devices to be used in diverse fields such as drug delivery, nanomedicine and tissue engineering. This paper describes a novel material based on thermo-responsive PNIPAm nanopillars with amplified surface properties through the incorporation of Fe3O4 nanoparticles. The incorporation of magnetic nanoparticles into the nanopillars, prepared via surface-initiated atom-transfer radical polymerization in anodized aluminum oxide templates, sharply increased their stiffness and hydrophobicity when increasing the temperature above the volume phase transition temperature. Furthermore, their magnetic response turned out to be proportional to the amount of the incorporated nanoparticles. The possibility of sharply increasing the stiffness with a temperature variation close to the human body temperature paves the way to the application of these substrates as "smart" scaffolds for cell culture. Additionally, the presence of superparamagnetic nanoparticles in the nanopillars offers the possibility of using these nanostructured systems for magnetic hyperthermia.

16.
Plant Reprod ; 31(2): 159-169, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29236154

RESUMO

KEY MESSAGE: In vitro tomato pollen tubes show a cytoplasmic calcium gradient that oscillates with the same period as growth. Pollen tube growth requires coordination between the tip-focused cytoplasmic calcium concentration ([Ca2+]cyt) gradient and the actin cytoskeleton. This [Ca2+]cyt gradient is necessary for exocytosis of small vesicles, which contributes to the delivery of new membrane and cell wall at the pollen tube tip. The mechanisms that generate and maintain this [Ca2+]cyt gradient are not completely understood. Here, we studied calcium dynamics in tomato (Solanum lycopersicum) pollen tubes using transgenic tomato plants expressing the Yellow Cameleon 3.6 gene under the pollen-specific promoter LAT52. We use tomato as an experimental model because tomato is a Solanaceous plant that is easy to transform, and has an excellent genomic database and genetic stock center, and unlike Arabidopsis, tomato pollen is a good system to do biochemistry. We found that tomato pollen tubes showed an oscillating tip-focused [Ca2+]cyt gradient with the same period as growth. Then, we used a pharmacological approach to disturb the intracellular Ca2+ homeostasis, evaluating how the [Ca2+]cyt gradient, pollen germination and in vitro pollen tube growth were affected. We found that cyclopiazonic acid (CPA), a drug that inhibits plant PIIA-type Ca2+-ATPases, increased [Ca2+]cyt in the subapical zone, leading to the disappearance of the Ca2+ oscillations and inhibition of pollen tube growth. In contrast, 2-aminoethoxydiphenyl borate (2-APB), an inhibitor of Ca2+ released from the endoplasmic reticulum to the cytoplasm in animals cells, completely reduced [Ca2+]cyt at the tip of the tube, blocked the gradient and arrested pollen tube growth. Although both drugs have antagonistic effects on [Ca2+]cyt, both inhibited pollen tube growth triggering the disappearance of the [Ca2+]cyt gradient. When CPA and 2-APB were combined, their individual inhibitory effects on pollen tube growth were partially compensated. Finally, we found that GsMTx-4, a peptide from spider venom that blocks stretch-activated Ca2+ channels, inhibited tomato pollen germination and had a heterogeneous effect on pollen tube growth, suggesting that these channels are also involved in the maintenance of the [Ca2+]cyt gradient. All these results indicate that tomato pollen tube is an excellent model to study calcium dynamics.


Assuntos
ATPases Transportadoras de Cálcio/antagonistas & inibidores , Cálcio/metabolismo , Calmodulina/metabolismo , Proteínas Luminescentes/metabolismo , Tubo Polínico/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Solanum lycopersicum/metabolismo , Compostos de Boro/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio , Citoplasma/metabolismo , Retículo Endoplasmático/metabolismo , Indóis/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular , Solanum lycopersicum/efeitos dos fármacos , Solanum lycopersicum/crescimento & desenvolvimento , Peptídeos/farmacologia , Proteínas de Plantas/antagonistas & inibidores , Tubo Polínico/efeitos dos fármacos , Tubo Polínico/crescimento & desenvolvimento , Venenos de Aranha/farmacologia
17.
Soft Matter ; 13(13): 2453-2464, 2017 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-28287232

RESUMO

The surface properties of soft nanostructured hydrogels are crucial in the design of responsive materials that can be used as platforms to create adaptive devices. The lower critical solution temperature (LCST) of thermo-responsive hydrogels such as poly(N-isopropylacrylamide) (PNIPAm) can be modified by introducing a hydrophilic monomer to create a wide range of thermo-responsive micro-/nano-structures in a large temperature range. Using surface initiation atom-transfer radical polymerization in synthesized anodized aluminum oxide templates, we designed, fabricated, and characterized thermo-responsive nanopillars based on PNIPAm hydrogels with tunable mechanical properties by incorporating acrylamide monomers (AAm). In addition to their LCST, the incorporation of a hydrophilic entity in the nanopillars based on PNIPAm has abruptly changed the topological and mechanical properties of our system. To gain an insight into the mechanical properties of the nanostructure, its hydrophilic/hydrophobic behavior and topological characteristics, atomic force microscopy, molecular dynamics simulations and water contact angle studies were combined. When changing the nanopillar composition, a significant and opposite variation was observed in their mechanical properties. As temperature increased above the LCST, the stiffness of PNIPAm nanopillars, as expected, did so too, in contrast to the stiffness of PNIPAm-AAm nanopillars that decreased significantly. The molecular dynamics simulations proposed a local molecular rearrangement in our nanosystems at the LCST. The local aggregation of NIPAm segments near the center of the nanopillars displaced the hydrophilic AAm units towards the surface of the structure leading to contact with the aqueous environment. This behavior was confirmed via contact angle measurements below and above the LCST.

18.
Rev Sci Instrum ; 88(1): 013703, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28147641

RESUMO

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.


Assuntos
Adesões Focais , Mecanotransdução Celular , Proteínas/química , Fenômenos Fisiológicos Celulares
19.
Plant Direct ; 1(4): e00012, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31245661

RESUMO

Many of mRNAs synthesized during pollen development are translated after germination, and we hypothesize that they are stored in cytoplasmic granules. We analyzed the cellular localization of the SKS14 and AT59 Arabidopsis mRNAs, which are orthologues of the tobacco NTP303 and tomato LAT59 pollen mRNAs, respectively, by artificially labeling the transcripts with a MS2-GFP chimera. A MATLAB-automated image analysis helped to identify the presence of cytoplasmic SKS14 and AT59 mRNA granules in mature pollen grains. These mRNA granules partially colocalized with VCS and DCP1, two processing body (PB) proteins. Finally, we found a temporal correlation between SKS14 protein accumulation and the disappearance of SKS14 mRNA granules during pollen germination. These results contribute to unveil a mechanism for translational regulation in Arabidopsis thaliana pollen.

20.
ACS Appl Mater Interfaces ; 9(1): 930-943, 2017 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-28004915

RESUMO

Hierarchical assembly of hard/soft nanoparticles holds great potential as reinforcements for polymer nanocomposites with tailored properties. Here, we present a facile strategy to integrate polystyrene-grafted carbon nanotubes (PSgCNT) (0.05-0.3 wt %) and poly(styrene-b-[isoprene-ran-epoxyisoprene]-b-styrene) block copolymer (10 wt %) into epoxy coatings using an ultrasound-assisted noncovalent functionalization process. The method leads to cured nanocomposites with core-shell block copolymer (BCP) nanodomains which are associated with carbon nanotubes (CNT) giving rise to CNT-BCP hybrid structures. Nanocomposite energy dissipation and reduced Young's Modulus (E*) is determined from force-distance curves by atomic force microscopy operating in the PeakForce QNM imaging mode and compared to thermosets modified with BCP and purified carbon nanotubes (pCNT). Remarkably, nanocomposites bearing PSgCNT-BCP conjugates display an increase in energy dissipation of up to 7.1-fold with respect to neat epoxy and 53% more than materials prepared with pCNT and BCP at the same CNT load (0.3 wt %), while reduced Young's Modulus shows no significant change with CNT type and increases up to 25% compared to neat epoxy E* at a CNT load of 0.3 wt %. The energy dissipation performance of nanocomposites is also reflected by the lower wear coefficients of materials with PSgCNT and BCP compared to those with pCNT and BCP, as determined by abrasion tests. Furthermore, scanning electron microscopy (SEM) images taken on wear surfaces show that materials incorporating PSgCNT and BCP exhibit much more surface deformation under shear forces in agreement with their higher ability to dissipate more energy before particle release. We propose that the synergistic effect observed in energy dissipation arises from hierarchical assembly of PSgCNT and BCP within the epoxy matrix and provides clues that the CNT-BCP interface has a significant role in the mechanisms of energy dissipation of epoxy coating modified by CNT-BCP conjugates. These findings provide a means to design epoxy-based coatings with high-energy dissipation performance.

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