RESUMEN
Understanding cells' response to the macroscopic and nanoscale properties of biomaterials requires studies in model systems with the possibility to tailor their mechanical properties and different length scales. Here, we describe an interpenetrating network (IPN) design based on a stiff PEGDA host network interlaced within a soft 4-arm PEG-Maleimide/thiol (guest) network. We quantify the nano- and bulk mechanical behavior of the IPN and the single network hydrogels by single-molecule force spectroscopy and rheological measurements. The IPN presents different mechanical cues at the molecular scale, depending on which network is linked to the probe, but the same mechanical properties at the macroscopic length scale as the individual host network. Cells attached to the interpenetrating (guest) network of the IPN or to the single network (SN) PEGDA hydrogel modified with RGD adhesive ligands showed comparable attachment and spreading areas, but cells attached to the guest network of the IPN, with lower molecular stiffness, showed a larger number and size of focal adhesion complexes and a higher concentration of the Hippo pathway effector Yes-associated protein (YAP) than cells linked to the PEGDA single network. The observations indicate that cell adhesion to the IPN hydrogel through the network with lower molecular stiffness proceeds effectively as if a higher ligand density is offered. We claim that IPNs can be used to decipher how changes in ECM design and connectivity at the local scale affect the fate of cells cultured on biomaterials.
RESUMEN
Microbial resistant coatings have raised considerable interest in the biotechnological industry and clinical scenarios to combat the spreading of infections, in particular in implanted medical devices. Polymer brushes covalently attached to surfaces represent a useful platform to identify ideal compositions for preventing bacterial settlement by quantifying bacteria-surface interactions. In this work, a series of polymer brushes with different charges, positively charged poly[2-(methacryloyloxy)ethyl trimethylammonium chloride] (PMETAC), negatively charged poly(3-sulfopropyl methacrylate potassium salt) (PSPMA), and neutral poly(2-hydroxyethyl methacrylate) (PHEMA) were grafted onto glass surfaces by surface-initiated atom transfer radical polymerization in aqueous conditions. The antimicrobial activity of the polymer brushes against Gram-negative Escherichia coli was tested at the nano- and microscopic level on different time scales, that is, from nm to 100 µm, and ms to 24 h, respectively. The interaction between the polymer brushes and E. coli was studied by single-cell force spectroscopy (SCFS) and by quantification of the bacterial density on surfaces incubated with bacterial suspensions. E. coli firmly attached to positive PMETAC brushes with high work required for de-adhesion of 28 ± 9 nN·nm, but did not significantly bind to negatively charged PSPMA and neutral PHEMA brushes. Our studies of bacterial adhesion using polymer brushes with controllable chemistry provide essential insights into bacterial surface interactions and the origins of bacterial adhesion.
Asunto(s)
Antibacterianos/química , Adhesión Bacteriana , Escherichia coli/crecimiento & desarrollo , Polihidroxietil Metacrilato , Metacrilatos/química , Polihidroxietil Metacrilato/síntesis química , Polihidroxietil Metacrilato/química , Polimerizacion , Propiedades de SuperficieRESUMEN
Engineering of biomaterials with specific biological properties has gained momentum as a means to control stem cell behavior. Here, we address the effect of bifunctionalized hydrogels comprising polylysine (PL) and a 19-mer peptide containing the laminin motif IKVAV (IKVAV) on embryonic and adult neuronal progenitor cells under different stiffness regimes. Neuronal differentiation of embryonic and adult neural progenitors was accelerated by adjusting the gel stiffness to 2 kPa and 20 kPa, respectively. While gels containing IKVAV or PL alone failed to support long-term cell adhesion, in bifunctional gels, IKVAV synergized with PL to promote differentiation and formation of focal adhesions containing ß1-integrin in embryonic cortical neurons. Furthermore, in adult neural stem cell culture, bifunctionalized gels promoted neurogenesis via the expansion of neurogenic clones. These data highlight the potential of synthetic matrices to steer stem and progenitor cell behavior via defined mechano-adhesive properties.
Asunto(s)
Hidrogeles/farmacología , Laminina/química , Células-Madre Neurales/citología , Neurogénesis , Fragmentos de Péptidos/química , Animales , Células Cultivadas , Elasticidad , Adhesiones Focales/metabolismo , Hidrogeles/química , Laminina/farmacología , Ratones , Ratones Endogámicos C57BL , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/efectos de los fármacos , Células-Madre Neurales/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Polilisina/química , Polilisina/farmacología , Ingeniería de Tejidos/métodosRESUMEN
The contact geometry of microstructured adhesive surfaces is of high relevance for adhesion enhancement. Theoretical considerations indicate that the stress distribution in the contact zone is crucial for the detachment mechanism, but direct experimental evidence is missing so far. In this work, we propose a method that allows, for the first time, the detection of local stresses at the contact area of biomimetic adhesive microstructures during contact formation, compression and detachment. We use a mechano-sensitive polymeric layer, which turns mechanical stresses into changes of fluorescence intensity. The biomimetic surface is brought into contact with this layer in a well-defined fashion using a microcontact printer, while the contact area is monitored with fluorescence microscopy in situ. Thus, changes in stress distribution across the contact area during compression and pull-off can be visualized with a lateral resolution of 1 µm. We apply this method to study the enhanced adhesive performance of T-shaped micropillars, compared to flat punch microstructures. We find significant differences in the stress distribution of the both differing contact geometries during pull-off. In particular, we find direct evidence for the suppression of crack nucleation at the edge of T-shaped pillars, which confirms theoretical models for the superior adhesive properties of these structures.
RESUMEN
Two photoremovable protecting groups, namely, nitroveratryloxycarbonyl (NVo) and diethylamino-coumarin-4-yl (DEACM), have been tested for wavelength-selective, independent removal. The chromophores were attached to the amine group of aminopropyltriethoxysilane and used for the modification of silica surfaces. A photolytic experiment on the photosensitive layers allowed us to identify the irradiation conditions for the selective cleavage of the chromophores. UV measurements revealed that the photolabile DEACM group can be cleaved off with UV light at 412 nm without damaging the NVo group. The NVo group could then be removed at 365 nm. Masked irradiation of substrates modified with a 1:1 molar mixture of both silanes allowed the generation of bifunctional patterns after the selective cleavage of DEACM and NVo in a sequential irradiation process. The deprotection reaction was confirmed by coupling two different fluorescent dyes to the liberated amine groups. The expected two-color pattern could be observed by fluorescence microscopy.
Asunto(s)
Luz , Fotoquímica/métodos , Química Física/métodos , Diseño de Equipo , Fluorescencia , Colorantes Fluorescentes/farmacología , Microscopía Fluorescente/métodos , Modelos Químicos , Silanos/química , Espectrofotometría Ultravioleta/métodos , Propiedades de Superficie , Rayos UltravioletaRESUMEN
Inspired by biological attachment systems, micropatterned elastomeric surfaces with pillars of different heights (between 2.5 and 80 microm) and radii (between 2.5 and 25 microm) were fabricated. Their adhesion properties were systematically tested and compared with flat controls. Micropatterned surfaces with aspect ratios above 0.5 were found to be more compliant than flat surfaces. The adhesion significantly increases with decreasing pillar radius and increasing aspect ratio of the patterned features. A preload dependence of the adhesion force has been identified and demonstrated to be crucial for obtaining adhesives with tunable adherence.