Porous Nanofilm Biomaterials Via Templated Layer‐by‐Layer Assembly

Abstract

Hydrogel‐like biomaterials are often too soft to support robust cell adhesion, yet methods to increase mechanical rigidity (e.g., covalent cross‐linking the gel matrix) can compromise bioactivity by suppressing the accessibility or activity of embedded biomolecules. Nanoparticle templating is reported here as a strategy toward porous, layer‐by‐layer assembled, thin polyelectrolyte films of sufficient mechanical rigidity to promote strong initial cell adhesion, and that are capable of high bioactive species loading. Latex nanoparticles are incorporated during layer‐by‐layer assembly, and following 1‐ethyl‐3‐[3‐dimethylaminopropyl]carbodiimide/N‐hydroxysulfosuccinimide (EDC‐NHS) cross‐linking of the polyelectrolyte film, are removed via exposure to tetrahydrofuran (THF). THF exposure results in only a partial reduction in film thickness (as observed by ellipsometry), suggesting the presence of internal pore space. The attachment, spreading, and metabolic activity of pre‐osteoblastic MC3T3‐E1 cells cultured on templated, cross‐linked films are statistically similar to those on non‐templated films, and much greater than those on non‐cross‐linked films. Laser scanning confocal microscopy and quartz crystal microgravimetry indicate a high capacity for bioactive species loading (ca. 10% of film mass) in nanoparticle templated films. Porous nanofilm biomaterials, formed via layer‐by‐layer assembly with nanoparticle templating, promote robust cell adhesion and exhibit high bioactive species loading, and thus appear to be excellent candidates for cell‐contacting applications.

Nanotemplated polyelectrolyte films as porous biomolecular delivery systems

“…Prior work shows THF treatment of native films (i.e., without crosslinking) containing latex nanoparticles to result in significant film collapse, suggesting removal of the nanoparticles from the film 20 . In contrast, cross-linked films did not exhibit such a collapse, suggesting in this case the particle removal to be accompanied by formation of pore space 20 .…”

“…The penultimate layer of these films contains a fluorescently labeled polycation (PLL-FITC). PLL-FITC (as well as unlabeled PLL) has been previously shown to freely diffuse throughout the film, 25 and so the entire film appears green 20 . When subjected to red labeled BMP-2, both films appear mainly yellow, and the solution above red.…”

“…Prior work shows THF treatment of native films (i.e., without crosslinking) containing latex nanoparticles to result in significant film collapse, suggesting removal of the nanoparticles from the film 20 . In contrast, cross-linked films did not exhibit such a collapse, suggesting in this case the particle removal to be accompanied by formation of pore space 20 . UV spectra of THF used to dissolve the nanoparticles from the film exhibit peaks that are also found in a latex sample, suggesting the presence of latex in the dissolving solution and hence nanoparticle removal from the film (see Supplemental Materials).…”

“…We previously developed a cross-linked, nanoparticle templated film where the porosity is introduced after the dissolution of carboxyl latex nanoparticles introduced during film assembly 20 . We showed these films to be sufficiently rigid so as to promote initial cell adhesion, spreading and viability, and to be capable of significant biomolecular species loading.…”

“…Both films are subjected to chemical cross-linking with 40 mM 1-ethyl-3-(3 [dimethylamino]propyl)carbodiimide (EDC, Sigma) and 100 mM, N -hydroxysulfosuccinimide (sulfo-NHS, Sigma) for 16h. The films containing the carboxyl functionalized latex nanoparticles (NP, Invitrogen) are incubated in tetrahydrofurane (THF) for 16h in order to remove the NP from the films and rinsed three times for 20 min in buffer, as previously described 20 . Native (PLL-PGA) n films are formed as described above, but without chemical EDC-NHS cross-linking.…”