Joshua S. Katz

Light-responsive biomaterials: development and applications.

Novel biomaterials are beneficial to the growing fields of drug delivery, cell biology, micro-devices, and tissue engineering. With recent advances in chemistry and materials science, light is becoming an attractive option as a method to control biomaterial behavior and properties. In this Feature Article, we explore some of the early and recent advances in the design of light-responsive biomaterials. Particular attention is paid to macromolecular assemblies for drug delivery, multi-component surface patterning for advanced cell assays, and polymer networks that undergo chemical or shape changes upon light exposure. We conclude with some remarks about future directions of the field.

Sequential crosslinking to control cellular spreading in 3-dimensional hydrogels

With advanced understanding of how manipulations in material chemistry and structure influence cellular interactions, material control over cellular behavior (e.g., spreading) is becoming increasingly possible. In this example, we developed a novel process that utilizes different crosslinking mechanisms to provide gel environments that are either permissive or inhibitory to cellular spreading. To accomplish this, a multi-acrylated macromer (i.e., acrylated hyaluronic acid) was first crosslinked with an addition reaction using a matrix metalloprotease (MMP) cleavable peptide containing thiol groups. When an adhesive peptide was also coupled to the network, this environment permitted the spreading of encapsulated human mesenchymal stem cells (hMSCs), whereas control systems did not. If all acrylates were not consumed during the initial crosslinking step, a photoinitiated radical polymerization could be used to crosslink the remaining acrylates and inhibit cellular spreading with the production of covalent barriers. Variations in the ratio of the two crosslink types in individual constructs controlled the degradation and mechanical properties of the hydrogels, as well as the degree of spreading of encapsulated cells. Cell spreading was further controlled spatially with the use of photomasks. Overall, this new technology is an exciting and potentially valuable tool, both to provide new insights into the relationships between gel structure and cell behavior, and for eventual tissue-engineering applications where spatial control over cells is desired.

Modular synthesis of biodegradable diblock copolymers for designing functional polymersomes.

Polymer vesicles, or polymersomes, are promising candidates for applications in drug delivery and tissue imaging. While a vast variety of polymers have been explored for their ability to assemble into polymersomes, relatively little research on the functionalization of these polymers has been reported. We present here a novel route for the synthesis of poly(caprolactone)-b-poly(ethylene glycol) (PCL-b-PEG) diblock copolymers that allows for the insertion of functional groups at the block junctions and the assembly of functional membranes. This modular synthesis has been developed on the basis of solid-phase peptide synthesis techniques and is accomplished through the formation of two peptide bonds, one between an amine-terminated PEG and the carboxyl moiety of the functional group and the other between the functional group amine and a carboxy-terminated PCL. As a demonstration of the potential utility of the resulting vesicles, we incorporated two different amino acid functional groups at the junction. 2-Nitrophenylalanine was utilized to create UV-responsive membranes in which the vesicles were destabilized and released encapsulated contents upon irradiation. A fluorescein-conjugated lysine was also utilized to create stable fluorescent membranes in which the fluorescence was built into the polymer. This method should contribute to our ability to further develop smart, functional membranes.

Addressable micropatterning of multiple proteins and cells by microscope projection photolithography based on a protein friendly photoresist.

We report a new method for the micropatterning of multiple proteins and cells with micrometer-scale precision. Microscope projection photolithography based on a new protein-friendly photoresist, poly(2,2-dimethoxy nitrobenzyl methacrylate-r-methyl methacrylate-r-poly(ethylene glycol) methacrylate) (PDMP), was used for the fabrication of multicomponent protein/cell arrays. Microscope projection lithography allows precise registration between multiple patterns as well as facile fabrication of microscale features. Thin films of PDMP became soluble in near-neutral physiological buffer solutions upon UV exposure and exhibited excellent resistance to protein adsorption and cell adhesion. By harnessing advantages in microscope projection photolithography and properties of PDMP thin films, we could successfully fabricate protein arrays composed of multiple proteins. Furthermore, we could extend this method for the patterning of two different types of immune cells for the potential study of immune cell interactions. This technique will in general be useful for protein chip fabrication and high-throughput cell-cell communication study.

Hydrogel mediated delivery of trophic factors for neural repair.

Neurotrophins have been implicated in a variety of diseases and their delivery to sites of disease and injury has therapeutic potential in applications including spinal cord injury, Alzheimers disease, and Parkinsons disease. Biodegradable polymers, and specifically, biodegradable water-swollen hydrogels, may be advantageous as delivery vehicles for neurotrophins because of tissue-like properties, tailorability with respect to degradation and release behavior, and a history of biocompatibility. These materials may be designed to degrade via hydrolytic or enzymatic mechanisms and can be used for the sustained delivery of trophic factors in vivo. Hydrogels investigated to date include purely synthetic to purely natural, depending on the application and intended release profiles. Also, flexibility in material processing has allowed for the investigation of injectable materials, the development of scaffolding and porous conduits, and the use of composites for tailored molecule delivery profiles. It is the objective of this review to describe what has been accomplished in this area thus far and to remark on potential future directions in this field. Ultimately, the goal is to engineer optimal biomaterials to deliver molecules in a controlled and dictated manner that can promote regeneration and healing for numerous neural applications.

Membrane stabilization of biodegradable polymersomes.

Biodegradable polymersomes are promising vehicles for a range of applications. Their stabilization would improve many properties, including the retention and controlled release of polymersome contents, yet this has not been previously accomplished. Here, we present the first example of stabilizing fully biodegradable polymersomes through acrylation of the hydrophobic terminal end of polymersome-forming poly(caprolactone-b-ethylene glycol). Exposure of the resulting polymersomes loaded with a hydrophobic photoinitiator to ultraviolet light polymerized the acrylates, without affecting polymersome morphology or cell cytotoxicity. These stabilized polymersomes were more resistant to surfactant disruption and degradation. As an example of stabilized polymersome utility, the unintended release of doxorubicin (DOX) due to leakage from polymersomes decreased with membrane stabilization and slower sustained release was observed. Finally, DOX-loaded polymersomes retained their cytotoxicity following stabilization.

Photocleavable side groups to spatially alter hydrogel properties and cellular interactions

Hydrogel systems with inducible variations in mechanical and chemical properties are of interest to many aspects of tissue engineering. Substrate-tethered hydrogel films, fabricated by copolymerization of hydroxyethylacrylate with the photolabile monomer 2-nitrobenzyl acrylate (2-NBA) and a crosslinker were shown to be responsive to UV light through cleavage of the 2-NBA moiety. The surface and bulk properties of the films were characterized by infrared spectroscopy and atomic force and confocal microscopy. At long irradiation times, a tripling of the swelling ratio and an order of magnitude decrease in gel modulus were observed, coupled with an increase in surface wettability. Following short exposures, the gels became resistant to protein and cell adhesion, a trend that was reversed with longer exposure times. Finally, high-fidelity patterns of gel swelling were fabricated through spatially selective irradiation by employing basic photomasks. These materials are useful for future studies for which spatial and temporal control of material properties and cellular interactions are desirable.

Effects of membrane rheology on leuko-polymersome adhesion to inflammatory ligands

A strategy for treating inflammatory disease is to create micro-particles with the adhesive properties of leukocytes. The underlying rheology of deformable adhesive microspheres would be an important factor in the adhesive performance of such particles. In this work the effect of particle deformability on the selectin-mediated rolling of polymer vesicles (polymersomes) is evaluated. The rheology of the polymersome membrane was modulated by cross-linking unsaturated side-chains within the hydrophobic core of the membrane. Increased membrane rigidity resulted in decreased rates of particle recruitment rather than decreased average rolling velocities. Reflective interference contrast microscopy of rolling vesicles confirmed that neither flaccid nor rigid vesicles sustained close contacts with the substrate during rolling adhesion. A variable-shear rate parallel-plate flow chamber was employed to evaluate individual vesicles rolling on substrates under different flow conditions. Analysis of the trajectories of single flaccid vesicles revealed several distinct populations of rolling vesicles; however, some of these populations disappear when the vesicle membranes are made rigid. This work shows that membrane mechanics affects the capture, but not the rolling dynamics, of adherent leuko-polymersomes.

Biodegradable Polymersomes for the Delivery of Gemcitabine to Panc-1 Cells

Traditional anticancer chemotherapy often displays toxic side effects, poor bioavailability, and a low therapeutic index. Targeting and controlled release of a chemotherapeutic agent can increase drug bioavailability, mitigate undesirable side effects, and increase the therapeutic index. Here we report a polymersome-based system to deliver gemcitabine to Panc-1 cells in vitro. The polymersomes were self-assembled from a biocompatible and completely biodegradable polymer, poly(ethylene oxide)-poly(caprolactone), PEO-PCL. We showed that we can encapsulate gemcitabine within stable 200 nm vesicles with a 10% loading efficiency. These vesicles displayed a controlled release of gemcitabine with 60% release after 2 days at physiological pH. Upon treatment of Panc-1 cells in vitro, vesicles were internalized as verified with fluorescently labeled polymersomes. Clonogenic assays to determine cell survival were performed by treating Panc-1 cells with varying concentrations of unencapsulated gemcitabine (FreeGem) and polymersome-encapsulated gemcitabine (PolyGem) for 48 hours. 1 μM PolyGem was equivalent in tumor cell toxicity to 1 μM FreeGem, with a one log cell kill observed. These studies suggest that further investigation on polymersome-based drug formulations is warranted for chemotherapy of pancreatic cancer.