Photocrosslinkable Hyaluronic Acid Research Articles

Tumor environment differentiated “nanodepot” programmed for site-specific drug shuttling and combinative therapy on metastatic cancer

Metastatic cancer is difficult to defeat with current treatments due to lack of etiological therapeutics and efficient delivery platforms. Employing tumor microenvironment in programming intelligent nanosystems has attracted considerable attention for combinative antitumor therapy. Herein, we proposed a core-shell based drug depot consisting of micellar core and crosslinked-gel shell for site-specific shuttling of paclitaxel (PTX) and KIAA1199 specific shRNA (shKIAA). Poly (e-caprolactone) were grafted with branched polyethylenimine (PEI-PCL) as micellar core, into which hydrophobic PTX was embedded; while shKIAA, a reliable RNAi regimen for metastatic cell inhibition was condensed with PEI through electrostatic interaction; and then photo-crosslinked hyaluronic acid (m-HA) was further coated as shell. The nanoscale drug depot shared HAase-triggered charge switching and desirable release profile. Upon reaching tumor region, HA shell was degraded by concentrated HAase, and facilitated drug shuttling to individual subcellular targeting site. Rapid intracellular trafficking of micellar core achieved endo/lysosomal escape and cytoplasmic liberation. The half-maximal inhibitory concentration (IC50) of “nanodepot” toward human breast cancer cell line MDA-MB-231 was 0.016 μg/mL (PTX concentration), approximately 3-fold decrease compared to that of monotherapy group (0.043 μg/mL). The tumor weight inhibition (TWI) is 83.30% in xenografted MDA-MB-231 tumor model and metastasis was effectively inhibited in 4T1 orthotopic tumors. Moreover, knockdown of KIAA1199 via sustainable RNAi affected a broad range of cellular functions including apoptosis, migration and invasion. Collectively, tumor environment differentiated spatiotemporal co-delivery fashion holds a great promise for combinative treatment with enhanced efficacy on metastatic cancer cases.

Osteochondral Regeneration Induced by TGF-β Loaded Photo Cross-Linked Hyaluronic Acid Hydrogel Infiltrated in Fused Deposition-Manufactured Composite Scaffold of Hydroxyapatite and Poly (Ethylene Glycol)-Block-Poly(ε-Caprolactone).

The aim of this study was to report the fabrication of porous scaffolds with pre-designed internal pores using a fused deposition modeling (FDM) method. Polycaprolactone (PCL) is a suitable material for the FDM method due to the fact it can be melted and has adequate flexural modulus and strength to be formed into a filament. In our study, the filaments of methoxy poly(ethylene glycol)-block-poly(ε-caprolactone) having terminal groups of carboxylic acid were deposited layer by layer. Raw materials having a weight ratio of hydroxyapatite (HAp) to polymer of 1:2 was used for FDM. To promote cell adhesion, amino groups of the Arg-Gly-Asp(RGD) peptide were condensed with the carboxylic groups on the surface of the fabricated scaffold. Then the scaffold was infiltrated with hydrogel of glycidyl methacrylate hyaluronic acid loading with 10 ng/mL of TGF-β1 and photo cross-linked on the top of the scaffolds. Serious tests of mechanical and biological properties were performed in vitro. HAp was found to significantly increase the compressive strength of the porous scaffolds. Among three orientations of the filaments, the lay down pattern 0°/90° scaffolds exhibited the highest compressive strength. Fluorescent staining of the cytoskeleton found that the osteoblast-like cells and stem cells well spread on RGD-modified PEG-PCL film indicating a favorable surface for the proliferation of cells. An in vivo test was performed on rabbit knee. The histological sections indicated that the bone and cartilage defects produced in the knees were fully healed 12 weeks after the implantation of the TGF-β1 loaded hydrogel and scaffolds, and regenerated cartilage was hyaline cartilage as indicated by alcian blue and periodic acid-schiff double staining.

Photo-crosslinked hyaluronic acid coated upconverting nanoparticles

Hyaluronic acid (HA)-coated inorganic nanoparticles display enhanced interaction with the CD44 receptors which are overexpressed in many types of cancer cells. Here, we describe a modification of core-shell β-NaY0.80Yb0.18Er0.02F4@NaYF4 nanoparticles (UCNP) by HA derivative bearing photo-reactive groups. UCNP capped with oleic acid were firstly transferred to aqueous phase by an improved protocol using hydrochloric acid or lactic acid treatment. Subsequently, HA bearing furanacryloyl moieties (HA-FU) was adsorbed on the nanoparticle surface and crosslinked by UV irradiation. The crosslinking resulted in stable HA coating, and no polymer desorption was observed. As-prepared UCNP@HA-FU show a hydrodynamic diameter of about 180 nm and are colloidally stable in water and cell culture media. The cellular uptake by normal human fibroblasts and MDA MB-231 cancer cell line was investigated by upconversion luminescence imaging.

Photo-crosslinked HAMA hydrogel with cordycepin encapsulated chitosan microspheres for osteoarthritis treatment.

Autophagy is a protective mechanism in normal cartilage. The present study aimed to investigate the synergistic therapeutic effect of promotion of chondrocyte autophagy via exposure to cordycepin encapsulated by chitosan microspheres (CM-cordycepin) and photo-crosslinked hyaluronic acid methacrylate (HAMA) hydrogel, with the goal of evaluating CM-cordycepin as a treatment for patients with osteoarthritis. First, we developed and evaluated the characteristics of HAMA hydrogels and chitosan microspheres. Next, we measured the effect of cordycepin on cartilage matrix degradation induced by IL1-β in chondrocytes and an ex vivo model. Cordycepin protects cartilage from degradation partly by activation of autophagy. Moreover, we surgically induced osteoarthritis in mice, which were injected intra-articularly with CM-cordycepin and HAMA. The combination of CM-cordycepin and HAMA hydrogel retarded the progression of surgically induced OA. Cordycepin ameliorated cartilage matrix degradation at least partially by inducing autophagy in vivo. Our results demonstrate that the combination of cordycepin encapsulated by CMs and photo-crosslinked HAMA hydrogel could be a promising strategy for treating patients with osteoarthritis.

Injectable photocrosslinkable hyaluronic acid hydrogels incorporated with platelet lysate enhance the dentinogenic differentiation of human dental pulp stem cells

Injectable photocrosslinkable hyaluronic acid hydrogels incorporated with platelet lysate enhance the dentinogenic differentiation of human dental pulp stem cells

Effects of permeability and living space on cell fate and neo-tissue development in hydrogel-based scaffolds: a study with cartilaginous model.

One bottleneck in tissue regeneration with hydrogel scaffolds is the limited understanding of the crucial factors for controlling hydrogel's physical microenvironments to regulate cell fate. Here, the effects of permeability and living space of hydrogels on encapsulated cells' behavior were evaluated, respectively. Three model hydrogel-based constructs are fabricated by using photo-crosslinkable hyaluronic acid as precursor and chondrocytes as model cell type. The better permeable hydrogels facilitate better cell viability and rapid proliferation, which lead to increased production of extracellular matrix (ECM), e.g. collagen, glycosaminoglycan. By prolonged culture, nano-sized hydrogel networks inhibit neo-tissue development, and the presence of macro-porous living spaces significantly enhance ECM deposition via forming larger cell clusters and eventually induce formation of scaffold-free neo-tissue islets. The results of this work demonstrate that the manipulation and optimization of hydrogel microenvironments, namely permeability and living space, are crucial to direct cell fate and neo-tissue formation.

Phenotypic stability, matrix elaboration and functional maturation of nucleus pulposus cells encapsulated in photocrosslinkable hyaluronic acid hydrogels

Degradation of the nucleus pulposus (NP) is an early hallmark of intervertebral disc degeneration. The capacity for endogenous regeneration in the NP is limited due to the low cellularity and poor nutrient and vascular supply. Towards restoring the NP, a number of biomaterials have been explored for cell delivery. These materials must support the NP cell phenotype while promoting the elaboration of an NP-like extracellular matrix in the shortest possible time. Our previous work with chondrocytes and mesenchymal stem cells demonstrated that hydrogels based on hyaluronic acid (HA) are effective at promoting matrix production and the development of functional material properties. However, this material has not been evaluated in the context of NP cells. Therefore, to test this material for NP regeneration, bovine NP cells were encapsulated in 1%w/vol HA hydrogels at either a low seeding density (20×106cellsml−1) or a high seeding density (60×106cellsml−1), and constructs were cultured over an 8week period. These NP cell-laden HA hydrogels showed functional matrix accumulation, with increasing matrix content and mechanical properties with time in culture at both seeding densities. Furthermore, encapsulated cells showed NP-specific gene expression profiles that were significantly higher than expanded NP cells prior to encapsulation, suggesting a restoration of phenotype. Interestingly, these levels were higher at the lower seeding density compared to the higher seeding density. These findings support the use of HA-based hydrogels for NP tissue engineering and cellular therapies directed at restoration or replacement of the endogenous NP.

Functional properties of bone marrow-derived MSC-based engineered cartilage are unstable with very long-term in vitro culture

The success of stem cell-based cartilage repair requires that the regenerate tissue reach a stable state. To investigate the long-term stability of tissue engineered cartilage constructs, we assessed the development of compressive mechanical properties of chondrocyte and mesenchymal stem cell (MSC)-laden three dimensional agarose constructs cultured in a well defined chondrogenic in vitro environment through 112 days. Consistent with previous reports, in the presence of TGF-β, chondrocytes outperformed MSCs through day 56, under both free swelling and dynamic culture conditions, with MSC-laden constructs reaching a plateau in mechanical properties between days 28 and 56. Extending cultures through day 112 revealed that MSCs did not simply experience a lag in chondrogenesis, but rather that construct mechanical properties never matched those of chondrocyte-laden constructs. After 56 days, MSC-laden constructs underwent a marked reversal in their growth trajectory, with significant declines in glycosaminoglycan content and mechanical properties. Quantification of viability showed marked differences in cell health between chondrocytes and MSCs throughout the culture period, with MSC-laden construct cell viability falling to very low levels at these extended time points. These results were not dependent on the material environment, as similar findings were observed in a photocrosslinkable hyaluronic acid (HA) hydrogel system that is highly supportive of MSC chondrogenesis. These data suggest that, even within a controlled in vitro environment that is conducive to chondrogenesis, there may be an innate instability in the MSC phenotype that is independent of scaffold composition, and may ultimately limit their application in functional cartilage repair.

A biomimetic extracellular matrix for cartilage tissue engineering centered on photocurable gelatin, hyaluronic acid and chondroitin sulfate

The development of hydrogels tailored for cartilage tissue engineering has been a research and clinical goal for over a decade. Directing cells towards a chondrogenic phenotype and promoting new matrix formation are significant challenges that must be overcome for the successful application of hydrogels in cartilage tissue therapies. Gelatin–methacrylamide (Gel-MA) hydrogels have shown promise for the repair of some tissues, but have not been extensively investigated for cartilage tissue engineering. We encapsulated human chondrocytes in Gel-MA-based hydrogels, and show that with the incorporation of small quantities of photocrosslinkable hyaluronic acid methacrylate (HA-MA), and to a lesser extent chondroitin sulfate methacrylate (CS-MA), chondrogenesis and mechanical properties can be enhanced. The addition of HA-MA to Gel-MA constructs resulted in more rounded cell morphologies, enhanced chondrogenesis as assessed by gene expression and immunofluorescence, and increased quantity and distribution of the newly synthesized extracellular matrix (ECM) throughout the construct. Consequently, while the compressive moduli of control Gel-MA constructs increased by 26kPa after 8weeks culture, constructs with HA-MA and CS-MA increased by 114kPa. The enhanced chondrogenic differentiation, distribution of ECM, and improved mechanical properties make these materials potential candidates for cartilage tissue engineering applications.

Photo-crosslinked hyaluronic acid nanoparticles with improved stability for in vivo tumor-targeted drug delivery

One of the major hurdles of the nanoparticles as drug carriers is the unintended burst release of loaded drugs during blood circulation. To surmount this issue, we developed photo-crosslinked hyaluronic acid nanoparticles (c-HANPs) with improved stability for tumor-targeted drug delivery. They were readily prepared via UV-triggered chemical crosslinking with the acrylate groups in the polymer backbone. The size of c-HANPs was not much different from that of uncrosslinked HANPs. However, c-HANPs exhibited significantly high stability in a physiological buffer and released the loaded drug, paclitaxel (PTX), in a sustained manner. It is noteworthy that the drug release rate from c-HANPs remarkably increased in the presence of hyaluronidase, an enzyme abundant at the intracellular compartments of the tumor cells. It was found from in vitro cellular uptake tests that c-HANPs were rapidly taken up by the tumor cells via the receptor (CD44)-mediated endocytosis, which was not inhibited by photo-crosslinking. In non-invasive animal imaging results, they showed higher tumor-targeting ability than uncrosslinked HANPs because high stability of c-HANPs enabled their long circulation in the body. Owing to the sustained release of the drug and enhanced tumor-targeting ability, c-HANPs showed higher therapeutic efficacy compared to free PTX and uncrosslinked HANPs. These data implied the promising potential of c-HANP as tumor-targeting drug carriers and demonstrated the remarkable effect of the improved stability upon the biodistribution and therapeutic efficacy of drug-loaded nanoparticles.

Photocrosslinkable hyaluronic acid as an internal wetting agent in model conventional and silicone hydrogel contact lenses

Photocrosslinkable methacrylated hyaluronic acid (HA) was prepared and incorporated into model conventional and silicone hydrogel contact lenses as an internal wetting agent. The molecular weight of the HA, the degree of methacrylation as well as the amount (0.25 to 1.0 wt %) incorporated were varied. The HA-containing hydrogels were analyzed using a variety of techniques including water contact angles, equilibrium water content (EWC), and lysozyme sorption. The presence of HA could be detected in the materials using X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy-attenuated total reflectance. The materials containing methacrylated HA had improved hydrophilicity and reduced lysozyme sorption. Effects of modified HA on EWC were dependent upon the materials but generally increased water uptake. Increased mobility of the HA associated with a lower molecular weight and lower degree of methacrylation was found to be more effective in improving hydrophilicity and decreasing lysozyme sorption than the less mobile HA. All results found suggest that photocrosslinkable HA has significant potential in contact lens applications.

High mesenchymal stem cell seeding densities in hyaluronic acid hydrogels produce engineered cartilage with native tissue properties

Engineered cartilage based on adult mesenchymal stem cells (MSCs) is an alluring goal for the repair of articular defects. However, efforts to date have failed to generate constructs with sufficient mechanical properties to function in the demanding environment of the joint. Our findings with a novel photocrosslinked hyaluronic acid (HA) hydrogel suggest that stiff gels (high HA concentration, 5% w/v) foster chondrogenic differentiation and matrix production, but limit overall functional maturation due to the inability of the formed matrix to diffuse away from the point of production and form a contiguous network. In the current study, we hypothesized that increasing the MSC seeding density would decrease the required diffusional distance, and so expedite the development of functional properties. To test this hypothesis bovine MSCs were encapsulated at seeding densities of either 20,000,000 or 60,000,000cellsml–1 in 1%, 3%, and 5% (w/v) HA hydrogels. Counter to our hypothesis the higher concentration HA gels (3% and 5%) did not develop more rapidly with increased MSC seeding density. However, the biomechanical properties of the low concentration (1%) HA constructs increased markedly (nearly 3-fold with a 3-fold increase in seeding density). To ensure that optimal nutrient access was delivered, we next cultured these constructs under dynamic culture conditions (with orbital shaking) for 9weeks. Under these conditions 1% HA seeded at 60,000,000MSCsml–1 reached a compressive modulus in excess of 1MPa (compared with 0.3–0.4MPa for free swelling constructs). This is the highest level we have reported to date in this HA hydrogel system, and represents a significant advance towards functional stem cell-based tissue engineered cartilage.

Transient exposure to TGF-[formula omitted]3 improves the functional chondrogenesis of MSC-laden hyaluronic acid hydrogels

Tissue engineering with adult stem cells is a promising approach for the restoration of focal defects in articular cartilage. For this, progenitor cells would ideally be delivered to (and maintained within) the defect site via a biocompatible material and in combination with soluble factors to promote initial cell differentiation and subsequent tissue maturation in vivo. While growth factor delivery methods are continually being optimized, most offer only a short (days to weeks) delivery profile at high doses. To address this issue, we investigated mesenchymal stem cell (MSC) differentiation and maturation in photocrosslinkable hyaluronic acid (HA) hydrogels with transient exposure to the pro-chondrogenic molecule transforming growth factor-beta3 (TGF-β3), at varying doses (10, 50 and 100 ng/mL) and durations (3, 7, 21 and 63 days). Mechanical, biochemical, and histological outcomes were evaluated through 9 weeks of culture. Results showed that a brief exposure (7 days) to a very high level (100 ng/mL) of TGF-β3 was sufficient to both induce and maintain cartilage formation in these 3D constructs. Indeed, this short delivery resulted in constructs with mechanical and biochemical properties that exceeded that of continuous exposure to a lower level (10 ng/mL) of TGF-β3 over the entire 9-week time course. Of important note, the total TGF delivery in these two scenarios was roughly equivalent (200 vs. 180 ng), but the timing of delivery differed markedly. These data support the idea that acute exposure to a high dose of TGF will induce functional and long-term differentiation of stem cell populations, and further our efforts to improve cartilage repair in vivo.

Surface functionalization of hyaluronic acid hydrogels by polyelectrolyte multilayer films

Hyaluronic acid (HA), an anionic polysaccharide, is one of the major components of the natural extracellular matrix (ECM). Although HA has been widely used for tissue engineering applications, it does not support cell attachment and spreading and needs chemical modification to support cellular adhesion. Here, we present a simple approach to functionalize photocrosslinked HA hydrogels by deposition of poly( l-lysine) (PLL) and HA multilayer films made by the layer-by-layer (LbL) technique. PLL/HA multilayer film formation was assessed by using fluorescence microscopy, contact angle measurements, cationic dye loading and confocal microscopy. The effect of polyelectrolyte multilayer film (PEM) formation on the physicochemical and mechanical properties of hydrogels revealed polyelectrolyte diffusion inside the hydrogel pores, increased hydrophobicity of the surface, reduced equilibrium swelling, and reduced compressive moduli of the modified hydrogels. Furthermore, NIH-3T3 fibroblasts seeded on the surface showed improved cell attachment and spreading on the multilayer functionalized hydrogels. Thus, modification of HA hydrogel surfaces with multilayer films affected their physicochemical properties and improved cell adhesion and spreading on these surfaces. This new hydrogel/PEM composite system may offer possibilities for various biomedical and tissue engineering applications, including growth factor delivery and co-culture systems.

Controlling the adhesion and differentiation of mesenchymal stem cells using hyaluronic acid-based, doubly crosslinked networks

We have created hyaluronic acid (HA)-based, cell-adhesive hydrogels that direct the initial attachment and the subsequent differentiation of human mesenchymal stem cells (MSCs) into pre-osteoblasts without osteogenic supplements. HA-based hydrogel particles (HGPs) with an average diameter of 5–6 μm containing an estimated 2.2 wt% gelatin (gHGPs) were synthesized by covalent immobilization of gelatin to HA HGPs prepared via an inverse emulsion polymerization technique. Separately, a photocrosslinkable HA macromer (HAGMA) was synthesized by chemical modification of HA with glycidyl methacrylate (GMA). Doubly crosslinked networks (DXNs) were engineered by embedding gHGPs in a secondary network established by HAGMA at a particle concentration of 2.5 wt%. The resultant composite gels, designated as HA-gHGP, have an average compressive modulus of 21 kPa, and are non-toxic to the cultured MSCs. MSCs readily attached to these gels, exhibiting an early stage of stress fiber assembly 3 h post seeding. By day 7, stellate-shaped cells with extended filopodia were found on HA-gHGP gels. Moreover, cells had migrated deep into the matrix, forming a three dimensional, branched and interconnected cell community. Conversely, MSCs on the control gels lacking gelatin moieties formed isolated spheroids with rounded cell morphology. After 28 days of culture on HA-gHGP, Type I collagen production and mineral deposition were detected in the absence of osteogenic supplements, suggesting induction of osteogenic differentiation. In contrast, cells on the control gels expressed markers for adipogenesis. Overall, the HA-gHGP composite matrix has great promise for directing the osteogenic differentiation of MSCs by providing an adaptable environment through the spatial presentation of cell-adhesive modules.

Differential Maturation and Structure–Function Relationships in Mesenchymal Stem Cell- and Chondrocyte-Seeded Hydrogels

Degenerative disease and damage to articular cartilage represents a growing concern in the aging population. New strategies for engineering cartilage have employed mesenchymal stem cells (MSCs) as a cell source. However, recent work has suggested that chondrocytes (CHs) produce extracellular matrix (ECM) with superior mechanical properties than MSCs do. Because MSC-biomaterial interactions are important for both initial cell viability and subsequent chondrogenesis, we compared the growth of MSC- and CH-based constructs in three distinct hydrogels-agarose (AG), photocrosslinkable hyaluronic acid (HA), and self-assembling peptide (Puramatrix, Pu). Bovine CHs and MSCs were isolated from the same group of donors and seeded in AG, Pu, and HA at 20 million cells/mL. Constructs were cultured for 8 weeks with biweekly analysis of construct physical properties, viability, ECM content, and mechanical properties. Correlation analysis was performed to determine quantitative relationships between formed matrix and mechanical properties for each cell type in each hydrogel. Results demonstrate that functional chondrogenesis, as evidenced by increasing mechanical properties, occurred in each MSC-seeded hydrogel. Interestingly, while CH-seeded constructs were strongly dependent on the 3D environment in which they were encapsulated, similar growth profiles were observed in each MSC-laden hydrogel. In every case, MSC-laden constructs possessed mechanical properties significantly lower than those of CH-seeded AG constructs. This finding suggests that methods for inducing MSC chondrogenesis have yet to be optimized to produce cells whose functional matrix-forming potential matches that of native CHs.

In situ characterization of the degradation of PLGA microspheres in hyaluronic acid hydrogels by optical coherence tomography.

The polymeric implant material poly(lactide-co-glycolide) (PLGA) degrades by a process of bulk degradation, which allows it to be used for the controlled release of therapeutic molecules from implants and microspheres. The temporal characterization of PLGA microsphere degradation has been limited by the need to destructively monitor the samples at each time point. In this study, a noninvasive imaging technology, optical coherence tomography (OCT), was utilized to characterize the in situ degradation of PLGA microspheres suspended within photo-crosslinked hyaluronic acid (HA) hydrogels. Microspheres with differing degradation rates were loaded with bovine serum albumin (BSA) as a marker protein, and temporal release of protein was correlated with morphological changes observed during 3-D OCT imaging. As proof-of-principle, a microsphere-loaded hydrogel scaffold was implanted in a modified rat calvarial critical size defect model and imaged using OCT. This animal model presents the opportunity to monitor microsphere degradation over time in living animals.

Drug‐binding hydrogels of hyaluronic acid functionalized with β‐cyclodextrin

Hyaluronic acid (HA) hydrogels are attractive materials for biomedical applications because they are porous, water-swelling, biocompatible, biodegradable, and resistant to non-specific cell adhesion. A limitation of HA hydrogels is that incorporation of bioactive drugs can be restricted by low solubility of drug within the hydrogel environment. Our goal was to synthesize HA hydrogels that bind drug through hydrophobic interactions as a method for increasing drug loading. We functionalized photocrosslinked HA hydrogels with a methacryloyl derivative of beta-cyclodextrin (betaCD). betaCD is a molecular "basket" with a hydrophilic exterior and a hydrophobic cavity. Inclusion complexes are formed when betaCD hosts all or part of a hydrophobic drug within the cavity. HA hydrogels functionalized with methacryloyl-betaCD monomer gained the property of inclusion complexation which greatly enhanced the uptake of a model hydrophobic drug, hydrocortisone. Pre-incubation of the hydrogels with adamantane carboxylic acid (ACA) inhibited hydrocortisone uptake by competition for betaCD cavities. In addition, control hydrogels of HA functionalized with alphaCD monomer were not efficient at hydrocortisone uptake because the alphaCD cavity is too small for efficient complexation. These experiments confirmed that the betaCD monomer enhances drug loading by the mechanism of inclusion complexation. Drug-binding HA-betaCD hydrogels may be further engineered to create HA-based biomaterials with a built in drug delivery capability.

Micromolding of photocrosslinkable hyaluronic acid for cell encapsulation and entrapment

Micropatterning of hydrogels is potentially useful for a variety of applications, including tissue engineering, fundamental biological studies, diagnostics, and high-throughput screening. Although synthetic polymers have been developed for these applications, natural polymers such as polysaccharides may have advantages for biological samples and cell-based devices because they are natural components of the in vivo microenvironment. In this study, we synthesized and used hyaluronic acid (HA) modified with photoreactive methacrylates to fabricate microstructures as functional components of microfabricated devices. To demonstrate the universality of this approach, two types of microstructures were formed. In the first approach, HA microstructures were fabricated and used as docking templates to enable the subsequent formation of cell microarrays within low shear stress regions of the patterns. Cells within these microwells remained viable, could generate spheroids, and could be retrieved using mechanical disruption. In the second approach, cells were encapsulated directly within the HA hydrogels. Arrays of viable embryonic stem (ES) cells or fibroblasts were encapsulated within HA hydrogels and could later be recovered using enzymatic digestion of the microstructures. These results demonstrate that it is possible to incorporate photocrosslinkable HA, a natural, versatile, degradable, and biocompatible biopolymer, into micro-electromechanical systems.