Era Jain

A comparison of cryogel scaffolds to identify an appropriate structure for promoting bone regeneration

To create an ideal graft substitute for regenerating bone, the scaffold should possess osteoconductive, osteoinductive, and osteogenic properties. Hydrogels are a very common scaffold, but the mechanical integrity and nanoporous structure are not advantageous for bone regeneration. Cryogelation is a technique in which the controlled freezing and thawing of a polymer creates a spongy, macroporous structure with ideal structural characteristics and promising mechanical stability. Hydrogels and cryogels of three different materials (chitosan–gelatin, N-vinyl-2-pyrrolidone, and silk fibroin (SF)) were compared to assess the optimal material and form of scaffold for this application. Cryogel and hydrogel structures were tested in parallel to evaluate porosity, swelling, mechanical integrity, cellular infiltration, and mineralization potential. Cryogels proved superior to hydrogels based on swelling potential and mechanical properties. Among the cryogels, SF demonstrated high pore diameter and area, mineralization upon cellular infiltration, and the largest presence of osteocalcin, a marker of bone formation. These results demonstrate the practicality of cryogels for a bone regeneration application and identify SF as a potential material choice.

Biomaterials for liver tissue engineering

Liver extracellular matrix (ECM) composition, topography and biomechanical properties influence cell-matrix interactions. The ECM presents guiding cues for hepatocyte phenotype maintenance, differentiation and proliferation both in vitro and in vivo. Current understanding of such cell-guiding cues along with advancement of techniques for scaffold fabrication has led to evolution of matrices for liver tissue culture from simple porous scaffolds to more complex 3D matrices with microarchitecture similar to in vivo. Natural and synthetic polymeric biomaterials fabricated in different topographies and porous matrices have been used for hepatocyte culture. Heterotypic and homotypic cell interactions are necessary for developing an adult liver as well as an artificial liver. A high oxygen demand of hepatocytes as well as graded oxygen distribution in liver is another challenging attribute of the normal liver architecture that further adds to the complexity of engineered substrate design. A balanced interplay of cell-matrix interactions along with cell-cell interactions and adequate supply of oxygen and nutrient determines the success of an engineered substrate for liver cells. Techniques devised to incorporate these features of hepatic function and mimic liver architecture range from maintaining liver cells in mm-sized tailor-made scaffolds to a more bottoms up approach that starts from building the microscopic subunit of the whole tissue. In this review, we discuss briefly various biomaterials used for liver tissue engineering with respect to design parameters such as scaffold composition and chemistry, biomechanical properties, topography, cell-cell interactions and oxygenation.

Design of electrohydrodynamic sprayed polyethylene glycol hydrogel microspheres for cell encapsulation

Electrohydrodynamic spraying (EHS) has recently gained popularity for microencapsulation of cells for applications in cell delivery and tissue engineering. Some of the polymers compatible with EHS are alginate, chitosan, and other similar natural polymers, which are subject to ionotropic or physical gelation. It is desirable to further extend the use of the EHS technique beyond such polymers for wider biofabrication applications. Here, building upon our previous work of making PEG microspheres via EHS, we utilized the principles of EHS to fabricate cell-laden polyethylene glycol (PEG) hydrogel microspheres. The gelation of PEG hydrogel microspheres was achieved by forming covalent crosslinks between multiarm PEG acrylate and dithiol crosslinkers via Michael-type addition. We conducted a detailed investigation of the critical parameters of EHS, such as the applied voltage, inner needle diameter (i.d. needle), and flow rate, to obtain PEG microspheres with high cell viability and tightly-controlled diameters in the range of 70-300 μm. The polydispersity of cell-laden PEG hydrogel microspheres as measured by % coefficient of variation was between 6% and 23% for all conditions tested. We established that our method was compatible with different cell types and that all tested cell types could be encapsulated at high densities of 106-109 and ≥90% encapsulation efficiency. We observed cell aggregation within the hydrogel microspheres at applied voltage >5 kV. Since PEG is a synthetic polymer devoid of cell attachment sites, we could overcome this limitation by tethering Arg-Gly-Asp-Ser (RGDS) peptide to the PEG hydrogel microspheres; upon RGDS tethering, we observed uniform cell dispersion. The microencapsulated cells could be cultured in the PEG hydrogel microspheres of different sizes for up to one week without significant loss in cell viability. In conclusion, the EHS technique developed here could be used to generate cell-laden PEG hydrogel microspheres of controlled sizes for potential applications in cell delivery and organoid cultures.

Whispering gallery mode resonator sensor for in situ measurements of hydrogel gelation

Whispering gallery mode (WGM) resonators are compact and ultrasensitive devices, which enable label-free sensing at the single-molecule level. Despite their high sensitivity, WGM resonators have not been thoroughly investigated for use in dynamic biochemical processes including molecular diffusion and polymerization. In this work, the first report of using WGM sensors to continuously monitor a chemical reaction (i.e. gelation) in situ in a hydrogel is described. Specifically, we monitor and quantify the gelation dynamics of polyacrylamide hydrogels using WGM resonators and compare the results to an established measurement method based on rheology. Rheology measures changes in viscoelasticity, while WGM resonators measure changes in refractive index. Different gelation conditions were studied by varying the total monomer concentration and crosslinker concentration of the hydrogel precursor solution, and the resulting similarities and differences in the signal from the WGM resonator and rheology are elucidated. This work demonstrates that WGM alone or in combination with rheology can be used to investigate the gelation dynamics of hydrogels to provide insights into their gelation mechanisms.

Control of gelation, degradation and physical properties of polyethylene glycol hydrogels through the chemical and physical identity of the crosslinker

Tuning hydrogel properties through minor modifications of the crosslinker structure is a beneficial approach for hydrogel design that could result in hydrogels with wide range of properties to match a desired application. In this study, we analyzed the relationship between the dithiol crosslinker chemical and physical structure and the resulting properties of polyethylene glycol (PEG) hydrogels formed via Michael-type addition reaction. Specifically, the dithiol crosslinker properties and chemical structure were correlated with gelation time, hydrolytic degradation rate, reaction rate constant, crosslink density and storage modulus of PEG hydrogels. By changing the properties and structure of the crosslinker, hydrogels with controlled degradation ranging from 10 h to 22 d were obtained. It was also established that hydrogel gelation times correlated closely with degradation times. By extensive characterization of the dithiol crosslinker chemical structure and physical properties, we identified two sets of conditions which yielded fast-gelling, fast-degrading hydrogels and slow-gelling, slow-degrading hydrogels. Uniquely, the hydrogel storage moduli could be controlled by the dithiol crosslinker chemical identity independent of the degradation time of the hydrogel or the mesh size.

Sustained release of multicomponent platelet‐rich plasma proteins from hydrolytically degradable PEG hydrogels

Platelet-rich plasma (PRP), an autologous blood derived product is a concentrated mix of multiple growth factors and cytokines. Direct injections of PRP are clinically used for treatment of various musculoskeletal disorders and in wound healing. However, PRP therapy has met with limited clinical success possibly due to unpredictable and premature bolus delivery of PRP growth factors. The objective of this study was to predictably control the bioavailability of PRP growth factors using a hydrolytically degradable polyethylene glycol (PEG) hydrogel. We used a step-growth polymerization based on a Michael-type addition reaction between a 6-arm PEG-acrylate and a dithiol crosslinker, which led to the formation of a homogenous hydrogel network under mild, physiologically relevant conditions. Specifically, to model the release of multicomponent PRP through PEG hydrogels, we examined bulk diffusion of PRP as well as model proteins in a size range corresponding to that of growth factors found in PRP. Our results indicated that protein size and hydrogel degradation controlled diffusion of all proteins and that secondary structure of proteins encapsulated during gelation remained unaffected post-release. Analysis of specific PRP proteins released from the hydrogel showed sustained release until complete hydrogel degradation. PRP released from hydrogels promoted proliferation of human dermal fibroblast, indicating retained bioactivity upon encapsulation and release. The versatile hydrogel system holds clinical potential as a therapeutic drug delivery depot of multicomponent mixtures like PRP.

Advances in bionanocomposites for biomedical applications

Bionanocomposites possess exceptional mechanical strength and bioactivity, which makes them a valuable candidate for various tissue engineering and drug delivery applications. The unique combination of hard and soft components in an engineered bionanocomposite resembles the amalgamation of different components in naturally tissues. Some of the greatest challenge in the designing of bionanocomposites for tissue regeneration and biomedical applications is to have high mechanical strength coupled with biocompatibility, bioactivity, and hierarchical structure of natural tissue. Several biopolymers have been combined with nanofillers in different categories such as silicate-based materials such as clays and silica nanoparticles, ceramics such as nanohydroxyapatite, inorganic nanoparticles, synthetic layered double hydroxides, carbon-based nanomaterials such as carbon nanotubes, and metal/metal oxides. Because of the extraordinary mechanical properties of bionanocomposites, much interest lies in mimicking the bone structure such as dental implants and other orthopedic applications, as they allow for multilevel integration of material, structural, and biological properties constituted by the polymer and nanofiller combination. Presence of nanosized fillers in bionanocomposites makes them ideally suited for drug delivery, as they usually present torturous diffusion path for encapsulated small molecule/drug forming effective barrier and sustained delivery. Furthermore, drug releasing bionanocomposites are highly suited for wound dressing applications, as they have high water uptake and noncytotoxicity together, high mucoadhesivity, and tear resistance making them ideal as wound dressing. The developments of bionanocomposites provide new avenues for fulfilling certain needs of the emerging technologies in matrix formation, tissue regeneration, drug delivery, and wound dressing and will be instrumental in accelerating evolution of new therapeutics.

Cell Microencapsulation in Polyethylene Glycol Hydrogel Microspheres Using Electrohydrodynamic Spraying

Microencapsulation of cells is beneficial for various biomedical applications, such as tissue regeneration and cell delivery. While a variety of techniques can be used to produce microspheres, electrohydrodynamic spraying (EHS) has shown promising results for the fabrication of cell-laden hydrogel microspheres in a wide range of sizes and in a relatively high-throughput manner. Here we describe an EHS technique for the fabrication of cell-laden polyethylene glycol (PEG) microspheres. We utilize mild hydrogel gelation chemistry and a combination of EHS parameters to allow for cell microencapsulation with high efficiency and viability. We also give examples on the effect of different EHS parameters such as inner diameter of the needle, voltage and flow rate on microsphere size and encapsulated cell viability.

UV Dose Governs UV-Polymerized Polyacrylamide Hydrogel Modulus

Polyacrylamide (PAA) hydrogels have become a widely used tool whose easily tunable mechanical properties, biocompatibility, thermostability, and chemical inertness make them invaluable in many biological applications, such as cell mechanosensitivity studies. Currently, preparation of PAA gels involves mixtures of acrylamide, bisacrylamide, a source of free radicals, and a chemical stabilizer. This method, while generally well accepted, has its drawbacks: long polymerization times, unstable and toxic reagents, and tedious preparation. Alternatively, PAA gels could be made by free radical polymerization (FRP) using ultraviolet (UV) photopolymerization, a method which is quicker, less tedious, and less toxic. Here, we describe a simple strategy based on total UV energy for determining the optimal UV crosslinking conditions that lead to optimal hydrogel modulus.