Injectable In Situ Crosslinking Hydrogel for Autologous Fat Grafting.

Autologous fat grafting is hampered by unpredictable outcomes due to high tissue resorption. Hydrogels based on enzymatically pretreated tunicate nanocellulose (ETC) and alginate (ALG) are biocompatible, safe, and present physiochemical properties capable of promoting cell survival. Here, we compared in situ and ex situ crosslinking of ETC/ALG hydrogels combined with lipoaspirate human adipose tissue (LAT) to generate an injectable formulation capable of retaining dimensional stability in vivo. We performed in situ crosslinking using two different approaches; inducing Ca2+ release from CaCO3 microparticles (CMPs) and physiologically available Ca2+ in vivo. Additionally, we generated ex situ-crosslinked, 3D-bioprinted hydrogel-fat grafts. We found that in vitro optimization generated a CMP-crosslinking system with comparable stiffness to ex situ-crosslinked gels. Comparison of outcomes following in vivo injection of each respective crosslinked hydrogel revealed that after 30 days, in situ crosslinking generated fat grafts with less shape retention than 3D-bioprinted constructs that had undergone ex situ crosslinking. However, CMP addition improved fat-cell distribution and cell survival relative to grafts dependent on physiological Ca2+ alone. These findings suggested that in situ crosslinking using CMP might promote the dimensional stability of injectable fat-hydrogel grafts, although 3D bioprinting with ex situ crosslinking more effectively ensured proper shape stability in vivo.

Assessing cell migration in hydrogels: An overview of relevant materials and methods.

Cell migration is essential in numerous living processes, including embryonic development, wound healing, immune responses, and cancer metastasis. From individual cells to collectively migrating epithelial sheets, the locomotion of cells is tightly regulated by multiple structural, chemical, and biological factors. However, the high complexity of this process limits the understanding of the influence of each factor. Recent advances in materials science, tissue engineering, and microtechnology have expanded the toolbox and allowed the development of biomimetic in vitro assays to investigate the mechanisms of cell migration. Particularly, three-dimensional (3D) hydrogels have demonstrated a superior ability to mimic the extracellular environment. They are therefore well suited to studying cell migration in a physiologically relevant and more straightforward manner than in vivo approaches. A myriad of synthetic and naturally derived hydrogels with heterogeneous characteristics and functional properties have been reported. The extensive portfolio of available hydrogels with different mechanical and biological properties can trigger distinct biological responses in cells affecting their locomotion dynamics in 3D. Herein, we describe the most relevant hydrogels and their associated physico-chemical characteristics typically employed to study cell migration, including established cell migration assays and tracking methods. We aim to give the reader insight into existing literature and practical details necessary for performing cell migration studies in 3D environments.

Alginate Microsphere Encapsulation of Drug-Loaded Nanoparticles: A Novel Strategy for Intraperitoneal Drug Delivery.

Alginate hydrogels have been broadly investigated for use in medical applications due to their biocompatibility and the possibility to encapsulate cells, proteins, and drugs. In the treatment of peritoneal metastasis, rapid drug clearance from the peritoneal cavity is a major challenge. Aiming to delay drug absorption and reduce toxic side effects, cabazitaxel (CAB)-loaded poly(alkyl cyanoacrylate) (PACA) nanoparticles were encapsulated in alginate microspheres. The PACAlg alginate microspheres were synthesized by electrostatic droplet generation and the physicochemical properties, stability, drug release kinetics, and mesothelial cytotoxicity were analyzed before biodistribution and therapeutic efficacy were studied in mice. The 450 µm microspheres were stable at in vivo conditions for at least 21 days after intraperitoneal implantation in mice, and distributed evenly throughout the peritoneal cavity without aggregation or adhesion. The nanoparticles were stably retained in the alginate microspheres, and nanoparticle toxicity to mesothelial cells was reduced, while the therapeutic efficacy of free CAB was maintained or improved in vivo. Altogether, this work presents the alginate encapsulation of drug-loaded nanoparticles as a promising novel strategy for the treatment of peritoneal metastasis that can improve the therapeutic ratio between toxicity and therapeutic efficacy.

MS-proteomics provides insight into the host responses towards alginate microspheres.

Protein adsorption to biomaterial surfaces is considered a determining factor for the host response. Here we detail the protein adsorption profiles of alginate hydrogel microspheres relevant for cell therapy using mass spectrometry (MS)-based proteomics. The investigated microspheres include sulfated alginate (SA), high G alginate (HiG), and poly-l-lysine coated alginate (AP), which previously have been shown to exhibit different inflammatory and fibrotic responses. The biological significance was assessed in lepirudin-anticoagulated human whole blood (hWB) by functional analysis of the acute-phase responses (complement and coagulation). Proteomic profiling revealed distinct signatures for the microspheres, wherein Ingenuity Pathway Analysis identified complement and coagulation as the top enriched canonical pathways. The levels of complement and coagulation activators and inhibitors were distinctly different, which was reflected in the functional hWB analyses: SA was highly enriched with inhibitory factors of complement and coagulation (e.g. C1 inhibitor, factor H, antithrombin-III, heparin cofactor 2), other heparin-binding proteins and factors promoting fibrinolysis (factor XII, plasma kallikrein), conforming to an anti-inflammatory and anti-fibrotic profile. HiG enriched moderate levels of complement inhibitors, conforming to a low-inflammatory and pro-fibrotic profile. AP showed the most prominent enrichment of complement activators (e.g. C3, properdin, C-reactive protein) and low levels of inhibitors, conforming to a pro-inflammatory and highly pro-fibrotic profile. In conclusion, the extensive enrichment of inhibitory acute-phase proteins on SA could be a determining factor for its reduced host response. The interactions between the plasma proteins and hydrogel surfaces shown herein point to proteomics as an important supplement to existing in vitro and in vivo methods for designing biocompatible alginate-based hydrogels.

Alginate and tunicate nanocellulose composite microbeads - Preparation, characterization and cell encapsulation.

Alginate has been used for decades for cell encapsulation. Cellulose nanofibrils (CNF) from tunicates are desirable in biomedicine due to high molecular weight, purity, crystallinity, and sustainable production. We prepared microbeads of 400-600 µm of alginate and tunicate CNF. Greater size, dispersity and aspect ratio were observed in microbeads with higher fractions of CNF. CNF content in Ca-crosslinked alginate microbeads decreased stability upon saline exposure, whereas crosslinking with calcium (50 mM) and barium (1 mM) yielded stable microbeads. The Young's moduli of gel cylinders decreased when exchanging alginate with CNF, and slightly increased permeability to dextran was observed in microbeads containing CNF. Encapsulation of MC3T3 cells revealed high cell viability after encapsulation (83.6 ± 0.4%) in beads of alginate and CNF. NHDFs showed lower viability but optimizing mixing and production techniques of microbeads increased cell viability (from 66.2 ± 5.3% to 72.7 ± 7.5%).

Carbohydr Polym Special Issue Invited contribution: Click chemistry for block polysaccharides with dihydrazide and dioxyamine linkers - A review.

Engineered block polysaccharides is a relatively new class of biomacromolecules consisting of chemical assembly of separate block structures at the chain termini. In contrast to conventional, laterally substituted polysaccharide derivatives, the block arrangement allows for much higher preservation of inherent chain properties such as biodegradability and stimuli-responsive self-assembly, while at the same time inducing new macromolecular properties. Abundant, carbon neutral, and even recalcitrant biomass is an excellent source of blocks, opening for numerous new uses of biomass for a wide range of novel biomaterials. Among a limited range of methodologies available for block conjugation, bifunctional linkers allowing for oxyamine and hydrazide 'click' reactions have recently proven useful additions to the repertoire. This article focuses the chemistry and kinetics of these reactions. It also presents some new data with the aim to provide useful protocols and methods for general use towards new block polysaccharides.

Pericapsular fibrotic overgrowth mitigated in immunocompetent mice through microbead formulations based on sulfated or intermediate G alginates.

Cell encapsulation in alginate microbeads is a promising approach to provide immune isolation in cell therapy without immunosuppression. However, the efficacy is hampered by pericapsular fibrotic overgrowth (PFO), causing encapsulated cells to lose function. Stability of the microbeads is important to maintain immune isolation in the long-term. Here, we report alginate microbeads with minimal PFO in immunocompetent C57BL/6JRj mice. Microbead formulations included either alginate with an intermediate (47 %) guluronate (G) content (IntG) or sulfated alginate (SA), gelled in Ca2+/Ba2+ or Sr2+. A screening panel of eleven microbead formulations were evaluated for PFO, yielding multiple promising microbeads. Two candidate formulations were evaluated for 112 days in vivo, exhibiting maintained stability and minimal PFO. Microbeads investigated in a human whole blood assay revealed low cytokine and complement responses, while SA microbeads activated coagulation. Protein deposition on microbeads explanted from mice investigated by confocal laser scanning microscopy (CLSM) showed minimal deposition of complement C3. Fibrinogen was positively associated with PFO, with a high deposition on microbeads of high G (68 %) alginate compared to IntG and SA microbeads. Overall, stable microbeads containing IntG or SA may serve in long-term therapeutic applications of cell encapsulation. STATEMENT OF SIGNIFICANCE: Alginate-based hydrogels in the format of micrometer size beads is a promising approach for the immunoisolation of cells in cell therapy. Clinical trials in type 1 diabetes have so far had limited success due to fibrotic responses that hinder the diffusion of nutrients and oxygen to the encapsulated cells, resulting in graft failure. In this study, minimal fibrotic response towards micrometer size alginate beads was achieved by chemical modification of alginate with sulfate groups. Also, the use of alginate with intermediate guluronic acid content resulted in minimally fibrotic microbeads. Fibrinogen deposition was revealed to be a good indicator of fibrosis. This study points to both new microsphere developments and novel insight in the mechanisms behind the fibrotic responses.

High resolution imaging of soft alginate hydrogels by atomic force microscopy.

This work explores the largely unknown surface microstructure and elastic modulus of soft calcium-alginate hydrogels (E = 100-4500 Pa) in their hydrated state by atomic force microscopy (AFM) in quantitative imaging mode. Alginate concentration influenced the surface topography with surface roughness measured to be 101 ± 6 nm and 57 ± 1 nm for 0.5 and 2.0% (w/v) alginate, respectively. The calculated range of pore sizes increased with decreasing alginate concentration, with radii smaller than 360 nm, 570 nm and 1230 nm for 2.0%, 1.0% and 0.5% alginate, respectively. Small changes in calcium concentration (from 20 to 25 mM, 1.5% alginate) did not induce changes in surface microstructure, although it increased the elastic modulus mean values and distribution. Introducing oxidized or peptide-grafted alginate in the gels resulted in rougher surfaces, larger pore sizes and lower elasticity than the respective hydrogels with no modified alginate.

Alginate hydrogels functionalized with ß-cyclodextrin as a local paclitaxel delivery system.

Modification of drug delivery materials with beta-cyclodextrins (ß-CyD) is known to increase solubility of poorly water-soluble drugs, protect drugs from degradation and sustain release. In this study, we developed a hydrogel drug delivery system for local paclitaxel delivery using the natural polysaccharide alginate functionalized with ß-CyD-moieties. Paclitaxel was chosen due to its ability to form inclusion complexes with cyclodextrins. The rheological and mechanical properties of the prepared hydrogels were characterized, as well as in vitro release of the paclitaxel and in vitro activity on PC-3 prostate cancer cells. Introduction of ß-CyD-moieties into the hydrogel reduces the mechanical properties of the gels compared to nonmodified gels. However, gelation kinetics were not markedly different. Furthermore, the ß-CyD-modified alginate helped to reduce undesired crystallization of the paclitaxel in the gel and facilitated paclitaxel diffusion out of the gel network. Remarkably, the ß-CyD grafted alginate showed increased capacity to complex paclitaxel compared to free HPß-CyD. Release of both paclitaxel and degradation products were measured from the gels and were shown to have cytotoxic effects on the PC-3 cells. The results indicate that functionalized alginate with ß-CyDs has potential as a material for drug delivery systems.

Sulfated Alginate Reduces Pericapsular Fibrotic Overgrowth on Encapsulated cGMP-Compliant hPSC-Hepatocytes in Mice.

Intra-peritoneal placement of alginate encapsulated human induced pluripotent stem cell-derived hepatocytes (hPSC-Heps) represents a potential new bridging therapy for acute liver failure. One of the rate-limiting steps that needs to be overcome to make such a procedure more efficacious and safer is to reduce the accumulation of fibrotic tissue around the encapsulated cells to allow the free passage of relevant molecules in and out for metabolism. Novel chemical compositions of alginate afford the possibility of achieving this aim. We accordingly used sulfated alginate and demonstrated that this material reduced fibrotic overgrowth whilst not impeding the process of encapsulation nor cell function. Cumulatively, this suggests sulfated alginate could be a more suitable material to encapsulate hPSC-hepatocyte prior to human use.

Injectable Gel Form of a Decellularized Bladder Induces Adipose-Derived Stem Cell Differentiation into Smooth Muscle Cells In Vitro.

Biologic scaffolds composed of extracellular matrix components have been proposed to repair and reconstruct a variety of tissues in clinical and pre-clinical studies. Injectable gels can fill and conform any three-dimensional shape and can be delivered to sites of interest by minimally invasive techniques. In this study, a biological gel was produced from a decellularized porcine urinary bladder by enzymatic digestion with pepsin. The enzymatic digestion was confirmed by visual inspection after dissolution in phosphate-buffered saline solution and Fourier-transform infrared spectroscopy. The rheological and biological properties of the gel were characterized and compared to those of the MatrigelTM chosen as a reference material. The storage modulus G' reached 19.4 ± 3.7 Pa for the 30 mg/mL digested decellularized bladder gels after ca. 3 h at 37 °C. The results show that the gel formed of the porcine urinary bladder favored the spontaneous differentiation of human and rabbit adipose-derived stem cells in vitro into smooth muscle cells to the detriment of cell proliferation. The results support the potential of the developed injectable gel for tissue engineering applications to reconstruct for instance the detrusor muscle part of the human urinary bladder.

Encapsulation boosts islet-cell signature in differentiating human induced pluripotent stem cells via integrin signalling.

Cell replacement therapies hold great therapeutic potential. Nevertheless, our knowledge of the mechanisms governing the developmental processes is limited, impeding the quality of differentiation protocols. Generating insulin-expressing cells in vitro is no exception, with the guided series of differentiation events producing heterogeneous cell populations that display mixed pancreatic islet phenotypes and immaturity. The achievement of terminal differentiation ultimately requires the in vivo transplantation of, usually, encapsulated cells. Here we show the impact of cell confinement on the pancreatic islet signature during the guided differentiation of alginate encapsulated human induced pluripotent stem cells (hiPSCs). Our results show that encapsulation improves differentiation by significantly reshaping the proteome landscape of the cells towards an islet-like signature. Pathway analysis is suggestive of integrins transducing the encapsulation effect into intracellular signalling cascades promoting differentiation. These analyses provide a molecular framework for understanding the confinement effects on hiPSCs differentiation while confirming its importance for this process.

Efficient Grafting of Cyclodextrin to Alginate and Performance of the Hydrogel for Release of Model Drug.

Controlling the rate of release of molecules from a hydrogel is of high interest for various drug delivery systems and medical devices. A strategy to alter the release profiles of soluble and poorly soluble active ingredients from hydrogels can be to combine the hydrogel forming ability of alginate with the inclusion forming ability of cyclodextrins (CyD). Here, ß-CyD was grafted to alginate in a three-step synthesis using periodate oxidation, reductive amination and copper(I)-catalyzed azide-alkyne cycloaddition. A grafting degree of 4.7% mol ß-CyD/mol sugar residues was obtained. The grafting degree was controlled by varying the reaction parameters where the amount of linker used in reductive amination was especially influential. Ca-alginate gel beads grafted with ß-CyD showed increased uptake of the model molecule methyl orange. Release experiments showed that the grafted material had a prolonged release of methyl orange and an increased total amount of released methyl orange. These results show that the ß-CyD grafted alginate is still able to form a hydrogel while the grafted cyclodextrins retain their ability to form inclusion complex with methyl orange. Further testing should be done with this system to investigate capability for drug delivery applications.

Mechanical Properties of Ca-Saturated Hydrogels with Functionalized Alginate.

In this work, the mechanical properties and stability of alginate hydrogels containing functionalized alginates (peptide and ß-cyclodextrin) were studied. There is an increasing interest in the modification of alginates to add functions such as cell attachment and increased solubility of hydrophobic drugs, for better performance in tissue engineering and drug release, respectively. Functionalization was achieved in this study via periodate oxidation followed by reductive amination, previously shown to give a high and controllable degree of substitution. Young's modulus and the stress at rupture of the hydrogels were in general lowered when exchanging native alginate with the modified alginate. Still, the gel strength could be adjusted by the fraction of modified alginate in the mixed hydrogels as well as the degree of oxidation. No notable difference in deformation at rupture was observed while syneresis was influenced by the degree of oxidation and possibly by the nature and amount of the grafted molecules. The mixed hydrogels were less stable than hydrogels with only native alginate, and modified alginate was released from the hydrogels. Furthermore, the hydrogels in general rather disintegrated than swelled upon saline treatments.

Alginate encapsulation as long-term immune protection of allogeneic pancreatic islet cells transplanted into the omental bursa of macaques.

The transplantation of pancreatic islet cells could restore glycaemic control in patients with type-I diabetes. Microspheres for islet encapsulation have enabled long-term glycaemic control in diabetic rodent models; yet human patients transplanted with equivalent microsphere formulations have experienced only transient islet-graft function, owing to a vigorous foreign-body reaction (FBR), to pericapsular fibrotic overgrowth (PFO) and, in upright bipedal species, to the sedimentation of the microspheres within the peritoneal cavity. Here, we report the results of the testing, in non-human primate (NHP) models, of seven alginate formulations that were efficacious in rodents, including three that led to transient islet-graft function in clinical trials. Although one month post-implantation all formulations elicited significant FBR and PFO, three chemically modified, immune-modulating alginate formulations elicited reduced FBR. In conjunction with a minimally invasive transplantation technique into the bursa omentalis of NHPs, the most promising chemically modified alginate derivative (Z1-Y15) protected viable and glucose-responsive allogeneic islets for 4 months without the need for immunosuppression. Chemically modified alginate formulations may enable the long-term transplantation of islets for the correction of insulin deficiency.

Current and Future Perspectives on Alginate Encapsulated Pancreatic Islet.

Transplantation of pancreatic islets in immune protective capsules holds the promise as a functional cure for type 1 diabetes, also about 40 years after the first proof of principal study. The concept is simple in using semipermeable capsules that allow the ingress of oxygen and nutrients, but limit the access of the immune system. Encapsulated human islets have been evaluated in four small clinical trials where the procedure has been evaluated as safe, but lacking long-term efficacy. Host reactions toward the biomaterials used in the capsules may be one parameter limiting the long-term function of the graft in humans. The present article briefly discusses important capsule properties such as stability, permeability and biocompatibility, as well as possible strategies to overcome current challenges. Also, recent progress in capsule development as well as the production of insulin-producing cells from human stem cells that gives promising perspectives for the transplantation of encapsulated insulin-producing tissue is briefly discussed. Stem Cells Translational Medicine 2017;6:1053-1058.

Mechanical Properties of Composite Hydrogels of Alginate and Cellulose Nanofibrils.

Alginate and cellulose nanofibrils (CNF) are attractive materials for tissue engineering and regenerative medicine. CNF gels are generally weaker and more brittle than alginate gels, while alginate gels are elastic and have high rupture strength. Alginate properties depend on their guluronan and mannuronan content and their sequence pattern and molecular weight. Likewise, CNF exists in various qualities with properties depending on, e.g., morphology and charge density. In this study combinations of three types of alginate with different composition and two types of CNF with different charge and degree of fibrillation have been studied. Assessments of the composite gels revealed that attractive properties like high rupture strength, high compressibility, high gel rigidity at small deformations (Young's modulus), and low syneresis was obtained compared to the pure gels. The effects varied with relative amounts of CNF and alginate, alginate type, and CNF quality. The largest effects were obtained by combining oxidized CNF with the alginates. Hence, by combining the two biopolymers in composite gels, it is possible to tune the rupture strength, Young's modulus, syneresis, as well as stability in physiological saline solution, which are all important properties for the use as scaffolds in tissue engineering.

Gelling kinetics and in situ mineralization of alginate hydrogels: A correlative spatiotemporal characterization toolbox.

UNLABELLED: Due to their large water content and structural similarities to the extracellular matrix, hydrogels are an attractive class of material in the tissue engineering field. Polymers capable of ionotropic gelation are of special interest due to their ability to form gels at mild conditions. In this study we have developed an experimental toolbox to measure the gelling kinetics of alginate upon crosslinking with calcium ions. A reaction-diffusion model for gelation has been used to describe the diffusion of calcium within the hydrogel and was shown to match experimental observations well. In particular, a single set of parameters was able to predict gelation kinetics over a wide range of gelling ion concentrations. The developed model was used to predict the gelling time for a number of geometries, including microspheres typically used for cell encapsulation. We also demonstrate that this toolbox can be used to spatiotemporally investigate the formation and evolution of mineral within the hydrogel network via correlative Raman microspectroscopy, confocal laser scanning microscopy and electron microscopy. STATEMENT OF SIGNIFICANCE: Hydrogels show great promise in cell-based tissue engineering, however new fabrication and modification methods are needed to realize the full potential of hydrogel based materials. The inclusion of an inorganic phase is one such approach and is known to affect both cell-material interactions and mechanical properties. This article describes the development of a correlative experimental approach where gel formation and mineralization has been investigated with spatial and temporal resolution by applying Raman microspectroscopy, optical and electron microscopy and a reaction-diffusion modeling scheme. Modeling allows us to predict gelling kinetics for other geometries and sizes than those investigated experimentally. Our experimental system enables non-destructive study of composite hydrogel systems relevant for, but not limited to, applications within bone tissue engineering.

A correlative spatiotemporal microscale study of calcium phosphate formation and transformation within an alginate hydrogel matrix.

UNLABELLED: The modification of soft hydrogels with hard inorganic components is a method used to form composite materials with application in non-load-bearing bone tissue engineering. The inclusion of an inorganic component may provide mechanical enhancement, introduce osteoconductive or osteoinductive properties, or change other aspects of interactions between native or implanted cells and the material. A thorough understanding of the interactions between such components is needed to improve the rational design of such biomaterials. To achieve this goal, model systems which could allow study of the formation and transformation of mineral phases within a hydrogel network with a range of experimental methods and high spatial and time resolution are needed. Here, we report a detailed investigation of the formation and transformation process of calcium phosphate mineral within an alginate hydrogel matrix. A combination of optical microscopy, confocal Raman microspectroscopy and electron microscopy was used to investigate the spatial distribution, morphology and crystal phase of the calcium phosphate mineral, as well as to study transformation of the mineral phases during the hydrogel mineralization process and upon incubation in a simulated body fluid. It was found, that under the conditions used in this work, mineral initially formed as a metastable amorphous calcium phosphate phase (ACP). The ACP particles had a distinctive spherical morphology and transformed within minutes into brushite in the presence of brushite seed crystals or into octacalcium phosphate, when no seeds were present in the hydrogel matrix. Incubation of brushite-alginate composites in simulated body fluid resulted in formation of hydroxyapatite. The characterization strategy presented here allows for non-destructive, in situ observation of mineralization processes in optically transparent hydrogels with little to no sample preparation. STATEMENT OF SIGNIFICANCE: The precipitation and transformations of calcium phosphates (CaP) is a complex process, where both formation kinetics and the stability of different mineral phases control the outcome. This situation is even more complex if CaP is precipitated in a hydrogel matrix, where one can expect the organic matrix to modulate crystallization by introducing supersaturation gradients or changing the nucleation and growth kinetics of crystals. In this study we apply a range of characterization techniques to study the mineral formation and transformations of CaP within an alginate matrix with spatiotemporal resolution. It demonstrates how a detailed investigation of the mineral precipitation and transformations can aid in the future rational design of hydrogel-based materials for bone tissue engineering and studies of biomineralization processes.

Efficient functionalization of alginate biomaterials.

Peptide coupled alginates obtained by chemical functionalization of alginates are commonly used as scaffold materials for cells in regenerative medicine and tissue engineering. We here present an alternative to the commonly used carbodiimide chemistry, using partial periodate oxidation followed by reductive amination. High and precise degrees of substitution were obtained with high reproducibility, and without formation of by-products. A protocol was established using l-Tyrosine methyl ester as a model compound and the non-toxic pic-BH3 as the reducing agent. DOSY was used to indirectly verify covalent binding and the structure of the product was further elucidated using NMR spectroscopy. The coupling efficiency was to some extent dependent on alginate composition, being most efficient on mannuronan. Three different bioactive peptide sequences (GRGDYP, GRGDSP and KHIFSDDSSE) were coupled to 8% periodate oxidized alginate resulting in degrees of substitution between 3.9 and 6.9%. Cell adhesion studies of mouse myoblasts (C2C12) and human dental stem cells (RP89) to gels containing various amounts of GRGDSP coupled alginate demonstrated the bioactivity of the material where RP89 cells needed higher peptide concentrations to adhere.