Encapsulated Stem Cells Research Articles

Alginate-Gelatin Microspheres Protect Human Mesenchymal Stem Cells During Deep Cryopreservation

Background: The need for on-demand biological products has been raised during the last decades. To prepare ready-to-use organ-related products, it necessitates a bulk cell reservoir. In this regard, stem cells, especially mesenchymal stem cells, have numerous therapeutic properties in tissue repair. Therefore, the advent of novel cell banking systems is inevitable to maintain cells ready-to-use. Objectives: In the current study, the cryoprotective effects of alginate-gelatin were investigated on human mesenchymal stem cells after seven days. Methods: Mesenchymal stem cells were classified into two groups; the Control and Encapsulated cells. Cells were encapsulated by using the mixture of alginate (1% v/v) and 2% gelatin and 1% CaCl2, a cross-linker, and vitrified in liquid nitrogen in freezing medium containing 10% dimethyl sulfoxide. After seven days, mesenchymal stem cells were thawed and decapsulated by 0.01 M sodium citrate solution. The cell survival rate was monitored by using MTT assay. Flow cytometry analysis of annexin-V/PI was used to determine the number of apoptotic cells. Results: These data showed that encapsulation had a superior effect on maintaining cell viability after the freeze-thaw procedure compared to the control group (P < 0.05). We found a significant decrease in the number of early- and late-stage apoptotic cells in mesenchymal stem cells inside the alginate-gelatin microspheres seven days after deep freezing (P < 0.05). As expected, cell cycle analysis specified the lack of dynamic activity in cells from both groups. Cells at phases S and G2/M reached zero. Conclusions: These findings showed that encapsulation of human mesenchymal stem cells with alginate-gelatin microspheres could reduce deep freeze/thaw insult and decrease the detrimental effect of cryoprotectant compared to non-capsulated cells.

Bioprinting stem cells: building physiological tissues one cell at a time.

Bioprinting aims to direct the spatial arrangement in three dimensions of cells, biomaterials, and growth factors. The biofabrication of clinically relevant constructs for the repair or modeling of either diseased or damaged tissues is rapidly advancing, resulting in the ability to three-dimensional (3D) print biomimetic platforms which imitate a large number of tissues in the human body. Primary tissue-specific cells are typically isolated from patients and used for the fabrication of 3D models for drug screening or tissue repair purposes. However, the lack of resilience of these platforms, due to the difficulties in harnessing, processing, and implanting patient-specific cells can limit regeneration ability. The printing of stem cells obviates these hurdles, producing functional in vitro models or implantable constructs. Advancements in biomaterial science are helping the development of inks suitable for the encapsulation and the printing of stem cells, promoting their functional growth and differentiation. This review specifically aims to investigate the most recent studies exploring innovative and functional approaches for the printing of 3D constructs to model disease or repair damaged tissues. Key concepts in tissue physiology are highlighted, reporting stem cell applications in biofabrication. Bioprinting technologies and biomaterial inks are listed and analyzed, including recent advancements in biomaterial design for bioprinting applications, commenting on the influence of biomaterial inks on the encapsulated stem cells. Ultimately, most recent successful efforts and clinical potentials for the manufacturing of functional physiological tissue substitutes are reported here, with a major focus on specific tissues, such as vasculature, heart, lung and airways, liver, bone and muscle.

Natural Multimerization Rules the Performance of Affinity-Based Physical Hydrogels for Stem Cell Encapsulation and Differentiation.

Tissue engineering and stem cell research greatly benefit from cell encapsulation within hydrogels as it promotes cell expansion and differentiation. Affinity-triggered hydrogels, an appealing solution for mild cell encapsulation, rely on selective interactions between the ligand and target and also on the multivalent presentation of these two components. Although these hydrogels represent a versatile option to generate dynamic, tunable, and highly functional materials, the design of hydrogel properties based on affinity and multivalency remains challenging and unstudied. Here, the avidin-biotin affinity pair, with the highest reported affinity constant, is used to address this challenge. It is demonstrated that the binding between the affinity hydrogel components is influenced by the multivalent display selected. In addition, the natural multivalency of the interaction must be obeyed to yield robust multicomponent synthetic protein hydrogels. The hydrogel's resistance to erosion depends on the right stoichiometric match between the hydrogel components. The developed affinity-triggered hydrogels are biocompatible and support encapsulation of induced pluripotent stem cells and their successful differentiation into a neural cell line. This principle can be generalized to other affinity pairs using multimeric proteins, yielding biomaterials with controlled performance.

Continuous microfluidic encapsulation of single mesenchymal stem cells using alginate microgels as injectable fillers for bone regeneration

The encapsulation of cells in microscale hydrogels can provide a mimic of a three-dimensional (3D) microenvironment to support cell viability and functions and to protect cells from the environmental stress, which have been widely used in tissue regeneration and cell therapies. Here, a microfluidics-based approach is developed for continuous encapsulation of mesenchymal stem cells (MSCs) at the single-cell level using alginate microgels. This microfluidic technique integrated on-chip encapsulation, gelation, and de-emulsification into a one-step fabrication process, which enables scalable cell encapsulation while retaining the viability and functionality of loaded cells. Remarkably, we observed MSCs encapsulated in Ca-alginate microgels at the single-cell level showed significantly enhanced osteogenesis and accelerated mineralization of the microgels which occurred only after 7 days of induction. Furthermore, MSCs laden in alginate microgels displayed significantly enhanced bone formation compared to MSCs mixed with microgels and acellular microgels in a rat tibial ablation model. To conclude, the current microfluidic technique represents a significant step toward continuous single cell encapsulation, fabrication, and purification. These microgels can boost bone regeneration by providing a controlled osteogenic microenvironment for encapsulated MSCs and facilitate stem cell therapy in the treatment of bone defects in a minimally invasive delivery way. Statement of SignificanceThe biological functions and therapeutic activities of single cells laden in microgels for tissue engineering remains less investigated. Here, we reported a microfluidic-based method for continuous encapsulation of single MSCs with high viability and functionality by integrating on-chip encapsulation, gelation, and de-emulsification into a one-step fabrication process. More importantly, MSCs encapsulated in alginate microgels at the single-cell level showed significantly enhanced osteogenesis, remarkably accelerated mineralization in vitro and bone formation capacity in vivo. Therefore, this single-cell encapsulation technique can facilitate stem cell therapy for bone regeneration and be potentially used in a variety of tissue engineering applications.

Main-Chain Liquid Crystalline Hydrogels that Support 3D Stem Cell Culture.

Hydrogels are frequently utilized as three-dimensional matrices for the culture and regeneration of soft tissues, but one challenge with the existing hydrogels is that, though the natural extracellular matrix of tissues may be ordered, there are few biocompatible ways to incorporate anisotropy within hydrogels. Liquid crystalline (LC) polymers are well suited for this because of their combination of molecular ordering and polymer elasticity; however, the hydrophobic nature of LC monomers has hindered their polymerization into hydrogels under cytocompatible conditions. This work reports on the generation of main-chain LC hydrogels in aqueous media and the ability of LC phases to affect mesenchymal stem cell behavior. The synthesis results in high gel fraction materials, and calorimetry, thermomechanical analysis, and X-ray scattering show that the networks organize into LC phases in the dry and hydrogel states. Human mesenchymal stem cells (hMSCs) cultured within the hydrogels show excellent viability, and hMSC proliferation proceeds at a faster rate in LC hydrogels compared to non-LC hydrogels. TThe result is a new synthetic approach for calamitic liquid crystalline hydrogels, which support the encapsulation and culture of human stem cells and are expected to enable applications as anisotropic and responsive substrates for tissue engineering and regenerative medicine.

Inverse Electron-Demand Diels-Alder Methylcellulose Hydrogels Enable the Co-delivery of Chondroitinase ABC and Neural Progenitor Cells.

A hydrogel that can deliver both proteins and cells enables the local microenvironment of transplanted cells to be manipulated with a single injection. Toward this goal, we designed a hydrogel suitable for the co-delivery of neural stem cells and chondroitinase ABC (ChABC), a potent enzyme that degrades the glial scar that forms after central nervous system (CNS) injury. We leveraged the inverse electron-demand Diels-Alder reaction between norbornene and methylphenyltetrazine to form rapidly gelling (<15 min) crosslinked methylcellulose (MC) hydrogels at physiological temperature and pH, with Young's modulus similar to that of brain tissue (1-3 kPa), and degradable, disulfide-containing crosslinkers. To achieve tunable, affinity-controlled release of a ChABC-Src homology 3 (SH3) fusion protein, we immobilized norbornene-functionalized SH3-binding peptides onto MC-methylphenyltetrazine and observed release of bioactive ChABC-SH3 over 4 days. We confirmed cytocompatibility by evaluating neural progenitor cell survival and proliferation. The combined encapsulation of neural stem cells and chondroitinase ABC from one hydrogel lays the framework for future in vivo studies to treat CNS injuries.

Emulsion-based encapsulation of pluripotent stem cells in hydrogel microspheres for cardiac differentiation.

Cardiovascular disease is the leading cause of death worldwide, and current treatments are ineffective or unavailable to majority of patients. Engineered cardiac tissue (ECT) is a promising treatment to restore function to the damaged myocardium; however, for these treatments to become a reality, tissue fabrication must be amenable to scalable production and be used in suspension culture. Here, we have developed a low-cost and scalable emulsion-based method for producing ECT microspheres from poly(ethylene glycol) (PEG)-fibrinogen encapsulated mouse embryonic stem cells (mESCs). Cell-laden microspheres were formed via water-in-oil emulsification; encapsulation occurred by suspending the cells in hydrogel precursor solution at cell densities from 5 to 60 million cells/ml, adding to mineral oil and vortexing. Microsphere diameters ranged from 30 to 570 μm; size variability was decreased by the addition of 2% poly(ethylene glycol) diacrylate. Initial cell encapsulation density impacted the ability for mESCs to grow and differentiate, with the greatest success occurring at higher cell densities. Microspheres differentiated into dense spheroidal ECTs with spontaneous contractions occurring as early as Day 10 of cardiac differentiation; furthermore, these ECT microspheres exhibited appropriate temporal changes in gene expression and response to pharmacological stimuli. These results demonstrate the ability to use an emulsion approach to encapsulate pluripotent stem cells for use in microsphere-based cardiac differentiation.

Single-Cell Encapsulation via Click-Chemistry Alters Production of Paracrine Factors from Neural Progenitor Cells.

Extracellular matrix (ECM) properties affect multiple cellular processes such as cell survival, proliferation, and protein synthesis. Thus, a polymeric‐cell delivery system with the ability to manipulate the extracellular environment can act as a fundamental regulator of cell function. Given the promise of stem cell therapeutics, a method to uniformly enhance stem cell function, in particular trophic factor release, can prove transformative in improving efficacy and increasing feasibility by reducing the total number of cells required. Herein, a click‐chemistry powered 3D, single‐cell encapsulation method aimed at synthesizing a polymeric coating with the optimal thickness around neural progenitor cells is introduced. Polymer encapsulation of neural stem cells significantly increases the release of neurotrophic factors such as VEGF and CNTF. Cell encapsulation with a soft extracellular polymer upregulates the ADCY8‐cAMP pathway, suggesting a mechanism for the increase in paracrine factors. Hence, the described single‐cell encapsulation technique can emerge as a translatable, nonviral cell modulation method and has the potential to improve stem cells' therapeutic effect.

Droplet Microfluidics for the ex Vivo Expansion of Human Primary Multiple Myeloma Cells.

We previously reported a new approach for micromanipulation and encapsulation of human stem cells using a droplet-based microfluidic device We demonstrated the possibility of encapsulating and culturing difficult-to-preserve primary human hematopoietic stem cells using an engineered double layered bead composed by an inner layer of alginate and an outer layer of puramatrix constructed using a soft technology without the use of any external force. In this work, we use this micro manipulation technique to build a 3D scaffold as a biomimetic model to recapitulate the niche of patient-derived multiple myeloma cells (MM cell) using a multilayered 3D tissue scaffold constructed in a microfluidic device and cultured in 10% FBS culture medium. In the current study, we included the use of this biomimetic model comprising supporting human Mesenchymal stem cells to show the mid-term survival of MM cells in the proposed structures. We found that the generated microniches were suitable for the maintenance of MM cells with and without supporting cells. Additionally, cultured MM cells in droplets were exposed to both Bortezomib and Lenalidomide to test their toxicity in the cultured patient derived cells. Results indicate that the maintained MM cells were consistently responding to the applied medication, opening a wide field of possibilities to use the presented micro device as an ex vivo platform for drug screening.

Continuous Microfluidic Encapsulation of Single Mesenchymal Stem Cells Using Alginate Microgels as Injectable Fillers for Bone Regeneration

Encapsulation of single cell in microgels in a biocompatible and scalable manner can enable major advances in applications such as tissue engineering, cell biology, and cell-based therapies. Here, a microfluidics-based approach is developed for continuous encapsulation of mesenchymal stem cells (MSCs) at single-cell level using alginate as the extracellular matrix. This microfluidic technique enables on-chip encapsulation by triggered gelation and direct extraction of cell-laden microgels from oil into aqueous phase, thus retaining the viability and functionality of the loaded cells. Osteogenic culture of MSCs in Ca-alginate microgels show significantly accelerated osteogenesis compared to conventional 2D culture. It is further demonstrated that microfludics-generated alginate microgels can act as delivery vehicles of allogeneic rat MSCs for inducing bone regeneration using the rat tibial ablation model. A significantly higher amount of new bone formation is observed in the bone medullary cavity treated by MSC-laden microgels as relevant to naked MSCs with microgels and empty microgels. To conclude, single cell-laden microgels boost therapeutic effects of bone regeneration compared to conventional cell-based tissue engineering therapy by providing a controlled osteogenic and immune-shielding microenvironment for allogeneic MSCs.

Design of an Adhesive Film-Based Microfluidic Device for Alginate Hydrogel-Based Cell Encapsulation.

To support the increasing translational use of transplanted cells, there is a need for high-throughput cell encapsulation technologies. Microfluidics is a particularly promising candidate technology to address this need, but conventional polydimethylsiloxane devices have encountered challenges that have limited their utility, including clogging, leaking, material swelling, high cost, and limited scalability. Here, we use a rapid prototyping approach incorporating patterned adhesive thin films to develop a reusable microfluidic device that can produce alginate hydrogel microbeads with high-throughput potential for microencapsulation applications. We show that beads formed in our device have high sphericity and monodispersity. We use the system to demonstrate effective cell encapsulation of mesenchymal stem cells and show that they can be maintained in culture for at least 28days with no measurable reduction in viability. Our approach is highly scalable and will support diverse translational applications of microencapsulated cells.

Reversible Dynamic Mechanics of Hydrogels for Regulation of Cellular Behavior

The mechanical properties of the native extracellular matrix play a key role in regulating cell behavior during developmental, healing and homeostatic processes. Since these properties change over time, it may be valuable to have the capacity to dynamically vary the mechanical properties of engineered hydrogels used in tissue engineering strategies to better mimic the dynamics mechanical behavior of native extracellular matrix. However, in situ repeatedly reversible dynamic tuning of hydrogel mechanics is still limited. In this study, we have engineered a hydrogel system with reversible dynamic mechanics using a dual-crosslinkable alginate hydrogel. The effect of reversible mechanical signals on encapsulated stem cells in dynamically tunable hydrogels has been demonstrated. In situ stiffening of hydrogels decreases cell spreading and proliferation, and subsequent softening of hydrogels gives way to an increase in cell spreading and proliferation. The hydrogel stiffening and softening, and resulting cellular responses are repeatedly reversible. This hydrogel system provides a promising platform for investigating the effect of repeatedly reversible changes in extracellular matrix mechanics on cell behaviors.

Alginate-encapsulated brain-derived neurotrophic factor–overexpressing mesenchymal stem cells are a promising drug delivery system for protection of auditory neurons

The cochlear implant outcome is possibly improved by brain-derived neurotrophic factor treatment protecting spiral ganglion neurons. Implantation of genetically modified mesenchymal stem cells may enable the required long-term brain-derived neurotrophic factor administration. Encapsulation of mesenchymal stem cells in ultra-high viscous alginate may protect the mesenchymal stem cells from the recipient’s immune system and prevent their uncontrolled migration. Alginate stability and survival of mesenchymal stem cells in alginate were evaluated. Brain-derived neurotrophic factor production was measured and its protective effect was analyzed in dissociated rat spiral ganglion neuron co-culture. Since the cochlear implant is an active electrode, alginate–mesenchymal stem cell samples were electrically stimulated and alginate stability and mesenchymal stem cell survival were investigated. Stability of ultra-high viscous-alginate and alginate–mesenchymal stem cells was proven. Brain-derived neurotrophic factor production was detectable and spiral ganglion neuron survival, bipolar morphology, and neurite outgrowth were increased. Moderate electrical stimulation did not affect the mesenchymal stem cell survival and their viability was good within the investigated time frame. Local drug delivery by ultra-high viscous-alginate-encapsulated brain-derived neurotrophic factor–overexpressing mesenchymal stem cells is a promising strategy to improve the cochlear implant outcome.

Molecular recognition-directed site-specific release of stem cell differentiation inducers for enhanced joint repair

The remarkable difference in cell type and matrix composition between two connected parts of a joint (cartilage and subchondral bone) makes it challenging to simultaneously regenerate both parts for joint repair. Thus we chemically designed a biphasic hydrogel made of two well-bonded shape-tunable hydrogel phases, termed bone-regenerating hydrogel (BRH) and cartilage-regenerating hydrogel (CRH). The BRH and CRH, encapsulating stem cells, were produced by photo-crosslinking bone and cartilage matrix-mimicking biopolymers and a nanobox (β-cyclodextrin) in situ in the subchondral bone defect and cartilage defect, respectively. The nanoboxes in BRH and CRH were loaded with osteogenic and chondrogenic differentiation inducers (melatonin and kartogenin) by host−guest interactions, respectively. Such interactions directed the sustained phase- and defect site-specific release of the inducers and subsequent site-specific stem cell differentiation into cartilage and bone forming cells for joint repair. The strategy opens up a new chemical approach to biomaterials with phase-specific drug release for tissue repair.

Covalently injectable chitosan/chondroitin sulfate hydrogel integrated gelatin/heparin microspheres for soft tissue engineering

Microspheres and injectable hydrogels derived from natural biopolymers have been extensively investigated as drug carriers and cell scaffolds. In this study, we report a preparation of composite scaffolds basing microspheres and hydrogel via the Schiff’s base reaction. Hybrid gelatin/heparin microspheres loading insulin-like growth factor-1 (IGF-1) with a diameter of 5–10 µm were fabricated using an emulsion cross-linking method. Synchronously, water-soluble carboxymethyl chitosan (CMC) and oxidized chondroitin sulfate (OCS) were prepared for cross-linking of hydrogels, which were embedded with microspheres to produce a composite microspheres/gel scaffold. The mechanism of scaffold cross-linking is attributed to the Schiff’s base reaction between amino and aldehyde groups of biopolymers. Currently, gelation rate, morphology, mechanical properties, swelling ratio, weight loss, and IGF-1 release of the composite scaffolds were examined in vitro. The results show that mechanical and bioactive properties of CMC-OCS hydrogel can be significantly improved by embedding gelatin/heparin microspheres containing IGF-1. Compressive modulus of composite gel scaffolds containing 3 wt% of microspheres was 16 kPa, which was higher than the control hydrogel without microspheres. Cumulative release of IGF-1 during 7 days from microspheres embedded hydrogel was 70%, which was significantly lower than those of microspheres and hydrogels. Moreover, the composite microspheres/gel scaffolds exhibited higher swelling ratio and slower degradation rate than the control. Potential of the composite scaffolds was demonstrated by encapsulation of human adipose-derived stem cells (ASCs) in vitro. Cell culture showed that this composite hydrogel could support survival and proliferation of ASCs. These results demonstrate the potential of gelatin/heparin microspheres embedded CMC-OCS hydrogels as an injectable scaffold in soft tissue engineering.

Bioactive glass-containing hydrogel delivery system for osteogenic differentiation of human dental pulp stem cells.

Osteoconductive hydrogels can be fabricated by incorporating necessary growth factors and bioactive particles or simply utilizing the ability of the hydrogel itself to induce bone regeneration. The osteogenic inductive potential of the bioactive glass microparticles (BG MPs) has been well-studied. However, the role of the hydrogel embedding the BG MPs on the osteogenic differentiation of the encapsulated stem cells has not been well established. Moreover, it has been reported that the dental pulp stem cell (DPSC) has the capability of regenerating the craniofacial bone tissue when receiving the necessary osteogenic signals from the microenvironment. In the current study, we have fabricated a composite hydrogel based on alginate and Matrigel containing BG MPs and evaluated the role of the BG MPs and Matrigel on osteogenic differentiation of the encapsulated DPSCs. Our results confirmed that presence of Matrigel enhances the osteogenic differentiation of the DPSCs regardless of the decrease in elasticity of the hydrogel in presence of the BG MPs. This strategy of modifying the microenvironment can be a promising approach for craniofacial bone tissue engineering.

Chondroprotective and osteogenic effects of silk-based bioinks in developing 3D bioprinted osteochondral interface

Attributing cell instructive features and multifunctionality to biological inks (bioinks) employed for three-dimensional (3D) printing strategies is very much essential to bring about a paradigm shift in developing next generation smart intuitive 3D bioprinted constructs. Giving perspective to this notion, we explore here the feasibilities in developing multifunctional silk-based cartilage and bone bioinks for recreating heterogeneous complicated tissue constructs such as the osteochondral interface. In this regard, the developed silk based bioinks exhibit shear thinning behaviour with quick thixotropic recovery (~90% recovery) aiding in printing self-standing structures with decent print fidelity. The hydrogel network within the 3D bioprinted constructs present good permeability enabling in forming an undulating demarcation region at the bioprinted osteochondral interface. Furthermore, the cartilage and bone inks used for the microextrusion based bioprinting of osteochondral constructs facilitate the spatial maturation and differentiation of encapsulated stem cells towards osteogenic and chondrogenic lineages. The incorporation of strontium doped nano-apatites activates hypoxia inducible factor (HIF-1α) related genes, conferring proangiogenic and chondroprotective traits to the bioinks. Involvement of strontium in down regulating cyclooxygenase-2 via inhibiting prostaglandins (PGE2) pathway enabled anti-osteoclastic activity while favouring M2 macrophage biasness. Altogether, these findings corroborate the potential of the developed nanocomposite bioinks for fabricating clinically viable grafts for osteochondral defect repair associated with osteoporosis.

Injectable supramolecular polymer-nanoparticle hydrogels enhance human mesenchymal stem cell delivery.

Stem cell therapies have emerged as promising treatments for injuries and diseases in regenerative medicine. Yet, delivering stem cells therapeutically can be complicated by invasive administration techniques, heterogeneity in the injection media, and/or poor cell retention at the injection site. Despite these issues, traditional administration protocols using bolus injections in a saline solution or surgical implants of cell‐laden hydrogels have highlighted the promise of cell administration as a treatment strategy. To address these limitations, we have designed an injectable polymer–nanoparticle (PNP) hydrogel platform exploiting multivalent, noncovalent interactions between modified biopolymers and biodegradable nanoparticles for encapsulation and delivery of human mesenchymal stem cells (hMSCs). hMSC‐based therapies have shown promise due to their broad differentiation capacities and production of therapeutic paracrine signaling molecules. In this work, the fundamental hydrogel mechanical properties that enhance hMSC delivery processes are elucidated using basic in vitro models. Further, in vivo studies in immunocompetent mice reveal that PNP hydrogels enhance hMSC retention at the injection site and retain administered hMSCs locally for upwards of 2 weeks. Through both in vitro and in vivo experiments, we demonstrate a novel scalable, synthetic, and biodegradable hydrogel system that overcomes current limitations and enables effective cell delivery.

Reversible Thermoresponsive Hydrogel Fabricated from Natural Biopolymer for the Improvement of Critical Limb Ischemia by Controlling Release of Stem Cells.

Stem cells therapy is an effective treatment for critical limb ischemia diseases (CLI), but is limited to low cells retention and poor target release in severe ischemia tissues. Due to the notable feature of CLI, namely, the temperature of ischemia tissues decreases with the severity of the lesions, a thermoresponsive and reversible hydrogel based on methylcellulose-salt system encapsulating stem cells is facilely prepared and successfully achieved the goal of releasing stem cells in lower temperature areas. The investigations show that the thermogel presents notable biocompatibility, thermoresponsiveness, and cytoprotection. Furthermore, the combined transplantation of hydrogel and stem cells system effectively inhibits the fibrosis and muscular atrophy of lower limb ischemia, accelerates the recovery of lower limb blood flow, and promotes angiogenesis, indicating that the reversible thermogel can promote vascular repair by controlling the release of loaded stem cells in the treatment of CLI.

First Aldol Cross-Linked Hyaluronic Acid Hydrogel: Fast and Hydrolytically Stable Hydrogel with Tissue Adhesive Properties

Currently, there are limited approaches to tailor 3D scaffolds cross-linked with a stable covalent C-C bond that does not require any catalysts or initiators. We present here the first hydrogels employing aldol condensation chemistry that exhibit exceptional physicochemical properties. We investigated the aldol-cross-linking chemistry using two types of aldehyde-modified hyaluronic acid (HA) derivatives, namely, an enolizable HA-aldehyde (HA-Eal) and a non-enolizable HA-aldehyde (HA-Nal). Hydrogels formed using HA-Eal demonstrate inferior cross-linking efficiency (due to intramolecular loop formation), when compared with hydrogels formed by mixing HA-Eal and HA-NaI leading to a cross-aldol product. The change in mechanical properties as a result of cross-linking at different pH values is determined using rheological measurements and is interpreted in terms of molecular weight between cross-links (Mc). The novel HA cross-aldol hydrogel demonstrate excellent hydrolytic stability and favorable mechanical properties but allow hyaluronidase-mediated enzymatic degradation. Interestingly, residual aldehyde functionality within the aldol product rendered the tissue-adhesive properties by bonding two bone tissues. The aldehyde functionality also facilitated facile post-synthetic modifications with nucleophilic reagents. Finally, we demonstrate that the novel hydrogel is biocompatible with encapsulated stem cells that show a linear rate of expansion in our 3-6 days of study.