Injectable Microgels Research Articles

Injectable dECM-enhanced hyaluronic microgels with spatiotemporal release of cartilage-specific molecules to improve osteoarthritic chondrocyte’s function

The interior environment of articular cartilage in osteoarthritis (OA) presents substantial hurdles, leading to the malfunction of chondrocytes and the breakdown of collagen II-enriched hyaline cartilage matrix. Despite this, most clinical treatments primarily provide temporary relief from OA discomfort without arresting OA progression. This study aimed to alleviate OA by developing intra-articular injectable dECM-enhanced hyaluronic (HE) microgels. The HE hydrogel was engineered and shaped into uniformly sized microgels using microfluidics and photopolymerization techniques. These microgels provided a spatiotemporal cascade effect, facilitating the rapid release of growth factors and a slower release of ECM macromolecules and proteins. This process assisted in the recovery of OA chondrocytes’ function, promoting cell proliferation, matrix synthesis, and cartilage-specific gene expression in vitro. It also effectively aided repair of the collagen II-enriched hyaline cartilage and significantly reduced the severity of OA, as demonstrated by radiological observation, gross appearance, histological/immunohistochemical staining, and analysis in an OA rat model in vivo. Collectively, the HE injectable microgels with spatiotemporal release of cartilage-specific molecules have shown promise as a potential candidate for a cell-free OA therapy approach.Graphical

ROS-responsive drug-releasing injectable microgels for ameliorating myocardial infarction

Despite of the recent advances in regulatory T cell (Treg) therapy, a limited number of available cells and specificity at the desired tissue site have severely compromised their efficacy. Herein, an injectable drug-releasing (MTK-TK-drug) microgel system in response to in situ stimulation by reactive oxygen species (ROS) was constructed with a coaxial capillary microfluidic system and UV curing. The spherical microgels with a size of 150 μm were obtained. The MTK-TK-drug microgels efficiently converted the pro-inflammatory Th17 cells into anti-inflammatory regulatory T cells (Treg) cells in vitro, and the ROS-scavenging materials synergistically enhanced the effect by modulating the inflammation microenvironment. Thus, the microgels significantly reduced cardiomyocyte apoptosis and decreased the inflammatory response in the early stages of post-myocardial infarction (MI) in vivo, thereby reducing fibrosis, promoting vascularization, and preserving cardiac function. Overall, our results indicate that the MTK-TK-drug microgels can attenuate the inflammatory response and improve MI therapeutic effects in vivo.

In vitro development and optimization of cell-laden injectable bioprinted gelatin methacryloyl (GelMA) microgels mineralized on the nanoscale

Bone defects may occur in different sizes and shapes due to trauma, infections, and cancer resection. Autografts are still considered the primary treatment choice for bone regeneration. However, they are hard to source and often create donor-site morbidity. Injectable microgels have attracted much attention in tissue engineering and regenerative medicine due to their ability to replace inert implants with a minimally invasive delivery. Here, we developed novel cell-laden bioprinted gelatin methacrylate (GelMA) injectable microgels, with controllable shapes and sizes that can be controllably mineralized on the nanoscale, while stimulating the response of cells embedded within the matrix. The injectable microgels were mineralized using a calcium and phosphate-rich medium that resulted in nanoscale crystalline hydroxyapatite deposition and increased stiffness within the crosslinked matrix of bioprinted GelMA microparticles. Next, we studied the effect of mineralization in osteocytes, a key bone homeostasis regulator. Viability stains showed that osteocytes were maintained at 98 % viability after mineralization with elevated expression of sclerostin in mineralized compared to non-mineralized microgels, showing that mineralization can effectively enhances osteocyte maturation. Based on our findings, bioprinted mineralized GelMA microgels appear to be an efficient material to approximate the bone microarchitecture and composition with desirable control of sample injectability and polymerization. These bone-like bioprinted mineralized biomaterials are exciting platforms for potential minimally invasive translational methods in bone regenerative therapies.

Injectable Microgels with Hybrid Exosomes of Chondrocyte-Targeted FGF18 Gene-Editing and Self-Renewable Lubrication for Osteoarthritis Therapy.

Abnormal silencing of fibroblast growth factor (FGF) signaling significantly contributes to joint dysplasia and osteoarthritis (OA); However, the clinical translation of FGF18-based protein drugs is hindered by their short half-life, low delivery efficiency and the need for repeated articular injections. This study proposes a CRISPR/Cas9-based approach to effectively activate the FGF18 gene of OA chondrocytes at the genome level in vivo, using chondrocyte-affinity peptide (CAP) incorporated hybrid exosomes (CAP/FGF18-hyEXO) loaded with an FGF18-targeted gene-editing tool. Furthermore, CAP/FGF18-hyEXO are encapsulated in methacrylic anhydride-modified hyaluronic (HAMA) hydrogel microspheres via microfluidics and photopolymerization to create an injectable microgel system (CAP/FGF18-hyEXO@HMs) with self-renewable hydration layers to provide persistent lubrication in response to frictional wear. Together, the injectable CAP/FGF18-hyEXO@HMs, combined with in vivo FGF18 gene editing and continuous lubrication, have demonstrated their capacity to synergistically promote cartilage regeneration, decrease inflammation, and prevent ECM degradation both in vitro and in vivo, holding great potential for clinical translation.

High‐Throughput Bioprinting of Geometrically‐Controlled Pre‐Vascularized Injectable Microgels for Accelerated Tissue Regeneration (Adv. Healthcare Mater. 22/2023)

High‐Throughput Bioprinting of Geometrically‐Controlled Pre‐Vascularized Injectable Microgels for Accelerated Tissue Regeneration (Adv. Healthcare Mater. 22/2023)

Galactose-functionalized injectable thermosensitive microgels for target specific uptake in hepatoma cells.

Microgels acts as a potential candidate for responsive composite materials, which has been favored by scientists because of its excellent colloid stability, easy integration, and most of their surface area can be used as support after modification. Specififically, microgels are fascinating capable of maintaining good biocompatibility and controlled-release in vivo and making the possible for applications in biomaterials and biomedicines. Besides, in the process of microgel synthesis, some targeting factors can be combined to achieve the purpose of cell targeting and uptake. Therefore, how to fundamentally design microgels is an urgent problem to be solved. In this study, we design and synthesize an injectable microgel P(DEGMA-co-OVNGal) that is made of 2-methyl-2-acrylate-2-(2-methoxy ethoxy) ethyl ester (DEGMA) and glycopolymer (OVNGal) that is thermoresponsive and contains galactose. When the content of crosslinking agent is regulated, the microgel will realize the transformation from sol to gel at the temperature around human body, causing the controlled release of the loaded drugs. With the increase of crosslinker content from 1% to 7%, the appearance of microgel changed from loose and ordered to compact and hard morphology, the swelling ratio of microgel decreased from 187% to 142%, and the phase volume transition temperature decreased from 29.2°C to 28°C. The results indicated that when the monomer ratio (DEGMA: OVNGal) increased from 2:1 to 40:1 with the amount of crosslinking agent at 1%, the particle size of microgel increased from 460nm to 660nm. In vitro released studies confirmed that the cumulative released DOX (doxorubicin, as a selected model drug) from the microgel can reach 50% after 7days. Furthermore, in vitro experiments demonstrated that the injectable microgel P(DEGMA-co-OVNGal) can target HepG2 cells effectively and meantime dispalys excellent biocompatibility. Therefore, the injected microgels based on P(DEGMA-co-OVNGal) have the potential to serve as a robust and promising drug delivery carrier for targeted cancer therapy.

High-Throughput Bioprinting of Geometrically-Controlled Pre-Vascularized Injectable Microgels for Accelerated Tissue Regeneration.

Successful integration of cell-laden tissue constructs with host vasculature depends on the presence of functional capillaries to provide oxygen and nutrients to the embedded cells. However, diffusion limitations of cell-laden biomaterials challenge regeneration of large tissue defects that require bulk-delivery of hydrogels and cells. Herein, a strategy to bioprint geometrically controlled, endothelial and stem-cell laden microgels in high-throughput is introduced, allowing these cells to form mature and functional pericyte-supported vascular capillaries in vitro, and then injecting these pre-vascularized constructs minimally invasively in-vivo. It is demonstrated that this approach offers both desired scalability for translational applications as well as unprecedented levels of control over multiple microgel parameters to design spatially-tailored microenvironments for better scaffold functionality and vasculature formation. As a proof-of-concept, the regenerative capacity of the bioprinted pre-vascularized microgels is compared with that of cell-laden monolithic hydrogels of the same cellular and matrix composition in hard-to-heal defects in vivo. The results demonstrate that the bioprinted microgels have faster and higher connective tissue formation, more vessels per area, and widespread presence of functional chimeric (human and murine) vascular capillaries across regenerated sites. The proposed strategy, therefore, addresses a significant issue in regenerative medicine, demonstrating a superior potential to facilitate translational regenerative efforts.

Swelling-Mediated Mechanical Stimulation Regulates Differentiation of Adipose-Derived Mesenchymal Stem Cells for Intervertebral Disc Repair Using Injectable UCST Microgels.

Mechanical stimulation is an effective approach for controlling stem cell differentiation in tissue engineering. However, its realization in in vivo tissue repair remains challenging since this type of stimulation can hardly be applied to injectable seeding systems. Here, it is presented that swelling of injectable microgels can be transformed to in situ mechanical stimulation via stretching the cells adhered on their surface. Poly(acrylamide-co-acrylic acid) microgels with the upper critical solution temperature property are fabricated using inverse emulsion polymerization and further coated with polydopamine to increase cell adhesion. Adipose-derived mesenchymal stem cells (ADSCs) adhered on the microgels can be omnidirectionally stretched along with the responsive swelling of the microgels, which upregulate TRPV4and Piezo1channel proteins and enhance nucleus pulposus (NP)-like differentiation of ADSCs. In vivo experiments reveal that the disc height and extracellular matrix content of NP are promoted after the implantation with the microgels. The findings indicate that swelling-induced mechanical stimulation has great potential for regulating stem cell differentiation during intervertebral disc repair.

Comparison of Four Different Preparation Methods for Making Injectable Microgels for Tissue Engineering and Cell Therapy

Comparison of Four Different Preparation Methods for Making Injectable Microgels for Tissue Engineering and Cell Therapy

Injectable and Self-healing microgels repairing infected irregular tubular fistula in anal and muscle

An anal fistula caused by a perianal abscess is a hollow tubular structure connecting anal canal with skin and often in irregular shapes. The surgical treatment causes damage to the surrounding tissues and results in complications. Current filling plugs in the clinic, derived from the porcine small intestinal submucosa, only seal the opening and cannot fill into the irregular fistula passage due to the poor liquidity, and fails to control the high-frequent infections around fistulas. Injecting gel-like material is a direct and effective way. To this end, we reported injectable microgels from elastic cryogels of biodegradable disulfide-based crosslinking. Microgels with shear-thinning and self-healing remain solid under low strain and fluid under high strain to ensure successful filling and repair the multiple-branched irregular canals. Nano-silver dopped microgels presented antibacterial efficacy. In rabbit models of anal and muscle fistula, the inflammatory cells significantly decreased. CD31 and α-smooth muscle actin (α-SMA) showed the fistula healing was improved considerably. These findings demonstrated that self-healable microgels have enormous potential to repair the abnormal and complicated tubular passages caused by ulcers and fistulas.

Hydrolytically Degradable Microgels with Tunable Mechanical Properties Modulate the Host Immune Response.

Hydrogel microparticles (microgels) are an attractive approach for therapeutic delivery because of their modularity, injectability, and enhanced integration with the host tissue. Multiple microgel fabrication strategies and chemistries have been implemented, yet manipulation of microgel degradability and its effect on in vivo tissue responses remains underexplored. Here, the authors report a facile method to synthesize microgels crosslinked with ester-containing junctions to afford tunable degradation kinetics. Monodisperse microgels of maleimide-functionalized poly(ethylene-glycol) are generated using droplet microfluidics crosslinked with thiol-terminated, ester-containing molecules. Tunable mechanics are achievable based on the ratio of degradable to nondegradable crosslinkers in the continuous phase. Degradation in an aqueous medium leads to microgel deformation based on swelling and a decrease in elastic modulus. Furthermore, degradation byproducts are cytocompatible and do not cause monocytic cell activation under noninflammatory conditions. These injectable microgels possess time-dependent degradation on the order of weeks in vivo. Lastly, the evaluation of tissue responses in a subcutaneous dorsal pocket shows a dynamic type-1 like immune response to the synthetic microgels, driven by interferon gamma (IFN-γ ) expression, which can be moderated by tuning the degradation properties. Collectively, this study demonstrates the development of a hydrolytic microgel platform that can be adapted to desired host tissue immune responses.

Stem Cell‐Recruiting Injectable Microgels for Repairing Osteoarthritis (Adv. Funct. Mater. 48/2021)

Injectable Microgels Inspired by the phenomenon where islands could recruit seabirds for nesting, in article number 2105084, Xiaoji Luo, Wenguo Cui, Wei Huang, and co-workers present an injectable porous microgel generated via microfluidic technology that is non-covalently loaded with PDGF-BB and TGF-β3, forming “cell island” microgels that can serve as a temporary “nest site” to recruit endogenous stem cells and presents a promising therapeutic effect for osteoarthritic cartilage damage.

Self-assembled low-molecular-weight gelator injectable microgel beads for delivery of bioactive agents.

We report the preparation of hybrid self-assembled microgel beads by combining the low molecular weight gelator (LMWG) DBS-CONHNH2 and the natural polysaccharide calcium alginate polymer gelator (PG). Microgel formulations based on LMWGs are extremely rare due to the fragility of the self-assembled networks and the difficulty of retaining any imposed shape. Our hybrid beads contain interpenetrated LMWG and PG networks, and are obtained by an emulsion method, allowing the preparation of spherical gel particles of controllable sizes with diameters in the mm or μm range. Microgels based on LMWG/alginate can be easily prepared with reproducible diameters <1 μm (ca. 800 nm). They are stable in water at room temperature for many months, and survive injection through a syringe. The rapid assembly of the LMWG on cooling plays an active role in helping control the diameter of the microgel beads. These LMWG microbeads retained the ability of the parent gel to deliver the bioactive molecule heparin, and in cell culture medium this enhanced the growth of human mesenchymal stem cells. Such microgels may therefore have future applications in tissue repair. This approach to fabricating LMWG microgels is a platform technology, which could potentially be applied to a variety of different functional LMWGs, and hence has wide-ranging potential.

Sustained release of dermal papilla-derived extracellular vesicles from injectable microgel promotes hair growth.

Hair regeneration has long captured researchers' attention because alopecia is a common condition and current therapeutic approaches have significant limitations. Dermal papilla (DP) cells serve as a signaling center in hair follicles and regulate hair formation and cycling by paracrine secretion. Secreted EVs are important signaling mediators for intercellular communication, and DP-derived extracellular vesicles (DP-EVs) may play an important role in hair regeneration. However, the instability of EVs in vivo and their low long-term retention after transplantation hinder their use in clinical applications.Methods: Human DP-EVs were encapsulated in partially oxidized sodium alginate (OSA) hydrogels, yielding OSA-encapsulated EVs (OSA-EVs), which act as a sustained-release system to increase the potential therapeutic effect of DP-EVs. The ability of the OSA-EVs to protect protein was assessed. The hair regeneration capacity of OSA-EVs, as well as the underlying mechanism, was explored in hair organ culture and a mouse model of depilation.Results: The OSA-EVs were approximately 100 μm in diameter, and as the hydrogel degraded, DP-EVs were gradually released. In addition, the hydrogel markedly increased the stability of vesicular proteins and increased the retention of EVs in vitro and in vivo. The OSA-EVs significantly facilitated proliferation of hair matrix cells, prolonged anagen phase in cultured human hairs, and accelerated the regrowth of back hair in mice after depilation. These effects may be due to upregulation of hair growth-promoting signaling molecules such as Wnt3a and β-catenin, and downregulation of inhibitory molecule BMP2.Conclusion: This study demonstrated that OSA hydrogels promote the therapeutic effects of DP-EVs, and indicate that our novel OSA-EVs could be used to treat alopecia.

Intra-articular Treatment of Osteoarthritis with Diclofenac-Conjugated Polymer Reduces Inflammation and Pain.

The most common treatment for osteoarthritis is daily oral administration of a nonsteroidal anti-inflammatory drug such as diclofenac. This daily dosage regime is often associated with severe side effects. In this study, we explored the potential of utilizing a high molecular weight cross-linked polyurethane polymer covalently linked to diclofenac (C-DCF-PU) for intra-articular administration. We aim to exploit the advantages of local drug delivery by developing an implant with improved efficacy and reduced side effects. The polymer was synthesized from a diclofenac-functionalized monomer unit in a simple one-pot reaction, followed by cross-linking. In vitro drug release studies showed zero-order drug release for 4 days, followed by a gradual decline in drug release rate until diclofenac was depleted after 15 days. The cross-linked polymer was triturated to yield an injectable microgel formulation for administration. Whole animal fluorescence imaging of the rhodamine-labeled C-DCF-RH-PU showed good retention of the polymer in the knee joints of healthy rats, with approximately 30% of the injected dose still present 2 weeks post intra-articular administration. In a reactivation arthritis animal model, the C-DCF-RH-PU formulation reduced pain and significantly reduced inflammation after a short lag phase, showing that this drug delivery system warrants further development for long-term treatment of osteoarthritis with the benefit of reduced side effects.

3D Printing Enabled Customization of Functional Microgels.

Injectable microgels show great promising applications in cell therapy and drug delivery. Currently, there remains a challenge to rapidly and cost-effectively fabricate customized microgels. Here, we present a digital light processing based three-dimensional (3D) printing process to fabricate microgels with tailored shapes and sizes. The microgels are constructed by the digital light controlled polymerization of photopolymerizable monomer solution within 2 s. By mixing nanoparticle-encapsulated drugs into the monomer solution, the microgels with sustained drug release can be readily prepared. Also, cells can be printed into microgels with survival and proliferation. In conclusion, this study provides a 3D printing process for customizing functional microgels containing drugs or cells with potential therapeutic applications.

Injectable microgels development for sustained GALNS enzyme replacement therapy for Morquio syndrome type A

Injectable microgels development for sustained GALNS enzyme replacement therapy for Morquio syndrome type A

Chitosan-gelatin-based microgel for sustained drug delivery

We describe here the proof-of-concept of synthesizing microgels via cross-linking of chitosan and gelatin with succinimide-end polyethylene glycol (PEG). The introduction of PEG provided thermo-responsiveness to microgels, and the size of microgels significantly changed with increasing temperature. Furthermore, the microgel was biocompatible as confirmed in vitro by MTT assay with NIH 3T3 fibroblast and degradable. Chitosan-based microgels exhibited strong affinity to hydrophobic drug and prolonged release of folic acid was observed at physiological temperature with varying pH. The proposed cost-effective injectable microgels have the potential to serve as the delivery vehicle of hydrophobic bioactive molecules.

Uncoupling angiogenesis and inflammation in peripheral artery disease with therapeutic peptide-loaded microgels

Peripheral artery disease (PAD) is characterized by vessel occlusion and ischemia in the limbs. Treatment for PAD with surgical interventions has been showing limited success. Moreover, recent clinical trials with treatment of angiogenic growth factors proved ineffective as increased angiogenesis triggered severe inflammation in a proportionally coupled fashion. Hence, the overarching goal of this research was to address this issue by developing a biomaterial system that enables controlled, dual delivery of pro-angiogenic C16 and anti-inflammatory Ac-SDKP peptides in a minimally-invasive way. To achieve the goal, a peptide-loaded injectable microgel system was developed and tested in a mouse model of PAD. When delivered through multiple, low volume injections, the combination of C16 and Ac-SDKP peptides promoted angiogenesis, muscle regeneration, and perfusion recovery, while minimizing detrimental inflammation. Additionally, this peptide combination regulated inflammatory TNF-α pathways independently of MMP-9 mediated pathways of angiogenesis in vitro, suggesting a potential mechanism by which angiogenic and inflammatory responses can be uncoupled in the context of PAD. This study demonstrates a translatable potential of the dual peptide-loaded injectable microgel system for PAD treatment.

Injectable multifunctional microgel encapsulating outgrowth endothelial cells and growth factors for enhanced neovascularization

Recent cell-based therapy approaches have employed both nanotechnologies and other biomedical technologies to enhance their therapeutic potential. A combined strategy using therapeutic stem/progenitor cells and angiogenic proteins is attractive for the treatment of vascular disease. In this study, we developed an injectable multifunctional micro-sized gel system (microgel), composed of arginine–glycine–aspartic acid (RGD)-conjugated alginate, for the delivery of both cells and growth factors in vivo. The microgels encapsulated with outgrowth endothelial cells (OECs) and growth factors (vascular endothelial growth factor, VEGF, and hepatocyte growth factor, HGF) were formed via electrospraying. Cells encapsulated within the microgel exhibited a time-dependent proliferation with enhanced cell viability, and the size-controlled microgels resulted in sustained release of growth factors for enhanced new vessel formation by tube formation and rat aorta sprouting in vitro. Increased angiogenesis was also estimated in mice treated with RGD-microgel containing OECs and growth factors. Furthermore, injection of the multifunctional microgel into a hindlimb ischemia model improved blood flow perfusion and increased the capillary density by histological analysis. Compared with hydrogel system, injectable microgel system was shown to be superior with no toxicity. Overall, our injectable multifunctional microgel system can be attributed to deliver potential therapeutic agents/cells for the treatment of vascular diseases.