The Synergistic Effect of Electrical Stimulation and Dermal Fibroblast Cells-Laden 3D Conductive Hydrogel for Full-Thickness Wound Healing.

Clinically, most patients with poor wound healing suffer from generalized skin damage, usually accompanied by other complications, so developing therapeutic strategies for difficult wound healing has remained extremely challenging until now. Current studies have indicated that electrical stimulation (ES) to cutaneous lesions enhances skin regeneration by activating intracellular signaling cascades and secreting skin regeneration-related cytokine. In this study, we designed different concentrations of graphene in gelatin-methacrylate (GelMa) to form the conductive composite commonly used in wound healing because of its efficiency compared to other conductive thermo-elastic materials. The results demonstrated the successful addition of graphene to GelMa while retaining the original physicochemical properties of the GelMa bioink. In addition, the incorporation of graphene increased the interactions between these two biomaterials, leading to an increase in mechanical properties, improvement in the swelling ratio, and the regulation of degradation characteristics of the biocomposite scaffolds. Moreover, the scaffolds exhibited excellent electrical conductivity, increasing proliferation and wound healing-related growth factor secretion from human dermal fibroblasts. Overall, the HDF-laden 3D electroconductive GelMa/graphene-based hydrogels developed in this study are ideal biomaterials for skin regeneration applications in the future.

Stiffness of photocrosslinkable gelatin hydrogel influences nucleus pulposus cell propertiesin vitro.

A key early sign of degenerative disc disease (DDD) is the loss of nucleus pulposus (NP) cells (NPCs). Accordingly, NPC transplantation is a treatment strategy for intervertebral disc (IVD) degeneration. However, in advanced DDD, due to structural damage of the IVD and scaffold mechanical properties, the transplanted cells are less viable and secrete less extracellular matrix, and thus, are unable to efficiently promote NP regeneration. In this study, we evaluated the encapsulation of NPCs in a photosensitive hydrogel made of collagen hydrolysate gelatin and methacrylate (GelMA) to improve NP regeneration. By adjusting the concentration of GelMA, we prepared hydrogels with different mechanical properties. After examining the mechanical properties, cell compatibility and tissue engineering indices of the GelMA-based hydrogels, we determined the optimal hydrogel concentration of the NPC-encapsulating GelMA hydrogel for NP regeneration as 5%. NPCs effectively combined with GelMA and proliferated. As the concentration of the GelMA hydrogel increased, the survival, proliferation and matrix deposition of the encapsulated NPCs gradually decreased, which is the opposite of NPCs grown on the surface of the hydrogel. The controllability of the GelMA hydrogels suggests that these NPC-encapsulating hydrogels are promising candidates to aid in NP tissue engineering and repairing endogenous NPCs.

Comparison of the Translational Potential of Human Mesenchymal Progenitor Cells from Different Bone Entities for Autologous 3D Bioprinted Bone Grafts.

Reconstruction of segmental bone defects by autologous bone grafting is still the standard of care but presents challenges including anatomical availability and potential donor site morbidity. The process of 3D bioprinting, the application of 3D printing for direct fabrication of living tissue, opens new possibilities for highly personalized tissue implants, making it an appealing alternative to autologous bone grafts. One of the most crucial hurdles for the clinical application of 3D bioprinting is the choice of a suitable cell source, which should be minimally invasive, with high osteogenic potential, with fast, easy expansion. In this study, mesenchymal progenitor cells were isolated from clinically relevant human bone biopsy sites (explant cultures from alveolar bone, iliac crest and fibula; bone marrow aspirates; and periosteal bone shaving from the mastoid) and 3D bioprinted using projection-based stereolithography. Printed constructs were cultivated for 28 days and analyzed regarding their osteogenic potential by assessing viability, mineralization, and gene expression. While viability levels of all cell sources were comparable over the course of the cultivation, cells obtained by periosteal bone shaving showed higher mineralization of the print matrix, with gene expression data suggesting advanced osteogenic differentiation. These results indicate that periosteum-derived cells represent a highly promising cell source for translational bioprinting of bone tissue given their superior osteogenic potential as well as their minimally invasive obtainability.

Investigating PEGDA and GelMA Microgel Models for Sustained 3D Heterotypic Dermal Papilla and Keratinocyte Co-Cultures.

Hair follicle morphogenesis is heavily dependent on reciprocal, sequential, and epithelial-mesenchymal interaction (EMI) between epidermal stem cells and the specialized cells of the underlying mesenchyme, which aggregate to form the dermal condensate (DC) and will later become the dermal papilla (DP). Similar models were developed with a co-culture of keratinocytes and DP cells. Previous studies have demonstrated that co-culture with keratinocytes maintains the in vivo characteristics of the DP. However, it is often challenging to develop three-dimensional (3D) DP and keratinocyte co-culture models for long term in vitro studies, due to the poor intercellular adherence between keratinocytes. Keratinocytes exhibit exfoliative behavior, and the integrity of the DP and keratinocyte co-cultured spheroids cannot be maintained over prolonged culture. Short durations of culture are unable to sufficiently allow the differentiation and re-programming of the keratinocytes into hair follicular fate by the DP. In this study, we explored a microgel array approach fabricated with two different hydrogel systems. Using poly (ethylene glycol) diacrylate (PEGDA) and gelatin methacrylate (GelMA), we compare their effects on maintaining the integrity of the cultures and their expression of important genes responsible for hair follicle morphogenesis, namely Wnt10A, Wnt10B, and Shh, over prolonged duration. We discovered that low attachment surfaces such as PEGDA result in the exfoliation of keratinocytes and were not suitable for long-term culture. GelMA, on the hand, was able to sustain the integrity of co-cultures and showed higher expression of the morphogens overtime.

High Throughput Screening of Cell Mechanical Response Using a Stretchable 3D Cellular Microarray Platform.

Cells in vivo are constantly subjected to multiple microenvironmental mechanical stimuli that regulate cell function. Although 2D cell responses to the mechanical stimulation have been established, these methods lack relevance as physiological cell microenvironments are in 3D. Moreover, the existing platforms developed for studying the cell responses to mechanical cues in 3D either offer low-throughput, involve complex fabrication, or do not allow combinatorial analysis of multiple cues. Considering this, a stretchable high-throughput (HT) 3D cell microarray platform is presented that can apply dynamic mechanical strain to cells encapsulated in arrayed 3D microgels. The platform uses inkjet-bioprinting technique for printing cell-laden gelatin methacrylate (GelMA) microgel array on an elastic composite substrate that is periodically stretched. The developed platform is highly biocompatible and transfers the applied strain from the stretched substrate to the cells. The HT analysis is conducted to analyze cell mechano-responses throughout the printed microgel array. Also, the combinatorial analysis of distinct cell behaviors is conducted for different GelMA microenvironmental stiffnesses in addition to the dynamic stretch. Considering its throughput and flexibility, the developed platform can readily be scaled up to introduce a wide range of microenvironmental cues and to screen the cell responses in a HT way.

Effects of Encapsulated Cells on the Physical-Mechanical Properties and Microstructure of Gelatin Methacrylate Hydrogels.

Gelatin methacrylate (GelMA) has been gaining popularity in recent years as a photo-crosslinkable biomaterial widely used in a variety of bioprinting and tissue engineering applications. Several studies have established the effects of process-based and material-based parameters on the physical-mechanical properties and microstructure of GelMA hydrogels. However, the effect of encapsulated cells on the physical-mechanical properties and microstructure of GelMA hydrogels has not been fully understood. In this study, 3T3 fibroblasts were encapsulated at different cell densities within the GelMA hydrogels and incubated over 96 h. The effects of encapsulated cells were investigated in terms of mechanical properties (tensile modulus and strength), physical properties (swelling and degradation), and microstructure (pore size). Cell viability was also evaluated to confirm that most cells were alive during the incubation. It was found that with an increase in cell density, the mechanical properties decreased, while the degradation and the pore size increased.

Vascularized Bone-Mimetic Hydrogel Constructs by 3D Bioprinting to Promote Osteogenesis and Angiogenesis.

Bone is a highly vascularized tissue with a unique and complex structure. Long bone consists of a peripheral cortical shell containing a network of channels for vascular penetration and an inner highly vascularized bone marrow space. Bioprinting is a powerful tool to enable rapid and precise spatial patterning of cells and biomaterials. Here we developed a two-step digital light processing technique to fabricate a bone-mimetic 3D hydrogel construct based on octacalcium phosphate (OCP), spheroids of human umbilical vein endothelial cells (HUVEC), and gelatin methacrylate (GelMA) hydrogels. The bone-mimetic 3D hydrogel construct was designed to consist of a peripheral OCP-containing GelMA ring to mimic the cortical shell, and a central GelMA ring containing HUVEC spheroids to mimic the bone marrow space. We further demonstrate that OCP, which is evenly embedded in the GelMA, stimulates the osteoblastic differentiation of mesenchymal stem cells. We refined the design of a spheroid culture device to facilitate the rapid formation of a large number of HUVEC spheroids, which were embedded into different concentrations of GelMA hydrogels. It is shown that the concentration of GelMA modulates the extent of formation of the capillary-like structures originating from the HUVEC spheroids. This cell-loaded hydrogel-based bone construct with a biomimetic dual ring structure can be potentially used for bone tissue engineering.

Fiber-Based Mini Tissue with Morphology-Controllable GelMA Microfibers.

The use of microscale fibers could facilitate nutrient diffusion in fiber-based tissue engineering and improve cell survival. However, in order to build a functional mini tissue such as muscle fibers, nerve conduits, and blood vessels, hydrogel microfibers should not only mimic the structural features of native tissues but also offer a cell-favorable environment and sufficient strength for tissue functionalization. Therefore, an important goal is to fabricate morphology-controllable microfibers with appropriate hydrogel materials to mimic the structural and functional complexity of native tissues. Here, gelatin methacrylate (GelMA) is used as the fiber material due to its excellent biological performance, and a novel coaxial bioprinting method is developed to fabricate morphology-controllable GelMA microfibers encapsulated in calcium alginate. By adjusting the flow rates, GelMA microfibers with straight, wavy, and helical morphologies could be obtained. By varying the coaxial nozzle design, more complex GelMA microfibers such as Janus, multilayered, and double helix structures could be fabricated. Using these microfibers, mini tissues containing human umbilical cord vein endothelial cells are built, in which cells gradually migrate and connect to form lumen resembling blood vessels. The merits of cytocompatibility, structural diversity, and mechanical tunability of the versatile microfibers may open more avenues for further biomedical research.

Facile One-step Micropatterning Using Photodegradable Methacrylated Gelatin Hydrogels for Improved Cardiomyocyte Organization and Alignment.

Hydrogels are often employed as temporary platforms for cell proliferation and tissue organization in vitro. Researchers have incorporated photodegradable moieties into synthetic polymeric hydrogels as a means of achieving spatiotemporal control over material properties. In this study protein-based photodegradable hydrogels composed of methacrylated gelatin (GelMA) and a crosslinker containing o-nitrobenzyl ester groups have been developed. The hydrogels are able to degrade rapidly and specifically in response to UV light and can be photopatterned to a variety of shapes and dimensions in a one-step process. Micropatterned photodegradable hydrogels are shown to improve cell distribution, alignment and beating regularity of cultured neonatal rat cardiomyocytes. Overall this work introduces a new class of photodegradable hydrogel based on natural and biofunctional polymers as cell culture substrates for improving cellular organization and function.

Microfabrication of Gelatin Methacrylate/Hydroxyapatite Composites by Utilizing Alternate Soaking Process.

To construct a complex three-dimensional (3D) structure mimicking bone microstructure, hydrogel models of polymerized gelatin methacrylate (pGelMA) were fabricated by using stereolithography and modified with hydroxyapatite (HAp) via an alternate soaking process (ASP) using a solution of calcium and phosphate ions. Fabricated pGelMA line models whose widths were designed as 100, 300, and 600 µm were modified with HAp by ASP by changing the immersion time and number of cycles. After ASP, all of the line models with widths of 100, 300, and 600 µm were successfully modified with HAp, and large amounts of HAp were covered with the fabricated models by increasing both the immersion time and the number of cycles in ASP. HAp was observed near the surface of the line model with a width of 600 µm after ASP at an immersion time of 10 s, while the entire model was modified with HAp using ASPs for longer immersion times. The adhesion and spread of mesenchymal stem cells (MSCs) on the pGelMA-HAp discs depended on the ASP conditions. Moreover, the HAp modification of 3D pyramid models without alteration of the microstructure was also conducted. This two-step fabrication method of first fabricating frameworks of hydrogel models by stereolithography and subsequently modifying the fabricated models with HAp will lead to the development of 3D cell culture systems to support bone grafts or to create biological niches, such as artificial bone marrow.

A photocrosslinkable and hemostatic bilayer wound dressing based on gelatin methacrylate hydrogel and polyvinyl alcohol foam for skin regeneration.

The enormous potential of multifunctional bilayer wound dressings in various medical interventions for wound healing has led to decades of exploration into this field of medicine. However, it is usually difficult to synthesize a single hydrogel with all the required capabilities simultaneously. This paper proposes a bilayer model with an outer layer intended for hydrogel wound treatment. By adding gelatin methacrylate (GelMA) and tannic acid (TA) to the hydrogel composition and using polyvinyl alcohol-carboxymethyl chitosan (PVA-CMCs) foam layer as supports, a photocrosslinkable hydrogel with an optimal formulation was created. The hydrogels were then examined using a range of analytical procedures, including mechanical testing, rheology, chemical characterization, and in vitro and in vivo tests. The resulting bilayer wound dressing has many desirable properties, namely uniform adhesion and quick crosslinking by UV light. When used against Gram-positive and Gram-negative bacterial strains, bilayer wound dressings demonstrated broad antibacterial efficacy. In bilayer wound dressings with GelMA and TA, better wound healing was observed. Those without these elements showed less effectiveness in healing wounds. Additionally, encouraging collagen production and reducing wound infection has a major therapeutic impact on wounds. The results of this study could have a significant impact on the development of better-performing wound dressings.

Neutrophil Granulopoiesis Optimized Through Ex Vivo Expansion of Hematopoietic Progenitors in Engineered 3D Gelatin Methacrylate Hydrogels.

Neutrophils are the first line of defense of the innate immune system. In response to methicillin-resistant Staphylococcus aureus infection in the skin, hematopoietic stem, and progenitor cells (HSPCs) traffic to wounds and undergo extramedullary granulopoiesis, producing neutrophils necessary to resolve the infection. This prompted the engineering of a gelatin methacrylate (GelMA) hydrogel that encapsulates HSPCs within a matrix amenable to subcutaneous delivery. The authors study the influence of hydrogel mechanical properties to produce an artificial niche for granulocyte-monocyte progenitors (GMPs) to efficiently expand into functional neutrophils that can populate infected tissue. Lin-cKIT+ HSPCs, harvested from fluorescent neutrophil reporter mice, are encapsulated in GelMA hydrogels of varying polymer concentration and UV-crosslinked to produce HSPC-laden gels of specific stiffness and mesh sizes. Softer 5% GelMA gels yield the most viable progenitors and effective cell-matrix interactions. Compared to suspension culture, 5% GelMA results in a twofold expansion of mature neutrophils that retain antimicrobial functions including degranulation, phagocytosis, and ROS production. When implanted dermally in C57BL/6J mice, luciferase-expressing neutrophils expanded in GelMA hydrogels are visualized at the site of implantation for over 5 days. They demonstrate the potential of GelMA hydrogels for delivering HSPCs directly to the site of skin infection to promote local granulopoiesis.

Decellularized matrix bioink with gelatin methacrylate for simultaneous improvements in printability and biofunctionality.

Gelatin methacrylate (GelMA) bioink has been widely used in bioprinting because it is a printable and biocompatible biomaterial. However, it is difficult to print GelMA bioink without any temperature control because it has a thermally-sensitive rheological property. Therefore, in this study, we developed a temperature-controlled printing system in real time without affecting the viability of the cells encapsulated in the bioink. In addition, a skin-derived decellularized extracellular matrix (SdECM) was printed with GelMA to better mimic the native tissue environment compared with solely using GelMA bioink with the enhancement of structural stability. The temperature setting accuracy was calculated to be 98.58 ± 1.8 % for the module and 99.48 ± 1.33 % for the plate from 5 °C to 37 °C. The group of the temperature of the module at 10 °C and the plate at 20 °C have 93.84 % cell viability with the printable range in the printability window. In particular, the cell viability and proliferation were increased in the encapsulated fibroblasts in the GelMA/SdECM bioink, relative to the GelMA bioink, with a morphology that significantly spread for seven days. The gene expression and growth factors related to skin tissue regeneration were relatively upregulated with SdECM components. In the bioprinting process, the rheological properties of the GelMA/SdECM bioink were successfully adjusted in real time to increase printability, and the native skin tissue mimicked components providing tissue-specific biofunctions to the encapsulated cells. The developed bioprinting strategies and bioinks could support future studies related to the skin tissue reconstruction, regeneration, and other medical applications using the bioprinting process.

3D-Bioprinted GelMA Scaffold with ASCs and HUVECs for Engineering Vascularized Adipose Tissue.

The purpose of tissue engineering is to reconstruct parts of injured tissues and to resolve the shortage of organ donations. However, the main concern is the limited size of engineered tissue due to insufficient oxygen and nutrition distribution in large three-dimensional (3D) tissue constructs. To provide better support for cells inside the scaffolds, the vascularization of blood vessels within the scaffold could be a solution. This study compared the effects of different culturing systems using human adipose tissue-derived stem/stromal cells (ASCs), human umbilical vein endothelial cells (HUVECs), and coculture of ASCs and HUVECs in 3D-bioprinted gelatin methacrylate (GelMA) hydrogel constructs. The in vitro results showed that the number of live cells was highest in the coculture of ASCs and HUVECs in the GelMA hydrogel after culturing for 21 days. Additionally, the tubular structure was the most abundant in the GelMA hydrogel, containing both ASCs and HUVECs. In the in vivo test, blood vessels were present in both the HUVECs and the coculture of ASCs and HUVECs hydrogels implanted in mice. However, the blood vessel density was the highest in the HUVEC and ASC coculture groups. These findings indicate that the 3D-bioprinted GelMA hydrogel coculture system could be a promising biomaterial for large tissue engineering applications.

Three-dimensional bioprinted GelMA/GO composite hydrogel for stem cell osteogenic differentiation both in vitro and in vivo.

In this study, we evaluated the use of graphene oxide (GO) mixed with methyl methacrylate gelatin (GelMA) for the construction of a microenvironmental implant to repair bone defects in orthopedic surgery. A scaffold containing a GelMA/GO composite with mesenchymal stem cells (MSCs) was constructed using three-dimensional bioprinting. The survival and osteogenic capacity of MSCs in the composite bioink were evaluated using cell viability and proliferation assays, osteogenesis-related gene expression analysis, and implantation under the skin of nude mice. The printing process had little effect on cell viability. We found that GO enhanced cell proliferation but had no significant effect on cell viability. In vitro experiments suggested that GO promoted material-cell interactions and the expression of osteogenesis-related genes. In vivo experiments showed that GO decreased the degradation time of the material and increased calcium nodule deposition. In contrast to pure GelMA, the addition of GO created a suitable microenvironment to promote the differentiation of loaded exogenous MSCs in vitro and in vivo, providing a basis for the repair of bone defects.

Methacrylated gelatin and platelet-rich plasma based hydrogels promote regeneration of critical-sized bone defects.

Physiological repair of large-sized bone defects requires instructive scaffolds with appropriate mechanical properties, biocompatibility, biodegradability, vasculogenic ability and osteo-inductivity. The objective of this study was to fabricate in situ injectable hydrogels using platelet-rich plasma (PRP)-loaded gelatin methacrylate (GM) and employ them for the regeneration of large-sized bone defects. We performed various biological assays as well as assessed the mechanical properties of GM@PRP hydrogels alongside evaluating the release kinetics of growth factors (GFs) from hydrogels. The GM@PRP hydrogels manifested sufficient mechanical properties to support the filling of the tissue defects. For biofunction assay, the GM@PRP hydrogels significantly improved cell migration and angiogenesis. Especially, transcriptome RNA sequencing of human umbilical vein endothelial cells and bone marrow-derived stem cells were performed to delineate vascularization and biomineralization abilities of GM@PRP hydrogels. The GM@PRP hydrogels were subcutaneously implanted in rats for up to 4 weeks for preliminary biocompatibility followed by their transplantation into a tibial defect model for up to 8 weeks in rats. Tibial defects treated with GM@PRP hydrogels manifested significant bone regeneration as well as angiogenesis, biomineralization, and collagen deposition. Based on the biocompatibility and biological function of GM@PRP hydrogels, a new strategy is provided for the regenerative repair of large-size bone defects.

MSCs-laden silk Fibroin/GelMA hydrogels with incorporation of platelet-rich plasma for chondrogenic construct.

Repair of osteochondral defects and regeneration of cartilage is a major challenge. In this work, the mesenchymal stem cells (MSCs)-laden hydrogel was designed using silk fibroin (SF) and gelatin methacrylate (GelMA), to encapsulate platelet-rich plasma (PRP). Initially, GelMA was synthesized, and SF was prepared using silkworm cocoon, then MSCs-laden SF/GelMA (SG) hydrogel was fabricated. The physicochemical properties of the hydrogels were evaluated using Fourier-transform infrared spectroscopy, scanning electron microscope, and rheometry. After hydrogel preparation, the viability of MSCs in the hydrogels was investigated via CCK-8 analysis and fluorescent images. The MSCs-laden SG hydrogel containing PRP was subsequently injected into the cartilage defect area in Sprague Dawley rats. Hematoxylin and eosin (H&E), Masson staining, and Mankin scores evaluation confirmed the new cartilage formation in 8 weeks. The results presented in the study, therefore, showed that the prepared MSCs-laden SG hydrogel loaded with PRP has the potential for cartilage reconstruction, which is crucial to the treatment of knee osteoarthritis.

Multifunctional and biodegradable methacrylated gelatin/Aloe vera nanofibers for endodontic disinfection and immunomodulation.

Currently employed approaches and materials used for vital pulp therapies (VPTs) and regenerative endodontic procedures (REPs) lack the efficacy to predictably achieve successful outcomes due to their inability to achieve adequate disinfection and/or lack of desired immune modulatory effects. Natural polymers and medicinal herbs are biocompatible, biodegradable, and present several therapeutic benefits and immune-modulatory properties; thus, standing out as a clinically viable approach capable of establishing a conducive environment devoid of bacteria and inflammation to support continued root development, dentinal bridge formation, and dental pulp tissue regeneration. However, the low stability and poor mechanical properties of the natural compounds have limited their application as potential biomaterials for endodontic procedures. In this study, Aloe vera (AV), as a natural antimicrobial and anti-inflammatory agent, was incorporated into photocrosslinkable Gelatin methacrylate (GelMA) nanofibers with the purpose of developing a highly biocompatible biomaterial capable of eradicating endodontic infection and modulating inflammation. Stable GelMA/AV nanofibers with optimal properties were obtained at the ratio of (70:30) by electrospinning. In addition to the pronounced antibacterial effect against Enterococcus faecalis, the GelMA/AV (70:30) nanofibers also exhibited a sustained antibacterial activity over 14 days and significant biofilm reduction with minimal cytotoxicity, as well as anti-inflammatory properties and immunomodulatory effects favoring healing. Our results indicate that the novel GelMA/AV (70:30) nanofibers hold great potential as a biomaterial strategy for endodontic infection eradication and enhanced healing.

Chitin nanocrystal-assisted 3D bioprinting of gelatin methacrylate scaffolds.

In recent years, there has been an increasing focus on the application of hydrogels in tissue engineering. The integration of 3D bioprinting technology has expanded the potential applications of hydrogels. However, few commercially available hydrogels used for 3D biological printing exhibit both excellent biocompatibility and mechanical properties. Gelatin methacrylate (GelMA) has good biocompatibility and is widely used in 3D bioprinting. However, its low mechanical properties limit its use as a standalone bioink for 3D bioprinting. In this work, we designed a biomaterial ink composed of GelMA and chitin nanocrystal (ChiNC). We explored fundamental printing properties of composite bioinks, including rheological properties, porosity, equilibrium swelling rate, mechanical properties, biocompatibility, effects on the secretion of angiogenic factors and fidelity of 3D bioprinting. The results showed that adding 1% (w/v) ChiNC to 10% (w/v) GelMA improved the mechanical properties and printability of the GelMA hydrogels, promoted cell adhesion, proliferation and vascularization and enabled the printing of complex 3D scaffolds. This strategy of incorporating ChiNC to enhance the performance of GelMA biomaterials could potentially be applied to other biomaterials, thereby expanding the range of materials available for use. Furthermore, in combination with 3D bioprinting technology, this approach could be leveraged to bioprint scaffolds with complex structures, further broadening the potential applications in tissue engineering.

Biodegradable double-network GelMA-ACNM hydrogel microneedles for transdermal drug delivery.

As a minimally invasive drug delivery platform, microneedles (MNs) overcome many drawbacks of the conventional transdermal drug delivery systems, therefore are favorable in biomedical applications. Microneedles with a combined burst and sustained release profile and maintained therapeutic molecular bioactivity could further broaden its applications as therapeutics. Here, we developed a double-network microneedles (DN MNs) based on gelatin methacrylate and acellular neural matrix (GelMA-ACNM). ACNM could function as an early drug release matrix, whereas the addition of GelMA facilitates sustained drug release. In particular, the double-network microneedles comprising GelMA-ACNM hydrogel has distinctive biological features in maintaining drug activity to meet the needs of application in treating different diseases. In this study, we prepared the double-network microneedles and evaluated its morphology, mechanical properties, drug release properties and biocompatibility, which shows great potential for delivery of therapeutic molecules that needs different release profiles in transdermal treatment.