Antibiofilm activity of polyethylene glycol-quercetin nanoparticles-loaded gelatin-N,O-carboxymethyl chitosan composite nanogels against Staphylococcus epidermidis.

BACKGROUND: Biofilms, such as those from Staphylococcus epidermidis, are generally insensitive to traditional antimicrobial agents, making it difficult to inhibit their formation. Although quercetin has excellent antibiofilm effects, its clinical applications are limited by the lack of sustained and targeted release at the site of S. epidermidis infection. OBJECTIVES: Polyethylene glycol-quercetin nanoparticles (PQ-NPs)-loaded gelatin-N,O-carboxymethyl chitosan (N,O-CMCS) composite nanogels were prepared and assessed for the on-demand release potential for reducing S. epidermidis biofilm formation. METHODS: The formation mechanism, physicochemical characterization, and antibiofilm activity of PQ-nanogels against S. epidermidis were studied. RESULTS: Physicochemical characterization confirmed that PQ-nanogels had been prepared by the electrostatic interactions between gelatin and N,O-CMCS with sodium tripolyphosphate. The PQ-nanogels exhibited obvious pH and gelatinase-responsive to achieve on-demand release in the micro-environment (pH 5.5 and gelatinase) of S. epidermidis. In addition, PQ-nanogels had excellent antibiofilm activity, and the potential antibiofilm mechanism may enhance its antibiofilm activity by reducing its relative biofilm formation, surface hydrophobicity, exopolysaccharides production, and eDNA production. CONCLUSIONS: This study will guide the development of the dual responsiveness (pH and gelatinase) of nanogels to achieve on-demand release for reducing S. epidermidis biofilm formation.

Creation of Porous, Perfusable Microtubular Networks for Improved Cell Viability in Volumetric Hydrogels.

The creation of large, volumetric tissue-engineered constructs has long been hindered due to the lack of effective vascularization strategies. Recently, 3D printing has emerged as a viable approach to creating vascular structures; however, its application is limited. Here, we present a simple and controllable technique to produce porous, free-standing, perfusable tubular networks from sacrificial templates of polyelectrolyte complex and coatings of salt-containing citrate-based elastomer poly(1,8-octanediol-co-citrate) (POC). As demonstrated, fully perfusable and interconnected POC tubular networks with channel diameters ranging from 100 to 400 µm were created. Incorporating NaCl particulates into the POC coating enabled the formation of micropores (∼19 µm in diameter) in the tubular wall upon particulate leaching to increase the cross-wall fluid transport. Casting and cross-linking gelatin methacrylate (GelMA) suspended with human osteoblasts over the free-standing porous POC tubular networks led to the fabrication of 3D cell-encapsulated constructs. Compared to the constructs without POC tubular networks, those with either solid or porous wall tubular networks exhibited a significant increase in cell viability and proliferation along with healthy cell morphology, particularly those with porous networks. Taken together, the sacrificial template-assisted approach is effective to fabricate tubular networks with controllable channel diameter and patency, which can be easily incorporated into cell-encapsulated hydrogels or used as tissue-engineering scaffolds to improve cell viability.

ICA/SDF-1α/PBMSCs loaded onto alginate and gelatin cross-linked scaffolds promote damaged cartilage repair.

A three-dimensional alginate-coated scaffold (GAIS) was constructed in the present study to showcase the multidifferentiation potential of peripheral blood mesenchymal stem cells (PBMSCs) and to investigate the role and mechanism by which Icariin (ICA)/stromal cell-derived factor (SDF-1α)/PBMSCs promote damaged articular repair. In addition, the ability of ICA, in combination with SDF-1α, to promote the migration and proliferation of stem cells was validated through the utilization of CCK-8 and migration experiments. The combination of ICA and SDF-1α inhibited the differentiation of PBMSCs into cartilage, as demonstrated by in vivo experiments and histological staining. Both PCR and western blot experiments showed that GAIS could upregulate the expression of particular genes in chondrocytes. In comparison to scaffolds devoid of alginate (G0), PBMSCs seeded into GAIS scaffolds exhibited a greater rate of proliferation, and the conditioned medium derived from scaffolds containing SDF-1α enhanced the capacity for cell migration. Moreover, after a 12-week treatment period, GAIS, when successfully transplanted into osteochondral defects of mice, was found to promote cartilage regeneration and repair. The findings, therefore, demonstrate that GAIS enhanced the in vitro capabilities of PBMSCs, including proliferation, migration, homing and chondrogenic differentiation. In addition, ICA and SDF-1α effectively collaborated to support cartilage formation in vivo. Thus, the ICA/SDF-1α/PBMSC-loaded biodegradable alginate-gelatin scaffolds showcase considerable potential for use in cartilage repair.

Injectable Nanoengineered Adhesive Hydrogel for Treating Enterocutaneous Fistulas.

Enterocutaneous fistula (ECF) is a severe medical condition where an abnormal connection forms between the gastrointestinal tract and skin. ECFs are, in most cases, a result of surgical complications such as missed enterotomies or anastomotic leaks. The constant leakage of enteric and fecal contents from the fistula site leads to skin breakdown and increases the risk of infection. Despite advances in surgical techniques and postoperative management, ECF accounts for significant mortality rates, estimated between 15-20%, and causes debilitating morbidity. Therefore, there is a critical need for a simple and effective method to seal and heal ECF. Injectable hydrogels with combined properties of robust mechanical properties and cell infiltration/proliferation have the potential to block and heal ECF. Herein, we report the development of an injectable nanoengineered adhesive hydrogel (INAH) composed of a synthetic nanosilicate (Laponite®) and a gelatin-dopamine conjugate for treating ECF. The hydrogel undergoes fast cross-linking using a co-injection method, resulting in a matrix with improved mechanical and adhesive properties. INAH demonstrates appreciable blood clotting abilities and is cytocompatible with fibroblasts. The adhesive properties of the hydrogel are demonstrated in ex vivo adhesion models with skin and arteries, where the volume stability in the hydrated internal environment facilitates maintaining strong adhesion. In vivo assessments reveal that the INAH is biocompatible, supporting cell infiltration and extracellular matrix deposition while not forming fibrotic tissue. These findings suggest that this INAH holds promising translational potential for sealing and healing ECF.

Insulin Conformation Changes in Hybrid Alginate-Gelatin Hydrogel Particles.

There is a strong need to develop an insulin delivery system suitable for oral administration and preserving natural (α-helix) insulin conformation. In this work, we fabricated alginate-gelatin hydrogel beads for insulin encapsulation. Altering matrix composition and crosslinking agents has resulted in various surface morphologies and internal spatial organization. The structures of the insulin-loaded matrices were studied using optical and field emission electronic microscopy. We use FTIR spectroscopy to identify insulin conformation changes as affected by the hydrogel matrices. It was found that blended alginate-gelatin matrices demonstrate better encapsulation efficiency and stronger swelling resistance to a simulated gastric environment than sodium alginate beads crosslinked with the CaCl2. FTIR measurements reveal conformation changes in insulin. It is also confirmed that in the presence of gelatin, the process of insulin fibrinogenesis ceases due to intermolecular interaction with the gelatin. Performed molecular modeling shows that dipole-dipole interactions are the dominating mechanism that determines insulin behavior within the fabricated matrix.

Adipose-derived stem cells loaded photocurable and bioprintable bioinks composed of GelMA, HAMA and PEGDA crosslinker to differentiate into smooth muscle phenotype.

Developing a polymer-based photocrosslinked 3D printable scaffolds comprised of gelatin methacryloyl (G) and hyaluronic acid methacryloyl (H) incorporated with two molecular weights of polyethylene glycol diacrylate (P) of various concentrations that enables rabbit adipose-derived stem cells (rADSCs) to survive, grow, and differentiate into smooth muscle cells (SMCs). Then, the chemical modification and physicochemical properties of the PGH bioinks were evaluated. The cell viability was assessed via MTT, CCK-8 assay and visualized employing Live/Dead assay. In addition, the morphology and nucleus count of differentiated SMCs were investigated by adopting TRAP (tartrate-resistant acid phosphatase) staining, and quantitative RT-PCR analysis was applied to detect gene expression using two different SMC-specific gene markers α-SMA and SM-MHC. The SMC-specific protein markers namely α-SMA and SM-MHC were applied to investigate SMC differentiation ability by implementing Immunocytofluorescence staining (ICC) and western blotting. Moreover, the disk, square, and tubular cellular models of PGH7 (GelMA/HAMA=2/1) + PEGDA-8000 Da, 3% w/v) hybrid bioink were printed using an extrusion bioprinting and cell viability of rADSCs was also analysed within 3D printed square construct practising Live/Dead assay. The results elicited the overall viability of SMCs, conserving its phenotype in biocompatible PGH7 hybrid bioink revealing its great potential to regenerate SMCs associated organs repair.

Fabrication of gelatin-polyester based biocomposite scaffold via one-step functionalization of melt electrowritten polymer blends in aqueous phase.

The rapid manufacturing of biocomposite scaffold made of saturated-Poly(ε-caprolactone) (PCL) and unsaturated Polyester (PE) blends with gelatin and modified gelatin (NCO-Gel) is demonstrated. Polyester blend-based scaffold are fabricated with and without applying potential in the melt electrowriting system. Notably, the applied potential induces phase separation between PCL and PE and drives the formation of PE rich spots at the interface of electrowritten fibers. The objective of the current study is to control the phase separation between saturated and unsaturated polyesters occurring in the melt electro-writing process and utilization of this phenomenon to improve efficiency of biofunctionalization at the interface of scaffold via Aza-Michael addition reaction. Electron-deficient triple bonds of PE spots on the fibers exhibit good potential for the biofunctionalization via the aza-Michael addition reaction. PE spots are found to be pronounced in which blend compositions are PCL-PE as 90:10 and 75:25 %. The biofunctionalization of scaffold is monitored through CN bond formation appeared at 400 eV via X-ray photoelectron spectroscopy (XPS) and XPS chemical mapping. The described biofunctionalization methodology suggest avoiding use of multi-step chemical modification on additive manufacturing products and thereby rapid prototyping of functional polymer blend based scaffolds with enhanced biocompatibility and preserved mechanical properties. Additionally one-step additive manufacturing method eliminates side effects of toxic solvents and long modification steps during scaffold fabrication.

Nitric oxide-releasing thiolated starch nanoparticles embedded in gelatin sponges for wound dressing applications.

This study introduces a novel wound dressing by combining nitric oxide-releasing thiolated starch nanoparticles (NO-TS NPs) with gelatin. First, starch was thiolated (TS), and then its nanoparticles were prepared (TS NPs). Subsequently, NPs were covalently bonded to sodium nitrite to obtain NO-releasing TS NPs (NO-TS-NPs) that were incorporated into gelatin sponges at various concentrations. The resulting spherical TS NPs had a mean size of 85.42 ± 5.23 nm, which rose to 100.73 ± 7.41 nm after bonding with sodium nitrite. FTIR spectroscopy confirmed S-nitrosation on the NO-TS NPs' surface, and morphology analysis showed well-interconnected pores in all sponges. With higher NO-TS NPs content, pore size, porosity, and water uptake increased, while compressive modulus and strength decreased. Composites exhibited antibacterial activity, particularly against E. coli, with enhanced efficacy at higher NPs' concentrations. In vitro release studies demonstrated Fickian diffusion, with faster NO release in sponges containing more NPs. The released NO amounts were non-toxic to fibroblasts, but samples with fewer NO-TS NPs exhibited superior cellular density, cell attachment, and collagen secretion. Considering the results, including favorable mechanical strength, release behavior, antibacterial and cellular properties, gelatin sponges loaded with 2 mg/mL of NO-TS NPs can be suitable for wound dressing applications.

Poly-L-lactic acid/gelatin electrospun membrane-loaded bone marrow-derived mesenchymal stem cells attenuate erectile dysfunction caused by cavernous nerve injury.

Radical prostatectomy (RP) can cause neurogenic erectile dysfunction (ED), which negatively affects the quality of life of patients with prostate cancer. Currently, there is a dearth of effective therapeutic strategies. Although stem cell therapy is promising, direct cell transplantation to injured cavernous nerves is constrained by poor cell colonization. In this study, poly-L-lactic acid (PLLA)/gelatin electrospun membranes (PGEM) were fabricated to load bone marrow-derived mesenchymal stem cells (BM-MSCs) as a patch to be placed on injured nerves to alleviate ED. This study aimed to establish a promising and innovative approach to mitigate neurogenic ED post-RP and lay the foundation for modifying surgical procedures. Electrospinning and molecular biotechnology were performed in vitro and in vivo, respectively. It was observed that PGEM enhanced the performance of BM-MSCs and Schwann cells due to their excellent mechanical properties and biocompatibility. The transplanted PGEM and loaded BM-MSCs synergistically improved bilateral cavernous nerve injury-related ED and the corresponding histopathological changes. Nevertheless, transplantation of BM-MSCs alone has been verified to be ineffective. Overall, PGEM can serve as an ideal carrier to supply a more suitable survival environment for BM-MSCs and Schwann cells, thereby promoting the recovery of injured cavernous nerves and erectile function.

Surface modification of additive manufactured Ti6Al4V scaffolds with gelatin/alginate- IGF-1 carrier: An effective approach for healing bone defects.

The study investigates the potential of porous scaffolds with Gel/Alg-IGF-1 coatings as a viable candidate for orthopaedic implants. The scaffolds are composed of additively manufactured Ti6Al4V lattices, which were treated in an alkali solution to obtain the anatase and rutile phases. The treated surface exhibited hydrophilicity of <11.5°. A biopolymer carrier containing Insulin-like growth factor 1 was coated on the samples using immersion treatment. This study showed that the surface-modified porous Ti6Al4V scaffolds increased cell viability and proliferation, indicating potential for bone regeneration. The results demonstrate that surface modifications can enhance the osteoconduction and osteoinduction of Ti6Al4V implants, leading to improved bone regeneration and faster recovery. The porous Ti6Al4V scaffolds modified with surface coating of Gel/Alg-IGF-1 exhibited a noteworthy increase in cell viability (from 80.7 to 104.1%viability) and proliferation. These results suggest that the surface modified scaffolds have potential for use in treating bone defects.

Skeletal muscle regeneration with 3D bioprinted hyaluronate/gelatin hydrogels incorporating MXene nanoparticles.

There has been significant progress in the field of three-dimensional (3D) bioprinting technology, leading to active research on creating bioinks capable of producing structurally and functionally tissue-mimetic constructs. Ti3C2Tx MXene nanoparticles (NPs), promising two-dimensional nanomaterials, are being investigated for their potential in muscle regeneration due to their unique physicochemical properties. In this study, we integrated MXene NPs into composite hydrogels made of gelatin methacryloyl (GelMA) and hyaluronic acid methacryloyl (HAMA) to develop bioinks (namely, GHM bioink) that promote myogenesis. The prepared GHM bioinks were found to offer excellent printability with structural integrity, cytocompatibility, and microporosity. Additionally, MXene NPs within the 3D bioprinted constructs encouraged the differentiation of C2C12 cells into skeletal muscle cells without additional support of myogenic agents. Genetic analysis indicated that representative myogenic markers both for early and late myogenesis were significantly up-regulated. Moreover, animal studies demonstrated that GHM bioinks contributed to enhanced regeneration of skeletal muscle while reducing immune responses in mice models with volumetric muscle loss (VML). Our results suggest that the GHM hydrogel can be exploited to craft a range of strategies for the development of a novel bioink to facilitate skeletal muscle regeneration because these MXene-incorporated composite materials have the potential to promote myogenesis.

Storage stability of Asian seabass oil-in-water Pickering emulsion packed in pouches made from electrospun and solvent casted bilayer films from poly lactic acid/chitosan-gelatin blend containing epigallocatechin gallate.

Bilayer pouches were fabricated with chitosan (CS)-fish gelatin (FG) mixture containing epigallocatechin gallate (EGCG) deposited over the poly lactic acid (PLA) film through solvent casting and electrospinning techniques. Pickering emulsions (PE) of Asian seabass depot fat oil stabilized by zein colloidal particles were packed in bilayer pouches and stored at 28 ± 2 °C. The PE packed in pouch containing EGCG had higher emulsion and oxidative stability after 30 days of storage as witnessed by the smaller droplet size and lower values of thiobarbituric acid reactive substances, peroxide, conjugated diene and volatile compounds in comparison with control (PE packed in monolayer PLA pouch) (P < 0.05). EGCG incorporated pouch retained more linoleic acid (C18:2 n-6) and linolenic acid (C18:3 n-9) in emulsion than PLA pouch. Therefore, pouch from bilayer PLA/CS-FG films comprising EGCG could serve as active packaging and extended the shelf life of Pickering emulsion.

A dual network cross-linked hydrogel with multifunctional Bletilla striata polysaccharide/gelatin/tea polyphenol for wound healing promotion.

Wound healing is a dynamic and complex biological process, and traditional biological excipients cannot meet the needs of the wound healing process, and there is an urgent need for a biological dressing with multifunctionality and the ability to participate in all stages of wound healing. This study developed tea polyphenol (TP) incorporated multifunctional hydrogel based on oxidized Bletilla striata polysaccharide (OBSP) and adipic acid dihydrazide modified gelatin (Gel-ADH) with antimicrobial, antioxidant hemostatic, and anti-inflammatory properties to promote wound healing. The composite OBSP, Gel-ADH, TP (OBGTP) hydrogels prepared by double crosslinking between OBSP, TP and Gel-ADH via Schiff base bonding and hydrogen bonding had good rheological and swelling properties. The introduction of TP provided the composite hydrogel with excellent antioxidant antibacterial activities against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coil). In the rat liver hemorrhage model and skin injury model, the OBGTP composite hydrogel had significant (p < 0.001) hemostatic ability, and had the ability to accelerate collagen deposition, reduce the expression of inflammatory factors, and promote rapid wound healing. In addition, OBGTP hydrogels had adhesive properties and good biocompatibility. In conclusion, OBGTP multifunctional composite hydrogels have great potential for wound healing applications.

Fabrication of multifunctional ethyl cellulose/gelatin-based composite nanofilm for the pork preservation and freshness monitoring.

In this study, an active and intelligent nanofilm for monitoring and maintaining the freshness of pork was developed using ethyl cellulose/gelatin matrix through electrospinning, with the addition of natural purple sweet potato anthocyanin. The nanofilm exhibited discernible color variations in response to pH changes, and it demonstrated a higher sensitivity towards volatile ammonia compared with casting film. Notably, the experimental findings regarding the wettability and pH response performance indicated that the water contact angle between 70° and 85° was more favorable for the smart response of pH sensitivity. Furthermore, the film exhibited desirable antioxidant activities, water vapor barrier properties and also good antimicrobial activities with the incorporation of ε-polylysine, suggesting the potential as a food packaging film. Furthermore, the application preservation outcomes revealed that the pork packed with the nanofilm can prolong shelf life to 6 days, more importantly, a distinct color change aligned closely with the points indicating the deterioration of the pork was observed, changing from light pink (indicating freshness) to light brown (indicating secondary freshness) and then to brownish green (indicating spoilage). Hence, the application of this multifunctional film in intelligent packaging holds great potential for both real-time indication and efficient preservation of the freshness of animal-derived food items.

Enhanced antibacterial activity of porous chitosan-based hydrogels crosslinked with gelatin and metal ions.

Addressing the increasing drug resistance in pathogenic microbes, a significant threat to public health, calls for the development of innovative antibacterial agents with versatile capabilities. To enhance the antimicrobial activity of non-toxic biomaterials in this regard, this study focuses on novel, cost-effective chitosan (CS)-based hydrogels, crosslinked using gelatin (GEL), formaldehyde, and metallic salts (Ag+, Cu2+, and Zn2+). These hydrogels are formed by mixing CS and GEL with formaldehyde, creating iminium ion crosslinks with metallic salts without hazardous crosslinkers. Characterization techniques like FTIR, XRD, FESEM, EDX, and rheological tests were employed. FTIR analysis showed metal ions binding to amino and hydroxyl groups on CS, enhancing hydrogelation. FESEM revealed that freeze-dried hydrogels possess a crosslinked, porous structure influenced by various metal ions. Antibacterial testing against gram-negative and gram-positive bacteria demonstrated significant bacterial growth inhibition. CS-based hydrogels containing metal ions showed reduced MIC and MBC values against Staphylococcus aureus (0.5, 8, 16 µg/mL) and Escherichia coli (1, 16, 8 µg/mL) for CS-g-GEL-Ag+, CS-g-GEL-Cu2+, and CS-g-GEL-Zn2+. MTT assay results confirmed high biocompatibility (84.27%, 85.24%, 84.96% viability at 10 µg/mL) for CS-based hydrogels towards HFF-1 cells over 48 h. Therefore, due to their non-toxic nature, these CS hydrogels are promising for antibacterial applications.

A tough and bioadhesive injectable hydrogel formed with maleimidyl alginate and pristine gelatin.

Injectable hydrogels have wide applications in clinical practice. However, the development of tough and bioadhesive ones based on biopolymers, along with biofriendly and robust crosslinking strategies, still represents a great challenge. Herein, we report an injectable hydrogel composed of maleimidyl alginate and pristine gelatin, for which the precursor solutions could self-crosslink via mild Michael-type addition without any catalyst or external energy upon mixing. This hydrogel is tough and bioadhesive, which can maintain intactness as well as adherence to the defect of porcine skin under fierce bending and twisting, warm water bath, and boiling water shower. Besides, it is biocompatible, bioactive and biodegradable, which could support the growth and remodeling of cells by affording an extracellular matrix-like environment. As a proof of application, we demonstrate that this hydrogel could significantly accelerate diabetic skin wound healing, thereby holding great potential in healthcare.

Measuring cell proliferation in bioprinting research.

Tissue-like constructs, intended for application in tissue engineering and regenerative medicine, can be produced by three-dimensional (3D) bioprinting of cells in hydrogels. It is essential that the viability and proliferation of the encapsulated cells can be reliably determined. Methods currently used to evaluate cell proliferation, such as quantification of DNA and measurement of metabolic activity, have been developed for application in 2D cultures and might not be suitable for bioinks. In this study, human fibroblasts were either cast or printed in gelatin methacryloyl (GelMA) or sodium alginate hydrogels and cell proliferation was assessed by AlamarBlue, PicoGreen and visual cell counts. Comparison of data extrapolated from standard curves generated from 2D cultures and 3D hydrogels showed potential inaccuracies. Moreover, there were pronounced discrepancies in cell numbers obtained from these assays; the different bioinks strongly influenced the outcomes. Overall, the results indicate that more than one method should be applied for better assessment of cell proliferation in bioinks.

Conductive and Enhanced Mechanical Strength of Mo2Ti2C3 MXene-Based Hydrogel Promotes Neurogenesis and Bone Regeneration in Bone Defect Repair.

Bone defects are common with increasing high-energy fractures, tumor bone invasion, and implantation revision surgery. Bone is an electroactive tissue that has electromechanical interaction with collogen, osteoblasts, and osteoclasts. Hydrogel provides morphological plasticity and extracellular matrix (ECM) 3D structures for cell survival, and is widely used as a bone engineering material. However, the hydrogels have poor mechanical intensity and lack of cell adhesion, slow gelation time, and limited conductivity. MXenes are novel nanomaterials with hydrophilic groups that sense cell electrophysiology and improve hydrogel electric conductivity. Herein, gelatin had multiple active groups (NH2, OH, and COOH) and an accelerated gelation time. Acrylamide has Schiff base bonds to cross-link with gelatin and absorb metal ions. Deacetylated chitosan improved cell adhesion and active groups to connect MXene and acrylamide. We constructed Mo2Ti2C3 MXene hydrogel with improved elastic modulus and viscosity, chemical cross-linking structure, electric conductivity, and good compatibility. Mo2Ti2C3 MXene hydrogel exhibits outstanding osteogenesis in vitro. Mo2Ti2C3 MXene hydrogel promotes osteogenesis via alkaline phosphatase (ALP) and alizarin red S (ARS) staining, improving osteogenic marker genes and protein expressions in vitro. Mo2Ti2C3 MXene hydrogel aids new bone formation in the in vivo calvarial bone defect model via micro-CT and histology. Mo2Ti2C3 MXene hydrogel facilitates neurogenesis factors nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) expression, and aids newly born neuron marker Tuj-1 and sensory neuron marker serotonin (5-HT) and osteogenesis pathway proteins, runt-related transcription factor 2 (Runx2), osteocalcin (OCN), SMAD family member 4 (SMAD4), and bone morphogenetic protein-2 (BMP2) in the bone defect repair process. Mo2Ti2C3 MXene hydrogel promotes osteogenesis and neurogenesis, which extends its biomedical application in bone defect reconstruction.

MicroRNA-200c Release from Gelatin-Coated 3D-Printed PCL Scaffolds Enhances Bone Regeneration.

The fabrication of clinically relevant synthetic bone grafts relies on combining multiple biodegradable biomaterials to create a structure that supports the regeneration of defects while delivering osteogenic biomolecules that enhance regeneration. MicroRNA-200c (miR-200c) functions as a potent osteoinductive biomolecule to enhance osteogenic differentiation and bone formation; however, synthetic tissue-engineered bone grafts that sustain the delivery of miR-200c for bone regeneration have not yet been evaluated. In this study, we created novel, multimaterial, synthetic bone grafts from gelatin-coated 3D-printed polycaprolactone (PCL) scaffolds. We attempted to optimize the release of pDNA encoding miR-200c by varying gelatin types, concentrations, and polymer crosslinking materials to improve its functions for bone regeneration. We revealed that by modulating gelatin type, coating material concentration, and polymer crosslinking, we effectively altered the release rates of pDNA encoding miR-200c, which promoted osteogenic differentiation in vitro and bone regeneration in a critical-sized calvarial bone defect animal model. We also demonstrated that crosslinking the gelatin coatings on the PCL scaffolds with low-concentration glutaraldehyde was biocompatible and increased cell attachment. These results strongly indicate the potential use of gelatin-based systems for pDNA encoding microRNA delivery in gene therapy and further demonstrate the effectiveness of miR-200c for enhancing bone regeneration from synthetic bone grafts.