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In regenerative medicine and tissue engineering, the repair and replacement of damaged or lost bone tissue remains an enduring challenge. Traditional approaches often fail to achieve desired outcomes, but recent advancements in material science have introduced innovative solutions to this issue. Gelatin methacryloyl (GelMA), which is derived from gelatin, has a unique combination of biocompatibility and tunable physical properties for bone tissue engineering applications. In this study, a novel composite was constructed to combine the characteristics of GelMA and bioactive layered double hydroxide (LDH) loaded calcein. The GelMA-LDH-calcein (GLC) scaffold is designed to promote the osteogenic differentiation of mesenchymal stem cells (MSCs) and provide a microenvironment conducive to osteogenic differentiation, ultimately increasing the osteogenic activity to support bone regeneration. Our data demonstrated that GLC promoted the osteogenic differentiation related gene (ALP, Runx2, and OPG) expression in bone marrow-derived MSCs (BMSCs) and angiogenesis of human vascular epithelial cells (HUVECs), making it particularly valuable in the application of bone regeneration in vivo. Through the design, synthesis, and optimization of GLC composites, the optimized scaffolds can provide structure supporting and biological activity for cell proliferation, which promoted bone defect regeneration and could be used for further clinical applications.

Gelatin methacrylate (GelMA) hydrogels have gained significant traction in diverse tissue engineering applications through the utilization of 3D printing technology. As an artificial hydrogel possessing remarkable processability, GelMA has emerged as a pioneering material in the advancement of tissue engineering due to its exceptional biocompatibility and degradability. The integration of 3D printing technology facilitates the precise arrangement of cells and hydrogel materials, thereby enabling the creation of in vitro models that simulate artificial tissues suitable for transplantation. Consequently, the potential applications of GelMA in tissue engineering are further expanded. In tissue engineering applications, the mechanical properties of GelMA are often modified to overcome the hydrogel material's inherent mechanical strength limitations. This review provides a comprehensive overview of recent advancements in enhancing the mechanical properties of GelMA at the monomer, micron, and nano scales. Additionally, the diverse applications of GelMA in soft tissue engineering via 3D printing are emphasized. Furthermore, the potential opportunities and obstacles that GelMA may encounter in the field of tissue engineering are discussed. It is our contention that through ongoing technological progress, GelMA hydrogels with enhanced mechanical strength can be successfully fabricated, leading to the production of superior biological scaffolds with increased efficacy for tissue engineering purposes.

The clinical treatments of bone defects remain a challenge. Hydrogels containing bone marrow mesenchymal stem cells (BMSCs) are extensively used to bone regeneration because of excellent biocompatibility and hydrophilicity. However, the insufficient osteo-induction capacity of the BMSC-loaded hydrogels limits their clinical applications. In this study, bio-active glass (BG) and BMSCs were combined with gelatin methacryloyl (GelMA) to fabricate composite hydrogels via photo-crosslinking, and the regulation of bone regeneration was investigated. In vitro experiments showed that the BG/BMSCs@GelMA hydrogel had excellent cytocompatibility and promoted osteogenic differentiation in BMSCs. Furthermore, the BG/BMSCs@GelMA hydrogel was injected into critical-sized calvarial defects, and the results further confirmed its excellent angiogenetic and bone regeneration capacity. In addition, BG/BMSCs@GelMA promoted the polarization of macrophages towards the M2 phenotype. In summary, this novel composite hydrogel demonstrated remarkable potential for application in bone regeneration due to its immunomodulatory, excellent angiogenetic as well as osteo-induction capacity.

Rationale: Chronic liver fibrosis is a common and irreversible disease in clinical practice, and Pirfenidone (PFD) has been shown to effectively inhibit the progression of liver fibrosis in multiple studies. However, some critically ill patients cannot take oral medications, and high dose and frequency of administration can cause higher physical burden to the critically ill. Therefore, we are committed to finding a material that can provide an efficient mode of drug delivery to address chronic disease. Methods: We have developed a hydrogel microneedle patch (MNP) based on GelMA material to provide sustained release of PFD for reducing dosing frequency, alleviating the physical burden, and enhancing therapeutic efficacy. In vitro experiments were conducted to verify the effectiveness of the microneedle patch, and a chronic liver fibrosis model was established in C57 mouse for further validation. Results: Gel-PFD MNP demonstrated good biocompatibility in vitro and in vivo studies. Cy7 fluorescence showed complete degradation of the MNP after 1 week in vivo. In vitro, Gel-PFD MNP inhibited fibroblast migration and proliferation. In mice treated with Gel-PFD, markers of fibrosis, inflammation and apoptosis in tissue or serum RNA and protein expression were relatively low, and liver enzymes and behavioral levels were also relatively normal. Conclusions: Gel-PFD has good biocompatibility and sustained-release abilities, which can significantly reduce the frequency of administration and has better anti-fibrosis ability than oral PFD. It has extremely positive implications for improving chronic liver injury in patients who are unable to eat or severely ill.

The key to critical bone regeneration in tissue engineering relies on an ideal bio-scaffold coated with a controlled release of growth factors. Gelatin methacrylate (GelMA) and Hyaluronic acid methacrylate (HAMA) have been a novel topic of interest in bone regeneration while introducing appropriate nano-hydroxyapatite (nHAP) to improve its mechanical properties. And the exosomes derived from human urine-derived stem cells (human USCEXOs) have also been reported to promote osteogenesis in tissue engineering. The present study aimed to design a new GelMA-HAMA/nHAP composite hydrogel as a drug delivery system. The USCEXOs were encapsulated and slow-released in the hydrogel for better osteogenesis. The characterization of the GelMA-based hydrogel showed excellent controlled release performance and appropriate mechanical properties. The in vitro studies showed that the USCEXOs/GelMA-HAMA/nHAP composite hydrogel could promote the osteogenesis of bone marrow mesenchymal stem cells (BMSCs) and the angiogenesis of endothelial progenitor cells (EPCs), respectively. Meanwhile, the in vivo results confirmed that this composite hydrogel could significantly promote the defect repair of cranial bone in the rat model. In addition, we also found that USCEXOs/GelMA-HAMA/nHAP composite hydrogel can promote the formation of H-type vessels in the bone regeneration area, enhancing the therapeutic effect. In conclusion, our findings suggested that this controllable and biocompatible USCEXOs/GelMA-HAMA/nHAP composite hydrogel may effectively promote bone regeneration by coupling osteogenesis and angiogenesis.

The presence of periosteum can greatly affect the repair of a bone fracture. An artificial periosteum imitates the biological function of natural periosteum, which is capable of protecting and maintaining bone tissue structure and promoting bone repair. In our artificial periosteum, biocompatible methacrylate gelatin was used to provide the mechanical support of the membrane, E7 peptide added bioactivity, and Wharton's jelly provided the biological activity support of the membrane, resulting in a hydrogel membrane (G-E-W) for the chemotactic recruitment of bone marrow mesenchymal stem cells (BMSCs) and promoting cell proliferation and osteogenic differentiation. In an in vitro experiment, the G-E-W membrane recruited BMSCs and promoted cell proliferation and osteogenic differentiation. After 4 weeks and 8 weeks of implantation in a rat skull defect, the group implanted with a G-E-W membrane was superior to the blank control group and single-component membrane group in both quantity and quality of new bone. The artificial G-E-W membrane recruits BMSC chemotaxis and promotes cell proliferation and osteogenic differentiation, thereby effectively improving the repair efficiency of fractures and bone defects.

Cryopreserved testicular tissue offers a promising method to restore fertility in male infertility patients. Current protocols rely on high concentrations of penetrating cryoprotectants (pCPAs), such as dimethyl sulfoxide (DMSO), which necessitating complex washing procedures and posing risks of toxicity. Hydrogel encapsulation presents a non-toxic alternative for cellular cryopreservation. This study investigates the effects of various types, concentrations, and thicknesses of hydrogel encapsulation on the cryopreservation of mouse testicular tissue. Testicular tissues loaded with varying concentrations of DMSO were encapsulated in alginate or gelatin-methacryloyl (GelMA) hydrogels. We evaluated hydrogels as potential CPAs to reduce pCPA concentrations and determine optimal combinations for cryopreservation. Post-cryopreservation, tissues were cultured using organ culture methods to assess spermatogenesis progression. Cryomicroscopy and differential scanning calorimetry (DSC) were used to examine ice crystal formation, melting enthalpy, and non-freezing water content in different hydrogels during cooling. Results indicate that 3 % alginate or 5 % GelMA hydrogel with thin encapsulation optimally preserves mouse testicular tissue. Using 20 % DMSO in 5 % GelMA thin encapsulation showed comparable apoptosis rates, improved morphology, higher mitochondrial activity, and enhanced antioxidant capacity compared to conventional 30 % DMSO without encapsulation. This suggests that hydrogel encapsulation reduces pCPA concentration by 10 %, thereby mitigating toxic damage. Hydrogel encapsulation can reduce basement membrane shrinkage of testicular tissue during cryopreservation. Moreover, frozen tissues remained viable with preserved germ cells after being cultured for one week on alginate methacryloyl (AlgMA) hydrogel using the gas-liquid interphase method. Cryomicroscopy and DSC studies confirmed the hydrogel's ability to inhibit ice crystal growth. In conclusion, this study introduces novel strategies for male fertility preservation and advances cryopreservation technology for clinical applications in assisted reproduction., Competing Interests: Declaration of Competing Interest The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Skin wounds, prevalent and fraught with complications, significantly impact individuals and society. Wound healing encounters numerous obstacles, such as excessive reactive oxygen species (ROS) production and impaired angiogenesis, thus promoting the development of chronic wound. Traditional clinical interventions like hemostasis, debridement, and surgery face considerable challenges, including the risk of secondary infections. While therapies designed to scavenge excess ROS and enhance proangiogenic properties have shown effectiveness in wound healing, their clinical adoption is hindered by high costs, complex manufacturing processes, and the potential for allergic reactions. Lotus root, distinguished by its natural micro and macro porous architecture, exhibits significant promise as a tissue engineering scaffold. This study introduced a novel scaffold based on hybridization of lotus root-inspired and Gelatin Methacryloyl (GelMA), verified with satisfactory physicochemical properties, biocompatibility, antioxidative capabilities and proangiogenic abilities. In vivo tests employing a full-thickness wound model revealed that these scaffolds notably enhanced micro vessel formation and collagen remodeling within the wound bed, thus accelerating the healing process. Given the straightforward accessibility of lotus roots and the cost-effective production of the scaffolds, the novel scaffolds with ROS scavenging, pro-angiogenesis and re-epithelialization abilities are anticipated to have clinical applicability for various chronic wounds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Gelatin methacryloyl (GelMA) holds significant potential in tissue engineering; however, its clinical applications are often constrained by its lack of functional groups. To overcome this limitation, recombinant proteins with multiple biofunctional domains present a promising strategy for GelMA functionalization, enhancing its biological properties. In this study, we developed a rationally designed recombinant collagen-like protein (RC) engineered with multiple biofunctional domains, which demonstrated the ability to upregulate collagen 1α (COL-1α) expression in NIH-3 T3 cells. By utilizing EDC/NHS chemistry, the purified RC was conjugated to GelMA, resulting in GelMA-RC hydrogels that significantly improved cell viability and migration compared to unmodified GelMA. Subsequent in vivo studies showed that RC-modified GelMA exhibited superior wound healing efficacy, largely attributed to enhanced expression of cytokeratin-14 (CK-14) and COL-1α. These findings underscore the potential of RC-functionalized GelMA in promoting diabetic wound repair and suggest broader applicability for functionalizing other biomaterials., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Xinyu Li reports financial support was provided by Inner Mongolia University. Xinyu Li reports a relationship with Inner Mongolia University that includes: employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cryopreservation of biological samples is an important technology for expanding their applications in the biomedical field. However, the quality and functionality of samples after rewarming are limited by the toxicity of commonly used cryoprotectant agents (CPAs). Here, we developed a novel preservation system by combining the natural amino acid l-proline (L-Pro) with gelatin methacryloyl (GelMA) hydrogels. Compared with dimethyl sulfoxide (DMSO), L-Pro and GelMA demonstrated excellent biocompatibility when co-culturing with cells. Cryopreservation procedures were optimized using 3T3 as model cells. The results showed that rapid cooling was the most suitable cooling procedure for L-Pro and GelMA among the three cooling procedures. Co-culturing with cells for 3 h before cryopreservation, 6 % L-Pro +7 % GelMA had the highest survival rate, reaching up to 80 %. Differential Scanning Calorimetry (DSC) analysis showed that 6 % L-Pro + 7 % GelMA lowered the freezing point of the solution to -4.2 °C and increased the unfrozen water content to 20 %. To the best of our knowledge, this is the first report of cell cryopreservation using a combination of L-Pro and GelMA hydrogels, which provides a new strategy for improving cell cryopreservation., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 Society for Cryobiology. Published by Elsevier Inc. All rights reserved.)

Aim: Three-dimensional (3D) cell culture systems perform better in resembling tissue or organism structures compared with traditional 2D models. Organs-on-chips (OoCs) are becoming more efficient 3D models. This study aimed to create a novel simplified dentin-on-a-chip using microfluidic chip technology and tissue engineering for screening dental materials., Methodology: A microfluidic device with three channels was designed for creating 3D dental tissue constructs using stem cells from the apical papilla (SCAP) and gelatin methacrylate (GelMA). The study investigated the effect of varying cell densities and GelMA concentrations on the layer features formed within the microfluidic chip. Cell viability and distribution were evaluated through live/dead staining and nuclei/F-actin staining. The osteo/odontogenic potential was assessed through ALP staining and Alizarin red staining. The impact of GelMA concentrations (5 %, 10 %) on the osteo/odontogenic differentiation trajectory of SCAP was also studied., Results: The 3D tissue constructs maintained high viability and favorable spreading within the microfluidic chip for 3-7 days. A cell seeding density of 2 × 10 4 cells/μL was found to be the most optimal choice, ensuring favorable cell proliferation and even distribution. GelMA concentrations of 5 % and 10 % proved to be most effective for promoting cell growth and uniform distribution. Within the 5 % GelMA group, SCAP demonstrated higher osteo/odontogenic differentiation than that in the 10 % GelMA group., Conclusion: In 3D culture, GelMA concentration was found to regulate the osteo/odontogenic differentiation of SCAP. The study recommends a seeding density of 2 × 10 4 cells/μL of SCAP within 5 % GelMA for constructing simplified dentin-on-a-chip., Clinical Significance: This study built up the 3D culture protocol, and induced odontogenic differentiation of SCAP, thus forming the simplified dentin-on-a-chip and paving the way to be used as a well-defined biological model for regenerative endodontics. It may serve as a potential testing platform for cell differentiation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Hydrogels incorporating natural biopolymer and adhesive substances have extensively been used to develop bioactive drugs and to design cells encapsulating sturdy structure for biomedical applications. However, the conjugation of the adhesive in most hydrogels is insufficient to maintain long-lasting biocompatibility inadequate to accelerate internal organ tissue repair in the essential native cellular microenvironment. The current work elaborates the synthesis of charged choline-catechol ionic liquid (BIL) adhesive and a hydrogel with an electronegative atom rich polyphenol (PU)-laden gelatinmethacryloyl (GelMA) to improve the structural bioactivities for in vivo tracheal repair by inducing swift crosslinking along with durable mechanical and tissue adhesive properties. It was observed that bioactive BIL and PU exhibited potent antioxidant (IC 50 % of 7.91 μg/mL and 24.55 μg/mL) and antibacterial activity against E. coli, P. aeruginosa and S. aureus. The novel integration of photocurable GelMA-BIL-PU revealed outstanding mechanical strength, biodegradability and sustained drug release. The in vitro study showed exceptional cell migration and proliferation in HBECs, while in vivo investigation of the GelMA-BIL-PU hydrogel on a rat's tracheal model revealed remarkable tracheal reconstruction, concurrently reducing tissue inflammation. Furthermore, the optimized GelMA-BIL-PU injectable adhesive bioink blend demonstrated superior MSCs migration and proliferation, which could be a strong candidate for developing stem cell-rich biomaterials to address multiple organ defects., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Peripheral nerve injuries (PNIs) can cause neuropathies and significantly affect the patient's quality of life. Autograft transplantation is the gold standard for conventional treatment; however, its application is limited by nerve unavailability, size mismatch, and local tissue adhesion. Tissue engineering, such as nerve guidance conduits, is an alternative and promising strategy to guide nerve regeneration for peripheral nerve repair; however, only a few conduits could reach the high repair efficiency of autografts. The healing process of PNI is frequently accompanied by not only axonal and myelination regeneration but also angiogenesis, which initializes nerve regeneration through vascular endothelial growth factor A (VEGF-A). In this study, a composite nerve conduit with a poly (lactic-co-glycolic acid) (PLGA) hollow tube as the outer layer and gelatin methacryloyl (GelMA) encapsulated with VEGF-A transfected Schwann cells (SCs) as the inner layer was established to evaluate its promising ability for peripheral nerve repair. A rat model of peripheral nerve defect was used to examine the efficiency of PLGA/GelMA-SC (VA) conduits, whereas autograft, PLGA, PLGA/GelMA, and PLGA/GelMA-SC (NC) were used as controls. VEGF-A-transfected SCs can provide a stable source for VEGF-A secretion. Furthermore, encapsulation in GelMA cannot only promote proliferation and tube formation of human umbilical vein endothelial cells but also enhance dorsal root ganglia and neuronal cell extension. Previous animal studies have demonstrated that the regenerative effects of PLGA/GelMA-SC (VA) nerve conduit were similar to those of autografts and much better than those of other conduits. These findings indicate that combination of VEGF-A-overexpressing SCs and PLGA/GelMA conduit-guided peripheral nerve repair provides a promising method that enhances angiogenesis and regeneration during nerve repair. STATEMENT OF SIGNIFICANCE: Nerve guidance conduits shows promise for peripheral nerve repair, while achieving the repair efficiency of autografts remains a challenge. In this study, a composite nerve conduit with a PLGA hollow tube as the outer layer and gelatin methacryloyl (GelMA) encapsulated with vascular endothelial growth factor A (VEGF-A)-transfected Schwann cells (SCs) as the inner layer was established to evaluate its potential ability for peripheral nerve repair. This approach preserves growth factor bioactivity and enhances material properties. GelMA insertion promotes Schwann cell proliferation and morphology extension. Moreover, transfected SCs serve as a stable VEGF-A source and fostering angiogenesis. This study offers a method preserving growth factor efficacy and safeguarding SCs, providing a comprehensive solution for enhanced angiogenesis and nerve regeneration., Competing Interests: Declaration of competing interest The author(s) declared no potential conflicts of interest with respect to the research, author - ship, and/or publication of this article., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Management of non-healing and slow to heal diabetic wounds is a major concern in healthcare across the world. Numerous techniques have been investigated to solve the issue of delayed wound healing, though, mostly unable to promote complete healing of diabetic wounds due to the lack of proper cell proliferation, poor cell-cell communication, and higher chances of wound infections. These challenges can be minimized by using hydrogel based wound healing patches loaded with bioactive agents. Gelatin methacrylate (GelMA) has been proven to be a highly cell friendly, cell adhesive, and inexpensive biopolymer for various tissue engineering and wound healing applications. In this study, S-Nitroso-N-acetylpenicillamine (SNAP), a nitric oxide (NO) donor, was incorporated in a highly porous GelMA hydrogel patch to improve cell proliferation, facilitate rapid cell migration, and enhance diabetic wound healing. We adopted a visible light crosslinking method to fabricate this highly porous biodegradable but relatively stable patch. Developed patches were characterized for morphology, NO release, cell proliferation and migration, and diabetic wound healing in a rat model. The obtained results indicate that SNAP loaded visible light crosslinked GelMA hydrogel patches can be highly effective in promoting diabetic wound healing.

Bone repair strategies, based on endogenous stem cell recruitment, can effectively avoid immune rejection and the low utilization of exogenous stem cells. Endogenous stem cells can be recruited to the implantation site by loading chemokines onto bone tissue-engineered scaffolds. However, challenges such as unstable chemokine activity and easy inactivation after implantation remain significant. In the present study, composite fiber scaffolds ((IL8@LIP)-GelMA) consisting of Interleukin 8 (IL8) -loaded liposomes and GelMA were constructed by electrospinning and photocrosslinking, and its ability to recruit bone marrow-derived mesenchymal stem cells (BMSCs) and immunomodulatory effect was investigated. Compared to GelMA loaded directly with IL8, scaffolds of (IL8@LIP)-GelMA demonstrated superior protection of IL8 activity, ensuring a slow and continuous release. Both in vivo and in vitro experiments demonstrated that the (IL8@LIP)-GelMA scaffolds effectively recruited BMSCs to the desired sites. Additionally, the (IL8@LIP)-GelMA scaffolds exhibited the capacity to recruit more macrophages to the implantation site. Importantly, they promoted the polarization of macrophages toward the M2 anti-inflammatory phenotype, facilitating the transition from the inflammatory stage to the tissue repair stage. Therefore, (IL8@LIP)-GelMA scaffolds show great potential for cell-free tissue engineering applications and provide insights into the loading mode of growth factors in scaffolds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Corneal transplantation serves as the standard clinical therapy for serious corneal disorders. However, rejection of grafts, significant expenditures, and most crucially, the global donor shortage, may affect the outcome. Recently, 3D bioprinting using biodegradable polymeric materials has become a suitable method for creating tissue replicas with identical architecture. One such most renowned material is GelMA, for its scaffold's three-dimensional structure, biocompatibility, robust mechanics, and favourable optical transmittance. However, GelMA's inadequate viscosity to print at body temperature with better form integrity remains an obstacle. The aim of this work is to create 3D printed GelMA/MC hydrogels for corneal stroma tissue engineering using MC's printability at room temperature and GelMA's irreversible photo cross-linking with UV irradiation. The print speed and pressure conditions for 3D GelMA/MC hydrogels were tuned. Thermal, morphological and physicochemical characteristics were studied for two distinct concentrations of GelMA/MC hydrogels. The hydrogels achieved a transparency of ~78 % (at 700 nm), which was on par with that of the normal cornea (80 %). The in vitro studies conducted using goat corneal stromal cells demonstrated the ability of both hydrogels to promote cell adhesion and proliferation. Expression of Vimentin and keratan sulphate validated the phenotype of keratocytes in the hydrogel. This 3D printed GelMA/MC hydrogel model mimics biophysical characteristics of the native corneal stroma, which may hold promise for clinical corneal stromal tissue engineering., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Testicular tissue culture in vitro is considered an important tool for the study of spermatogenesis and the treatment of male infertility. Although agarose hydrogel is commonly used in testicular tissue culture, the efficiency of spermatogenesis in vitro is limited. In this study, testicular tissues from adult mice were cultured using a gas-liquid interphase method based on agarose (Agarose), gelatin methacryloyl (GelMA), alginate methacryloyl (AlgMA), dextran methacryloyl (DexMA), and mixture GelMA-Agarose, AlgMA-Agarose, and DexMA-Agarose hydrogels, respectively, for 32 days in vitro. The integrity of the seminiferous tubules, the density and proportions of spermatogonia, spermatocytes, Sertoli cells, and testosterone concentrations were quantified and compared between groups. Properties of different hydrogels including compression modulus, Fourier Infrared Spectroscopy (FITR) spectra, pore size, water absorption, and water retention were tested to investigate how biochemical and physical properties of hydrogels affect the results of testicular tissue culture. The results indicate that testicular tissues cultured on AlgMA exhibited the highest seminiferous tubule integrity rate (0.835 ± 0.021), the presence of a high density of spermatocytes (2107.627 ± 232.082/mm 2 ), and a high proportion of SOX9-positive well-preserved seminiferous tubules (0.473 ± 0.047) compared to all remaining experimental groups on day 32. This may be due to the high water content of AlgMA reducing the toxic effect of oxygen on testicular tissue. In the later period of culture, testicular tissues cultured on DexMA, not DexMA-Agarose, produced significantly more testosterone (18.093 ± 3.302 ng/mL) than the other groups, suggesting that DexMA is friendly to Leydig cells. Our study provides a new idea for the optimization of the gas-liquid interphase method for achieving in vitro spermatogenesis, facilitating the future achievement of efficient in vitro spermatogenesis in more species, including humans., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cardiovascular diseases, particularly myocardial infarction (MI), remain a leading cause of morbidity and mortality worldwide. Current treatments for MI, more palliative than curative, have limitations in reversing the disease completely. Tissue engineering (TE) has emerged as a promising strategy to address this challenge and may lead to improved therapeutic approaches for MI. Gelatin-based scaffolds, including gelatin and its derivative, gelatin methacrylate (GelMA), have attracted significant attention in cardiac tissue engineering (CTE) due to their optimal physical and biochemical properties and capacity to mimic the native extracellular matrix (ECM). CTE mainly recruits two classes of gelatin/GelMA-based scaffolds: hydrogels and nanofibrous. This article reviews state-of-the-art gelatin/GelMA-based hybrid scaffolds currently applied for CTE and regenerative therapy. Hybrid scaffolds, fabricated by combining gelatin/GelMA hydrogel or nanofibrous scaffolds with other materials such as natural/synthetic polymers, nanoparticles, protein-based biomaterials, etc., are explored for enhanced cardiac tissue regeneration functionality. The engraftment of stem/cardiac cells, bioactive molecules, or drugs into these hybrid systems shows great promise in cardiac tissue repair and regeneration. Finally, the role of gelatin/GelMA scaffolds combined with the 3D bioprinting strategy in CTE will also be briefly highlighted., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Cardiac failure can be a life-threatening condition that, if left untreated, can have significant consequences. Functional hydrogel has emerged as a promising platform for cardiac tissue engineering. In the proposed study, gelatin methacrylate (GelMA) and alginate, as a primary matrix to maintain cell viability and proliferation, and polypyrrole and carboxyl-graphene, to improve mechanical and electrical properties, are thoroughly evaluated. Initially, a polymer blend of GelMA/Alginate (1:1) was prepared, and then the addition of 2-5 wt% of polypyrrole was evaluated. Next, the effect of incorporating graphene-carboxyl nanosheets (1, 2, and 3 wt%) within the optimized scaffold with 2 wt% polypyrrole was thoroughly studied. The electrical conductivity of the hydrogels was significantly enhanced from 0.0615 ± 0.007 S/cm in GelMA/alginate to 0.124 ± 0.04 S/cm with the addition of 5 wt% polypyrrole. Also, 3 wt% carboxyl graphene improved the electrical conductivity to 0.27 ± 0.09 S/cm. The compressive strength of carboxyl-graphene-containing hydrogel was in the range of 175 to 520 kPa, and tensile strength was 61 and 133 kPa. The simplicity and low-cost fabrication, tunable mechanical properties, optimal electrical conductivity, blood compatibility, and non-cytotoxicity of GelMA/alginate/polypyrrole/graphene biocomposite hydrogel is a promising construct for cardiac tissue engineering., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Methods in nerve tissue engineering for peripheral nerve regeneration have recently focused on the application of rapid three-dimensional (3D) printing with resorbable and degradable biomaterial ink. The technique aims to design and fabricate nerve guidance conduits (NGCs) to repair large-gap nerve injuries. In this study, Digital light processing (DLP)-based printing of multichannel NGCs for peripheral nerve regeneration by using gelatin methacrylate (GelMA) hydrogels was presented. Multichannel NGCs with different inner diameters were successfully fabricated using desired patterns, and optimal printing parameters were investigated. In vitro co-culturing of PC12 cells demonstrated that NGCs supported the survival, proliferation, and migration of neural cells along the longitudinal channel. The results also showed that neural crest stem cells could be induced to differentiate into neurons on fabricated conduits. This method will provide a rapid and accurate manner to fabricate complex NGCs, which can be potentially used to repair peripheral nerves.

Nanoliposomes are widely used as delivery vehicles for active compounds. Nanoliposomes from rapeseed phospholipids were incorporated into interpenetrating polymer network hydrogels of gelatin methacryloyl and alginate. The multiscale physicochemical properties of the hydrogels are studied both on the surface and through the thickness of the 3D network. The obtained composite hydrogels exhibited strong mechanical properties and a highly porous surface. The blend ratio, as well as the concentration of nanoliposomes, affects the properties of the hydrogels. Nanofunctionalized hydrogels induced keratinocyte ​growth. These advantageous characteristics may open up many applications of the developed hydrogels in drug delivery and tissue engineering.

This data article provides further detailed information related to our research article titled “Microfabricated Blood Vessels Undergo Neovascularization” (DiVito et al., 2017) [1], in which we report fabrication of human blood vessels using hydrodynamic focusing (HDF). Hydrodynamic focusing with advection inducing chevrons were used in concert to encase one fluid stream within another, shaping the inner core fluid into ‘bullseye-like” cross-sections that were preserved through click photochemistry producing streams of cellularized hollow 3-dimensional assemblies, such as human blood vessels (Daniele et al., 2015a, 2015b, 2014, 2016; Roberts et al., 2016) [2–6]. Applications for fabricated blood vessels span general tissue engineering to organ-on-chip technologies, with specific utility in in vitro drug delivery and pharmacodynamics studies. Here, we report data regarding the construction of blood vessels including cellular composition and cell positioning within the engineered vascular construct as well as functional aspects of the tissues.

The therapeutic role of tendon stem cells (TSCs) in tendon-related injuries has been well documented. Small extracellular vesicles (sEVs) are being increasingly used as new biotherapeutic agents for various diseases. Therefore, the potential function of TSC-sEVs in tendon injury repair warrants further investigation. In this study, we explored the effects of TSC-sEVs on TSC proliferation, migration, and differentiation in vitro in an autocrine manner. We further used a novel exosomal topical treatment with TSC-sEVs loaded with gelatin methacryloyl (GelMA) hydrogel in vivo; we mixed sufficient amounts of TSC-sEVs with GelMA hydrogel to cover the damaged molded Achilles tendon tissue and then exposed them to UV irradiation for coagulation. GelMA loading ensured that TSC-sEVs were slowly released at the injury site over a long period, thereby achieving their full local therapeutic effects. Treatment with TSC-sEVs loaded with GelMA significantly improved the histological score of the regenerated tendon by increasing the tendon expression while inhibiting the formation of excessive ossification and improving the mechanical properties of the tissue. Moreover, miRNA sequencing in TSC-sEVs, TSCs, and TSCs receiving sEVs revealed that TSC-sEVs altered the miRNA expression profile of TSCs, with increased expression of miR-145-3p. In conclusion, our study demonstrates that TSC-sEVs can play a key role in treating tendon injuries and that loading them with GelMA can enhance their effect in vivo. Moreover, miR-145-3p has a major functional role in the effect of TSC-sEVs. This study offers new therapeutic ideas for the local treatment of Achilles tendon injuries using sEVs. STATEMENT OF SIGNIFICANCE: In this study, we demonstrated that TSC-sEVs play a key role in treating tendon injuries and that loading them with GelMA hydrogel can act as a fixation and slow release in vivo. Moreover, it identifies the major functional role of miR-145-3p in the effect of TSCs that were identified and validated by miRNA sequencing. Our study provides a basis for further research on GelMA slow-release assays that have potential clinical applications. It offers new therapeutic ideas for the local treatment of Achilles tendon injuries using TSC-sEVs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Bioactive glass (BG) has been widely employed in the field of bone tissue engineering owing to its osteoconductive properties. These properties increase the stiffness and bioactivity of polymeric hydrogels, making them ideal for the repair, replacement, and regeneration of damaged bones. In this study, we investigated the effects of incorporating silanized 45S5 bioactive glass (Si-BG) into gelatin methacrylate (GelMA) hydrogel (GelMA/Si-BG) for potential bone tissue engineering. Our findings revealed that crosslinking GelMA with Si-BG had a striking increase in bioactivity with and without osteogenic induction of human mesenchymal stem cells (hMSCs) when compared to GelMA/BG hydrogels. Meanwhile, both GelMA/Si-BG and GelMA/BG hydrogels were able to maintain the cell viability of hMSC for up to 14 days. Additionally, GelMA/Si-BG hydrogels were shown to have a significantly higher compressive modulus than GelMA/BG hydrogels. This study has demonstrated the introduction of silanized 45S5 BG into GelMA hydrogel bioactivity and mechanical properties of GelMA hydrogels, exemplifying the potential application of silanization of BG in bone tissue engineering., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The incidence of osteochondral defect is increasing year by year, but there is still no widely accepted method for repairing the defect. Hydrogels loaded with bioactive molecules have provided promising alternatives for in-situ osteochondral regeneration. Kartogenin (KGN) is an effective and steady small molecule with the function of cartilage regeneration and protection which can be further boosted by TGF-β. However, the high cost, instability, and immunogenicity of TGF-β would limit its combined effect with KGN in clinical application. In this study, a composite hydrogel CM-KGN@GelMA, which contained TGF-β1 analog short peptide cytomodulin-10 (CM-10) and KGN, was fabricated. The results indicated that CM-10 modified on GelMA hydrogels exerted an equivalent role in enhancing chondrogenesis as TGF-β1, and this effect was also boosted when combined with KGN. Moreover, it was revealed that CM-10 and KGN had a synergistic effect on promoting the chondrogenesis of BMSCs by up-regulating the expression of RUNX1 and SOX9 at both mRNA and protein levels in vitro. Finally, the composite hydrogel exhibited a satisfactory osteochondral defect repair effect in vivo, showing similar structures close to the native tissue. Taken together, this study has revealed that CM-10 may serve as an alternative for TGF-β1 and can collaborate with KGN to accelerate chondrogenesis, which suggests that the fabricated CM-KGN@GelMA composite hydrogel can be acted as a potential scaffold for osteochondral defect regeneration. STATEMENT OF SIGNIFICANCE: Kartogenin and TGF-β have shown great value in promoting osteochondral defect regeneration, and their combined application can enhance the effect and show great potential for clinical application. Herein, a functional CM-KGN@GelMA hydrogel was fabricated, which was composed of TGF-β1 mimicking peptide CM-10 and KGN. CM-10 in hydrogel retained an activity like TGF-β1 to facilitate BMSC chondrogenesis and exhibited boosting chondrogenesis by up-regulating RUNX1 and SOX9 when being co-applied with KGN. In vivo, the hydrogel promoted cartilage regeneration and subchondral bone reconstruction, showing similar structures as the native tissue, which might be vital in recovering the bio-function of cartilage. Thus, this study developed an effective scaffold and provided a promising way for osteochondral defect repair., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Heart failure is a critical and ultimate phase of cardiovascular ailment that leads to a considerable incidence of disability and mortality. Among various factors contributing to heart failure, myocardial infarction is one of the most frequent and significant causes, which is still difficult to manage effectively. An innovative therapeutic strategy, namely a 3D bio-printed cardiac patch, has recently emerged as a promising approach to substitute damaged cardiomyocytes in a localized infarct region. Nevertheless, the efficacy of this treatment primarily relies on the long-term viability of the transplanted cells. In this study, we aimed to construct acoustically sensitive nano oxygen carriers to improve cell survival inside the bio-3D printed patch. In this study, we initially created nanodroplets capable of phase transition triggered by ultrasound and integrated them into GelMA (Gelatin Methacryloyl) hydrogels, which were then employed for 3D bioprinting. After adding nanodroplets and ultrasonic irradiation, numerous pores appeared inside the hydrogel with improved permeability. We further encapsulated hemoglobin into nanodroplets (ND-Hb) to construct oxygen carriers. Results of in vitro experiments showed the highest cell survival within the patch of ND-Hb irradiated by the low-intensity pulsed ultrasound (LIPUS) group. The genomic analysis discovered that the increased survival of seeded cells within the patch might be related to the protection of mitochondrial function owing to the improved hypoxic state. Eventually, in vivo studies revealed that the LIPUS+ND-Hb group had improved cardiac function and increased revascularization after myocardial infarction. To summarize, our study successfully improved the permeability of the hydrogel in a non-invasive and efficient manner, facilitating the exchange of substances in the cardiac patch. Moreover, ultrasound-controlled oxygen release augmented the viability of the transplanted cells and expedited the repair of infarcted tissues., Competing Interests: Declaration of competing interest All authors have no conflict of interest to declare., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Three-dimensional (3D) printing of soft biomaterials facilitates the progress of personalized medicine. The development for different forms of 3D-printable biomaterials can promotes the potential manufacturing for artificial organs and provides biomaterials with the required properties. In this study, gelatin methacryloyl (GelMA) and dialdehyde-functionalized polyurethane (DFPU) were combined to create a double crosslinking system and develop 3D-printable GelMA-PU biodegradable hydrogel and cryogel. The GelMA-PU system demonstrates a combination of self-healing ability and 3D printability and provides two distinct forms of 3D-printable biomaterials with smart functions, high printing resolution, and biocompatibility. The hydrogel was printed into individual modules through an 80 µm or larger nozzle and further assembled into complex structures through adhesive and self-healing abilities, which could be stabilized by secondary photocrosslinking. The 3D-printed hydrogel was adhesive, light transmittable, and could embed a light emitting diode (LED). Furthermore, the hydrogel laden with human mesenchymal stem cells (hMSCs) was successfully printed and showed cell proliferation. Meanwhile, 3D-printed cryogel was achieved by printing on a subzero temperature platform through a 210 µm nozzle. After secondary photocrosslinking and drying, the cryogel was deliverable through a 16-gage (1194 µm) syringe needle and can promote the proliferation of hMSCs. The GelMA-PU system extends the ink pool for 3D printing of biomaterials and has potential applications in tissue engineering scaffolds, minimally invasive surgery devices, and electronic wound dressings. STATEMENT OF SIGNIFICANCE: The 3D-printable biomaterials developed in this work are GelMA-based ink with smart funcitons and have potentials for various customized medical applications. The synthesized GelMA-polyurethane double network hydrogel can be 3D-printed into individual modules (e.g., 11 × 11 × 5 mm 3 ) through an 80 μm or larger size nozzle, which are then assembled into a taller structure over five times of the initial height by self-healing and secondary photocrosslinking. The hydrogel is adhesive, light transmittable, and biocompatible that can either carry human mesenchymal stem cells (hMSCs) as bioink or embed a red light LED (620 nm) with potential applications in electronic skin dressing. Meanwhile, the 3D-printed highly compressible cryogel (e.g., 6 × 6 × 1 mm 3 ) is deliverable by a 16-gage (1194 μm) syringe needle and supports the proliferation of hMSCs also., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Bone tissue is a natural composite, exhibiting complicated structures and unique mechanical/biological properties. With an attempt of mimicking the bone tissue, a novel inorganic-organic composite scaffolds (ZrO 2 -GM/SA) was designed and prepared via the vacuum infiltration method and the single/double cross-linking strategy by blending GelMA/alginate (GelMA/SA) interpenetrating polymeric network (IPN) into the porous zirconia (ZrO 2 ) scaffold. The structure, morphology, compressive strength, surface/interface properties, and biocompatibility of the ZrO 2 -GM/SA composite scaffolds were characterized to evaluate the performance of the composite scaffolds. Results showed that compared to ZrO 2 bare scaffolds with well-defined open pores, the composite scaffolds prepared by double cross-linking of GelMA hydrogel and sodium alginate (SA) presented a continuous, tunable and honeycomb-like microstructure. Meanwhile, GelMA/SA showed favorable and controllable water-uptake capacity, swelling property and degradability. After the introduction of IPN components, the mechanical strength of composite scaffolds was further improved. The compressive modulus of composite scaffolds was significantly higher than the bare ZrO 2 scaffolds. In addition, ZrO 2 -GM/SA composite scaffolds had highly biocompatibility and displayed a potent proliferation and osteogenesis of MC3T3-E1 pre-osteoblasts compared to bare ZrO 2 scaffolds and ZrO 2 -GelMA composite scaffolds. At the same time, ZrO 2 -10GM/1SA composite scaffold regenerated significantly greater bone than other groups in vivo. This study demonstrated that the proposed ZrO 2 -GM/SA composite scaffolds had great research and application potential in bone tissue engineering., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Electrically modulated delivery of proteins provides an avenue to target local tissues specifically and tune the dose to the application. This approach prolongs and enhances activity at the target site whilst reducing off-target effects associated with systemic drug delivery. The work presented here explores an electrically active composite material comprising of a biocompatible hydrogel, gelatin methacryloyl (GelMA) and a conducting polymer, poly(3,4-ethylenedioxythiophene), generating a conducting polymer hydrogel. In this paper, the key characteristics of electroactivity, mechanical properties, and morphology are characterized using electrochemistry techniques, atomic force, and scanning electron microscopy. Cytocompatibility is established through exposure of human cells to the materials. By applying different electrical-stimuli, the short-term release profiles of a model protein can be controlled over 4 h, demonstrating tunable delivery patterns. This is followed by extended-release studies over 21 days which reveal a bimodal delivery mechanism influenced by both GelMA degradation and electrical stimulation events. This data demonstrates an electroactive and cytocompatible material suitable for the delivery of protein payloads over 3 weeks. This material is well suited for use as a treatment delivery platform in tissue engineering applications where targeted and spatio-temporal controlled delivery of therapeutic proteins is required. STATEMENT OF SIGNIFICANCE: Growth factor use in tissue engineering typically requires sustained and tunable delivery to generate optimal outcomes. While conducting polymer hydrogels (CPH) have been explored for the electrically responsive release of small bioactives, we report on a CPH capable of releasing a protein payload in response to electrical stimulus. The composite material combines the benefits of soft hydrogels acting as a drug reservoir and redox-active properties from the conducting polymer enabling electrical responsiveness. The CPH is able to sustain protein delivery over 3 weeks, with electrical stimulus used to modulate release. The described material is well suited as a treatment delivery platform to deliver large quantities of proteins in applications where spatio-temporal delivery patterns are paramount., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Polymer-based composite scaffolds are an attractive class of biomaterials due to their suitable physical and mechanical performance as well as appropriate biological properties. When such composites contain osteoinductive ceramic nanopowders, it is possible, in principle, to stimulate the seeded cells to differentiate into osteoblasts. However, reproducibly fabricating and developing an appropriate niche for cells' activities in three-dimensional (3D) scaffolds remains a challenge using conventional fabrication techniques. Additive manufacturing provides a new strategy for the fabrication of complex 3D structures. Here, an extrusion-based 3D printing method was used to fabricate the Alginate (Alg)/Tri-calcium silicate (C 3 S) bone scaffolds. To improve physical and biological attributes, scaffolds were coated with gelatin methacryloyl (GelMA), a biocompatible viscose hydrogel. Conducting a combination of experimental techniques and molecular dynamics simulations, it is found that the composition ratio of Alg/C 3 S governs intermolecular interactions among the polymer and ceramic, affecting the product performance. Investigating the effects of various C 3 S amounts in the bioinks, the 90/10 composition ratio of Alg/C 3 S is known as the optimum content in developed bioinks. Accordingly, the printability of high-viscosity inks is boosted by improved hierarchical interactions among assemblies, which in turn leads to better nanoscale alignment in extruded macroscopic filaments. Conducting multiple tests on specimens, the GelMA-coated Alg/C 3 S scaffolds (with a composition ratio of 90/10) were shown to have improved mechanical qualities and cell adhesion, spreading, proliferation, and osteogenic differentiation, compared to the bare scaffolds, making them better candidates for further future research. Overall, the in-silico and in vitro studies of GelMA-coated 3D-printed Alg/C 3 S scaffolds open new aspects for biomaterials aimed at the regeneration of large- and complicated-bone defects through modifying the extrusion-based 3D-printed constructs., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2022. Published by Elsevier B.V.)

In bone tissue engineering, vascularization is one of the critical factors that limit the effect of biomaterials for bone repair. While various approaches have been tried to build vascular networks in bone grafts, lack of endothelialization still constitutes a major technical hurdle. In this study, we have developed a facile technique to fabricate endothelialized biomimetic microvessels (BMVs) from alginate-collagen composite hydrogels within a single step using microfluidic technology. BMVs with different sizes could be readily prepared by adjusting the flow rate of microfluids. All BMVs supported perfusion and outward penetration of substances in the tube. Endothelial cells could adhere and proliferate on the inner wall of tubes. It was also found that the expression of CD31 and secretion of BMP-2 and PDGF-BB were higher in the rat umbilical vein endothelial cells (RUVECs) in BMVs than those cultured on hydrogel. When co-cultured with bone marrow mesenchymal stem cells (BMSCs), endothelialized BMVs promoted the osteogenic differentiation of BMSCs compared to those in acellular BMV group. In vivo, markedly enhanced new bone formation was achieved by endothelialized BMVs in a rat critical-sized calvarial defect model compared to those with non-endothelialized BMVs or without BMVs. Together, findings from both in vitro and in vivo studies have proven that endothelialized BMVs function to facilitate osteogenesis and promote bone regeneration, and therefore might present an effective strategy in bone tissue engineering. STATEMENT OF SIGNIFICANCE: In bone tissue engineering, limited vascularization is one of the critical factors that limit the effect of biomaterials for bone repair. In this study, we developed a facile technique to fabricate endothelialized biomimetic microvessels (BMVs) from alginate-collagen composite hydrogels within a single step using microfluidic technology. Both in vitro and in vivo studies have proven that endothelialized BMVs function to facilitate osteogenesis and promote bone regeneration, and therefore might present an effective strategy in bone tissue engineering., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Ltd.)

Peripheral nerve injury can cause various degrees of damage to the morphological structure and physiological function of the peripheral nerve. At present, compared with "gold standard" autologous nerve transplantation, tissue engineering has certain potential for regeneration and growth; however, achieving oriented guidance is still a challenge. In this study, we used 3D bioprinting to construct a nerve scaffold of RSC96 cells wrapped in sodium alginate/gelatin methacrylate (GelMA)/bacterial nanocellulose (BNC) hydrogel. The 5% sodium alginate+5% GelMA+0.3% BNC group had the thinnest lines among all groups after printing, indicating that the inherent shape of the scaffold could be maintained after adding BNC. Physical and chemical property testing (Fourier transform infrared, rheometer, conductivity, and compression modulus) showed that the 5% alginate+5% GelMA+0.3% BNC group had better mechanical and rheological properties. Live/dead cell staining showed that no mass cell death was observed on days 1, 3, 5, and 7 after printing. In the 5% alginate+5% GelMA group, the cells grew and formed linear connections in the scaffold. This phenomenon was more obvious in the 5% alginate+5% GelMA+0.3% BNC group. In the 5% alginate+5% GelMA+0.3% BNC group, S-100β immunofluorescence staining and cytoskeleton staining showed oriented growth. Polymerase chain reaction (PCR) array results showed that mRNA levels of related neurofactors ASCL1, POU3F3, NEUROG1, DLL1, NOTCH1 and ERBB2 in the 5%GelMA+0.3%BNC group were higher than those of other groups. Four weeks after implantation in nude mice, RSC96 cells grew and proliferated well, blood vessels grew, and S-100β immunofluorescence was positive. These results indicate that a 3D-bioprinted sodium alginate/GelMA/BNC composite scaffold can improve cell-oriented growth, adhesion and the expression of related factors. This 3D-bioprinted composite scaffold has good biocompatibility and is expected to become a new type of scaffold material in the field of neural tissue engineering., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Tissue engineering strategies are widely used to model and study the bone marrow microenvironment in healthy and pathological conditions. Yet, while bone function highly depends on mechanical stimulation, the effects of biomechanical stimuli on the bone marrow niche, specifically on bone marrow adipose tissue (BMAT) is poorly understood due to a lack of representative in vitro loading models. Here, we engineered a BMAT analog made of a GelMA (gelatin methacryloyl) hydrogel/medical-grade polycaprolactone (mPCL) scaffold composite to structurally and biologically mimic key aspects of the bone marrow microenvironment, and exploited an innovative bioreactor to study the effects of mechanical loading. Highly reproducible BMAT analogs facilitated the successful adipogenesis of human mesenchymal bone marrow stem cells. Upon long-term intermittent stimulation (1 Hz, 2 h/day, 3 days/week, 3 weeks) in the novel bioreactor, cellular proliferation and lipid accumulation were similar to unloaded controls, yet there was a significant reduction in the secretion of adipokines including leptin and adiponectin, in line with clinical evidence of reduced adipokine expression following exercise/activity. Ultimately, this innovative loading platform combined with reproducibly engineered BMAT analogs provide opportunities to study marrow physiology in greater complexity as it accounts for the dynamic mechanical microenvironment context., (Copyright © 2021. Published by Elsevier B.V.)

Hydrogels were prepared by mixing protein and carbohydrate-based biopolymers to increase the mechanical properties and efficient cell adhesion and proliferation for wound healing applications. Microcrystalline cellulose (MCC) and its 6-deoxy-aminocellulose derivatives (6-deoxy-6-hydrazide Cellulose (Cell-Hyd), 6-deoxy-6-diethylamide Cellulose (Cell-DEA), and 6-deoxy-6-diethyltriamide Cellulose (Cell-DETA)) were embedded in methacrylated gelatin (GelMA). GelMA and 6-deoxy-aminocellulose derivatives were synthesized and characterized by spectroscopic techniques. MCC and cellulose derivatives embedded GelMA gels were characterized by FTIR, SEM and Tensile mechanical testing. SEM images revealed that, porosity of the amine MCC incorporated GelMA was decreased compared to GelMA and MCC incorporated GelMA. Tensile strain of GelMA 61.30% at break was increased to 64.3% in case of GelMA/Cell-HYD. In vitro cytocompatibility and cell proliferation using NIH-3T3 cell lines showed cell density trend on scaffold as GelMA/Cell-DETA>GelMA/Cell-Hyd > GelMA. Scratch assay for wound healing revealed that GelMA/Cell-DETA showed complete wound closure, while GelMA/Cell-Hyd and GelMA exhibited 85.7%, and 66.1% wound healing, respectively in 8 h. In vivo tests on rats revealed that GelMA/Cell-DETA exhibited 98% wound closure on day 9, whereas GelMA/Cell-Hyd exhibited 97.7% and GelMA 66.1% wound healing on day 14. Our findings revealed that GelMA embedded amine MCC derivatives hydrogels can be applied for achieving accelerated wound healing., (Copyright © 2021 Elsevier B.V. All rights reserved.)

We develop a robust micro-patterned double-layer film that can adhere firmly to the tissue and provide a sustained release of ascorbic acid (AA) for corneal regeneration. This double-layer film consists of a AA reservoir sodium alginate (SA) adhesive and an anisotropic layer made of micro-patterned silk nanofibrils (SNF) incorporated gelatin methacrylate (GelMA) (S/G). The S/G layer facilitates the adhesion and orientation of corneal stroma cells, depending on the pattern sizes (50 μm (P1) and 100 (P2) μm). Results reveal that more than 90% and 80% of the cells are located at angles close to the vertical axis (0-20°) in the sample with the smaller and larger pattern size, respectively. The mechanical robustness and 90% light transmission of this hybrid film originate from the micro-patterned S/G layer. However, the micro-pattern size does not show a significant role in the mechanical properties of hybrid films (tensile strength of S/G-SA, S/G-SA(P 1 ), and S/G-SA(P 2 ) is 3.4 ± 0.1 MPa, 3.6 ± 0.6 MPa and 3.3 ± 0.2 MPa, respectively). In addition, the strong adhesion to the tissue of this double-layer film is related to the alginate layer. AA can release in a controlled manner, which can significantly promote corneal stroma cells' attachment, alignment, and proliferation compared to the control (AA-free micro-patterned film). Our results reveal that this innovative multifunctional S/G-SA + AA film can be a proper candidate for use in stroma tissue engineering of the human cornea., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The human amniotic membrane (HAM) collagen matrix derived from human placenta can be decellularized (dHAM) to form a natural biocompatible material. dHAM has different bioactive substances and has been used widely in human tissue engineering research. However, dHAM has some disadvantages, e.g., poor mechanical properties, easy degradation and inconvenient operation and use, so it is not conducive to large-area or full-thickness skin defect healing. To overcome these limitations, for the first time, dHAM was grafted with methacrylic anhydride (MA) to form photocrosslinked dHAM methacrylate (dHAMMA); dHAMMA was then blended with methacrylated gelatin (GelMA), followed by the addition of a photosensitizer for photocrosslinking to obtain the fast-forming GelMA-dHAMMA composite hydrogel. Further, GelMA-dHAMMA was found to have the porous structure of a bicomponent polymer network and good physical and chemical properties. In vitro experiments, GelMA-dHAMMA was found to promote fibroblast proliferation and α-smooth muscle actin (α-SMA) expression. In vivo investigations also demonstrated that GelMA-dHAMMA promotes wound collagen deposition and angiogenesis, and accelerates tissue healing. GelMA-dHAMMA inherits the good mechanical properties of GelMA and maintains the biological activity of the amniotic membrane, promoting the reconstruction and regeneration of skin wounds. Thus, GelMA-dHAMMA can serve as a promising biomaterial in skin tissue engineering. STATEMENT OF SIGNIFICANCE: Since the early 20th century, the human amniotic membrane (HAM) has been successfully used for trauma treatment and reconstruction purposes. dHAM has different bioactive substances and has been used widely in human tissue-engineering research. In this work, the dHAM and gelatin were grafted and modified by using methacrylic anhydride (MA) to form photocrosslinked dHAMMA and methacrylated gelatin (GelMA). Then, the dHAMMA and GelMA were blended with a photosensitizer to form the GelMA-dHAMMA composite hydrogel derived from gelatin-dHAM. GelMA-dHAMMA exhibits a bicomponent-network (BCN) interpenetrating structure. dHAM dydrogel has advantages, e.g., good mechanical properties, slow degradation and convenient operation, so it is conducive to large-area or full-thickness skin defect healing., Competing Interests: Declaration of Competing Interest The authors declare no competing financial interest., (Copyright © 2021 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

A functional vascular supply is a key component of any large-scale tissue, providing support for the metabolic needs of tissue-remodeling cells. Although well-studied strategies exist to fabricate biomimetic scaffolds for bone regeneration, success rates for regeneration in larger defects can be improved by engineering microvascular capillaries within the scaffolds to enhance oxygen and nutrient supply to the core of the engineered tissue as it grows. Even though the role of calcium and phosphate has been well understood to enhance osteogenesis, it remains unclear whether calcium and phosphate may have a detrimental effect on the vasculogenic and angiogenic potential of endothelial cells cultured on 3D printed bone scaffolds. In this study, we presented a novel dual-ink bioprinting method to create vasculature interwoven inside CaP bone constructs. In this method, strands of a CaP ink and a sacrificial template material was used to form scaffolds containing CaP fibers and microchannels seeded with vascular endothelial and mesenchymal stem cells (MSCs) within a photo-crosslinkable gelatin methacryloyl (GelMA) hydrogel material. Our results show similar morphology of growing vessels in the presence of CaP bioink, and no significant difference in endothelial cell sprouting was found. Furthermore, our initial results showed the differentiation of hMSCs into pericytes in the presence of CaP ink. These results indicate the feasibility of creating vascularized bone scaffolds, which can be used for enhancing vascular formation in the core of bone scaffolds., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The aim of this investigation was to engineer metformin (MF)-loaded mesoporous silica nanospheres (MSNs)-laden gelatin methacryloyl (GelMA) photocrosslinkable hydrogels and test their effects on the mechanical properties, swelling ratio, drug release, cytocompatibility, and osteogenic differentiation of stem cells from human exfoliated deciduous teeth (SHEDs). As-received and carboxylated MSNs (MSNs-COOH) were characterized by scanning and transmission electron microscopies (SEM and TEM), as well as Fourier-transform infrared spectroscopy (FTIR) prior to hydrogel modification. MF-MSNs-COOH were obtained by loading MF into MSNs at a 1:1 mass ratio. Upon MSNs-COOH laden-hydrogels fabrication, the mechanical properties, swelling ratio and MF release were evaluated. SHEDs were seeded on the hydrogels and cytocompatibility was examined. The effects of the MF-MSNs-COOH/GelMA on the osteogenic differentiation of SHEDs were measured by ALP activity, Alizarin Red assay, and Real-time PCR. Statistics were performed using one-way ANOVA (α = 0.05). Morphological (SEM and TEM) analyses of pristine and carboxylated MSNs revealed a mean particle size of 200 nm and 218 nm, respectively. Importantly, an intrinsic nanoporous structure was noticed. Incorporation of MSNs-COOH at 1.5 mg/mL in GelMA led to the highest compressive modulus and swelling ratio. The addition of MSNs-COOH (up to 3 mg/mL) in GelMA did not impact cell viability. The presence of MF in MSNs-COOH/GelMA significantly promoted cell proliferation. Significant upregulation of osteogenic-related genes (except OCN) were seen for modified (MSNs-COOH and MF-MSNs-COOH) hydrogels when compared to GelMA. Altogether, the engineered MF-MSNs-COOH/GelMA shows great promise in craniomaxillofacial applications as an injectable, cell-free and bioactive therapeutics for bone regeneration., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Management of chronic diabetic ulcers remains as a major challenge in healthcare which requires extensive multidisciplinary approaches to ensure wound protection, management of excess wound exudates and promoting healing. Developing wound healing patches that can act as a protective barrier and support healing is highly needed to manage chronic diabetic ulcers. In order to boost the wound healing potential of patch material, bioactive agents such as growth factors can be used. Porous membranes made of nanofibers generated using electrospinning have potential for application as wound coverage matrices. However, electrospun membranes produced from several biodegradable polymers are hydrophobic and cannot manage the excess exudates produced by chronic wounds. Gelatin-methacryloyl (GelMA) hydrogels absorb excess exudates and provide an optimal biological environment for the healing wound. Epidermal growth factor (EGF) promotes cell migration, angiogenesis and overall wound healing. Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) membranes provide microbial, thermal and mechanical barrier properties to the wound healing patch. Herein, we developed a biodegradable polymeric patch based on the combination of mechanically stable electrospun PHBV, GelMA hydrogel and EGF for promoting diabetic wound healing. In vitro and in vivo studies were carried out to evaluate the effect of developed patches on cell proliferation, cell migration, angiogenesis and wound healing. Our results showed that EGF loaded patches can promote the migration and proliferation of multiple types of cells (keratinocytes, fibroblasts and endothelial cells) and enhance angiogenesis. In situ development of the patch and subsequent in vivo wound healing study in diabetic rats showed that EGF loaded patches provide rapid healing compared to control wounds. Interestingly, 100 ng EGF per cm 2 of the patches was enough to provide favourable cellular response, angiogenesis and rapid diabetic wound healing. Overall results indicate that EGF loaded PHBV-GelMA hybrid patch could be a promising approach to promote diabetic wound healing., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Carbon nanotubes (CN) have been studied to treat spinal cord injuries because of its electrical properties and nanometric dimensions. This work aims to develop a photopolymerizable hydrogel containing CN functionalized with an anti-inflammatory molecule to be used in situ on spinal cord injuries. The CN functionalization step was done using the drug (formononetin). The nanocomposites were characterized by morphological analysis, FTIR, Raman Spectroscopy, thermal analysis and cytotoxicity assays (MTT and HET-CAM). The nanocomposites were incorporated into gelatin methacryloyl hydrogel and exposed to UV light for photopolymerization. The volume of the formulation and the UV exposition time were also analyzed. The CN characterization showed that formononetin acted as a functionalization agent. The functionalized CN showed safe characteristics and can be incorporated in photocrosslinkable formulation. The UV exposition time for the formulation photopolymerization was compatible with the cell viability and also occurred in the injury site., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: Over the past decades, a variety of biomaterials have been investigated in terms of their suitability for oral mucosa tissue engineering. The aim of this study was to compare collagen and GelMA hydrogels as connective tissue scaffolds for fibroblasts and as substrates for seeding and culture of oral epithelial keratinocyte cells., Methods: Human primary oral fibroblast and keratinocyte cells were isolated from gingival biopsies. The mixture of fibroblasts with GelMA or collagen gel were aliquoted within six-well tissue culture plate inserts and cross-linked using visible light or reconstitution buffer/heat, respectively. The viability of fibroblasts in the hydrogels was investigated after one and three days of cultivation using the PrestoBlue assay. Following the addition and culture of oral keratinocytes onto the connective tissue constructs, the tissue-engineered oral mucosa was assessed histologically., Results: The tissue viability assay shows that collagen hydrogels encapsulating fibroblasts displayed significantly higher cell viability than cell-laden GelMA constructs after 24 and 72 h ( p  < 0.05). A stratified and differentiated epithelium has formed on the surface of cell-laden collagen hydrogel but not on the surface of the GelMA-based substrate., Conclusion: Collagen-based scaffold offers superior biological properties compared to GelMA hydrogel in terms of oral fibroblast growth, as well as epithelial cell adhesion and differentiation. Therefore, collagen-based hydrogels remain the preferred choice for oral mucosa tissue engineering., Competing Interests: None., (© 2020 Craniofacial Research Foundation. Published by Elsevier B.V. All rights reserved.)

Repairing large segmental bone defects above a critical size remains challenging with high risk of delayed union or even non-union. From the perspective of bone development and clinical experience, periosteum plays an indispensable role in bone repair and reconstruction. In this study, we explored the feasibility of using preosteoblast-derived matrix (pODM) as a biomimetic periosteum. By culturing MC3T3-E1 cell sheet on poly(dimethylsiloxane) and performing decellularization, an integral cell-free sheet of pODM could be readily harvested. Bone marrow mesenchymal stem cells (BMSCs) adhered and proliferated well on pODM. In addition, pODM exhibited a chemotactic effect on BMSCs in a concentration-dependent manner and also promoted osteogenic differentiation of BMSCs. Following that, pODM was wrapped around a gelatin methacryloyl (GelMA) hydrogel to construct an engineered periosteum-bone substitute. A rabbit radius segmental bone defect model was used to examine the bone repair efficacy of pODM/GelMA. Upon implantation of pODM/GelMA construct for 12 weeks, the critical-sized bone defects completely healed with remarkable full reconstruction of medullary cavity at the radial diaphysis. Together, this work proposes a high potency of using precursor cell-derived matrix as a biomimetic periosteum, which preserves the beneficial biological factors while avoids the limitations of using exogenous cells for bone regeneration. Combining precursor cell-derived matrix with hydrogel may provide a promising periosteum-bone biomimetic substitute for bone repair. STATEMENT OF SIGNIFICANCE: Repairing large segmental bone defects above a critical size remains challenging. As the periosteum plays an essential role in bone repair, this study aimed to explore the use of preosteoblast-derived matrix (pODM), harvested from decellularized MC3T3-E1 cell sheet, as a biomimetic periosteum to facilitate bone repair. We found that in vitro, pODM exhibited considerable chemotactic effect and osteogenic induction capability to bone marrow mesenchymal stem cells (BMSCs). In vivo, implantation of pODM/gelatin methacryloyl (GelMA) constructs as engineered periosteum-bone substitutes effectively repaired the critical-sized segmental bone defects at rabbit radius. Surprisingly, remarkable full reconstruction of medullary cavity at the diaphysis was achieved. Therefore, combining pODM with hydrogel may provide a promising biomimetic substitute for bone repair., (Copyright © 2020. Published by Elsevier Ltd.)

This study demonstrates the effect of substrate's geometrical cues on viability and the efficacy of an anti-cancer drug, doxorubicin (DOX), on breast cancer cells. It is hypothesized that the surface topographical properties can mediate the cellular drug intake. Pseudo-three dimensional (3D) platforms were fabricated using imprinting technique from polydimethylsiloxane (PDMS) and gelatin methacryloyl (GelMA) hydrogel to recapitulate topography of cells' membranes. The cells exhibited higher viability on the cell-imprinted platforms for both PDMS and GelMA materials compared to the plain/flat counterparts. For instance, MCF7 cells showed a higher metabolic activity (11.9%) on MCF7-imprinted PDMS substrate than plain PDMS. The increased metabolic activity for the imprinted GelMA was about 44.2% compared to plain hydrogel. The DOX response of cells was monitored for 24 h. Although imprinted substrates demonstrated enhanced biocompatibility, the cultured cells were more susceptible to the drug compared to the plain substrates. In particular, MCF7 cells on imprinted PDMS and GelMA substrates showed 37% and 50% higher in cell death compared to the corresponding plain PDMS and GelMA, respectively. Interestingly, the drug susceptibility of the cells on the imprinted hydrogel was about 70% higher than the cells cultured on imprinted PDMS substrates. Having MCF7 cell-imprinted substrates, DOX responses of two other breast cancer cell lines, SKBR3 and ZR-75-1, were also evaluated. The results support that cell membrane curvature developed by multiscale topography is able to mediate intracellular signaling and drug intake. STATEMENT OF SIGNIFICANCE: Research in biological sciences and drug discovery mostly rely on two dimensional (2D) cell culture techniques which cannot provide a reliable physiologically relevant environment. Lack of extracellular matrix and a large shift in physicochemical properties of conventional 2D substrates can induce aberrant cellular behaviors. While chemical composition, topographical, and mechanical properties of substrates have remarkable impacts on drug susceptibility, gene expression, and protein synthesis, the most cell culture plates are from rigid and plain substrates. A number of (bio)polymeric 3D-platforms have been introduced to resemble innate cell microenvironment. However, their intricate culture protocols restrain their applications in demanding high-throughput drug screening. To address the above concerns, in the present study, a hydrogel-based pseudo-3D substrate with imprinted cell features has been introduced., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier Ltd.)

Gelatin methacryloyl (GelMA) is a chemically modified extracellular matrix (ECM)-derived biopolymer that is widely used for 3D fabrication of tissue engineering scaffolds. However, its tendency for physical gelation limits its use in aqueous 3D printing resins to low concentrations, yielding a poor printing resolution in stereolithography (SLA). To obtain a GelMA-based resin that can be printed into high-resolution tissue scaffolds, we formulated resins of fish and porcine-derived GelMA in formamide using GelMA alone or mixed with star-shaped poly(ε-caprolactone) methacrylate (PCL-MA). We identified GelMA resins and GelMA/PCL-MA hybrid resins with a ratio of 70/30 wt-% to yield a suitable viscosity for SLA at 32 °C and demonstrated the resolution of the new resins in SLA by 3D printing acellular human small intestine-mimicking tissue scaffolds. The presence of PCL-MA in the hybrid resins improved the 3D printing fidelity compared to the neat GelMA resins, while GelMA provided the hybrid materials with enhanced swelling and proliferation of seeded cells. We further demonstrated the transferability of our resin formulation to native organ-derived materials by successfully replacing GelMA in the hybrid resin with solubilized, methacryloyl-functionalized decellularized liver ECM (dECM-MA) and by 3D printing multi-layer dECM/PCL-MA hydrogels., Competing Interests: Declaration of competing interest A European patent application entitled “Photocurable composition” (European patent office, 19189493.0) by Laura Elomaa and Marie Weinhart was submitted on 31.07.2019 based on the results in the current manuscript. The authors declare no other conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The current gold standard for nasal reconstruction after rhinectomy or severe trauma includes transposition of autologous cartilage grafts in conjunction with coverage using an autologous skin flap. Harvesting autologous cartilage requires a major additional procedure that may create donor site morbidity. Major nasal reconstruction also requires sculpting autologous cartilages to form a cartilage framework, which is complex, highly skill-demanding and very time consuming. These limitations have prompted facial reconstructive surgeons to explore different techniques such as tissue engineered cartilage. This work explores the use of multi-material 3D bioprinting with chondrocyte-laden gelatin methacrylate (GelMA) and polycaprolactone (PCL) to fabricate constructs that can potentially be used for nasal reconstruction. In this study, we have investigated the effect of 3D manufacturing parameters including temperature, needle gauge, UV exposure time, and cell carrier formulation (GelMA) on the viability and functionality of chondrocytes in bioprinted constructs. Furthermore, we printed chondrocyte-laden GelMA and PCL into composite constructs to combine biological and mechanical properties. It was found that 20% w/v GelMA was the best concentration for the 3D bioprinting of the chondrocytes without comprising the scaffold's porous structure and cell functionality. In addition, the 3D bioprinted constructs showed neocartilage formation and similar mechanical properties to nasal alar cartilage after a 50-day culture period. Neocartilage formation was also observed in the composite constructs evidenced by the presence of glycosaminoglycans and collagen type II. This study shows the feasibility of manufacturing neocartilage using chondrocytes/GelMA/PCL 3D bioprinted porous constructs which could be applied as a method for fabricating implants for nose reconstruction., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Interfacial migration is central to multiple processes including morphogenesis and wound healing. However, the sensitivity of interfacial migration to properties of the interfacial microenvironment has not been adequately explored. Here, we address this question by tracking motility of 3T3 fibroblasts at the interface of two hydrogels. By sandwiching cells between two adhesive gels (composed of methacrylated gelatin) or between an adhesive and a non-adhesive gel (composed of gellan), we show that cells are more motile in case of the latter. By tuning the bulk stiffness of the gellan gel, we then show that motility is tuned in a stiffness-dependent manner. Fastest motility observed in case of the stiffest gel was associated with increased cell height, suggestive of stiffness-mediated cytoskeletal assembly. Inhibition of cell motility by contractile agonists and actin depolymerizing drugs is indicative of a mode of migration wherein cells combine contractile tractions exerted at their base and actin-based pushing forces on the top surface to propel themselves forward. Together, our results suggest that dorso-ventral adhesion anisotropy and stiffness can be collectively tuned to engineer interfacial migration., Statement of Significance: It is increasingly understood that cells migrate in vivo through confining spaces which typically occur as pores in the matrix and through naturally occurring interfaces that exist between neighbouring ECM fibers, or between the stroma and the vasculature. Such interfaces are also created when treating wounds on the skin surface by covering the wounds with adhesives. How multiple cues impact interfacial migration has not been adequately addressed. By studying cell migratory behaviour at the interface of two hydrogel substrates, we identify adhesivity and stiffness as two critical factors that can be tuned to maximize cell migration. We foresee a potential use of this knowledge in the design of tissue adhesives for wound healing applications., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

A major challenge for clinical use of skin substitutes is insufficient host tissue integration leading to loosening and partial necrosis of the implant. In this present study, a three-dimensional (3D) coculture system constructed using human umbilical cord mesenchymal stem cells (uc-MSCs) and umbilical vein endothelial cells (HUVECs) encapsulated in gelatin methacryloyl (GelMA) hydrogels was evaluated to determine the outcomes of cell-cell interactions in vitro and in vivo. The results revealed that GelMA hydrogels displayed minor cytotoxicity on both cell types. An uc-MSC:HUVEC ratio of 50:50 demonstrated the highest cell proliferation and expression of angiogenic markers. The supplement of basic fibroblast growth factors (bFGF) in coculture system further induced cell proliferation and gene expression in vitro. In vivo transplantation of this cocultured constructs efficiently enhanced the implant and host tissue integration. Additionally, the proliferation of keratinocytes was well maintained on GelMA hydrogels and the gene expression related to cell proliferation and differentiation was significantly increased in coculture system comparing to monoculture. Mechanistically, AKT signaling pathways were activated in cocultures. Our findings suggest that coculturing MSC and EC in GelMA hydrogels could be a promising approach to substantially improve the integration of exogenous skin substitutes and host tissues., Statement of Significance: In this study, the co-culture of uc-MSCs and HUVECs in photocrosslinkable GelMA hydrogels significantly enhanced host tissue integration. Cell proliferation, ECM deposition and angiogenic genes expression were all substantially improved in vitro and the excellent host tissue integration into the implanted tissue was observed in vivo. When served as a dermal layer, the scaffold with co-cultured cells enhanced the proliferation and differentiation of keratinocytes. AKT signaling was proved to be involved in the regulation of cell survival and fate determination. This work demonstrated the importance of 3D cell co-culture to facilitate host tissue integration that can be a promising approach for long-term survival of skin substitutes., (Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)