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We developed insulin loaded biomimetic microsphere by laccase-mediated crosslinking using a microfluidic device in the water-in-oil emulsion system as an injectable vehicle for the repair of sciatic tissue. Aqueous polymeric solution of phenol-substituted hyaluronic acid (HAPh) and collagen (ColPh) containing insulin and laccase flowed from the inner channel into oil flow within an outer channel which leads formation of hydrogel microsphere. The physical properties of prepared specimens including swelling rate, mechanical resistance and the prolonged release rate of microspheres proved applicability of fabricated vehicles for tissue engineering and drug delivery systems. The growth profile and behavior of cells in microspheres indicated cytocompatibility of the method and prepared vehicles for microtissue development. Histopathological examination revealed a significant increase in axonal regeneration, and remyelination process in injured sciatic nerve following treatment with HAPh/ColPh microspheres containing insulin compared to control groups. Also, the functional characteristic of sciatic tissue showed that the presence of biomimetic microsphere and insulin simultaneously had improved sciatic tissue functions including functional sciatic index (SFI) values, reaction to hot plate and muscle weight of rats. In summary, the results proved that composite biomimetic microspheres containing insulin effectively improved nerve regeneration in the rat model.

Carbon nanotube (CNT) and gelatin (Gela) molecules are effective substrates in promoting engineered cardiac tissue functions. This study developed a microfluidic-based encapsulation process for biomimetic hydrogel microcapsule fabrication. The hydrogel microcapsule was produced through a coaxial double orifice microfluidic technique and a water-in-oil emulsion system in two sequential processes. The phenol (Ph) substituted Gela (Gela-Ph) and CNT (CNT-Ph), respectively as cell-adhesive and electrically conductive substrates were incorporated in hyaluronic acid (HA)-based hydrogel through laccase-mediated crosslinking. The Cardiomyocyte-enclosing microcapsule fabricated and cellular survival, function, and possible difference in the biological activity of encapsulated cells within micro vehicles were investigated. The coaxial microfluidic method and Lac-mediated crosslinking reaction resulted in spherical vehicle production in 183 μm diameter at 500 capsules/min speed. The encapsulation process did not affect cellular viability and harvested cells from microcapsule proliferated well likewise subcultured cells in tissue culture plate. The biophysical properties of the designed hydrogel, including mechanical strength, swelling, biodegradability and electroconductivity upregulated significantly for hydrogels decorated covalently with Gela-Ph and CNT-Ph. The tendency of the microcapsule for the spheroid formation of cardiomyocytes inside the proposed microcapsule occurred 3 days after encapsulation. Interestingly, immobilized Gela-Ph and CNT-Ph promote cellular growth and specific cardiac markers. Overall, the microfluidic-based encapsulation technology and synthesized biomimetic substrates with electroconductive properties demonstrate desirable cellular adhesion, proliferation, and cardiac functions for engineering cardiac tissue.

Abnormal craniofacial bone fusion can lead to the generation of several congenital malformations such as cleft palate, craniosynostosis and craniofacial skeletal hypoplasia, which physically and mentally affect patients. Conventional approaches for the treatment of craniofacial malformations such as the transplantation of autologous bone grafts are not completely efficient and usually, patients suffer from various complications. In line with these statements, the advent of novel therapeutic approaches in human medicine is mandatory. Regarding the extent, size and severity of the bone malformation, supplementation and release of oxygen molecules into the affected sites are critical issues for successful osteogenesis. Here, tissue engineering modalities associated with oxygen supplementation and novel approaches associated with hydrogel synthesis were highlighted in terms of craniofacial malformations.

Spinal cord injury (SCI) has been implicated in neural cell loss and consequently motor/sensory impairment. In this study, we propose a new formation for fibrous mat fabrication from derivatives of poly vinyl alcohol (PVA) and gelatin (Gela) through horseradish peroxidase (HRP) mediated crosslinking, which provide sustained release of methylprednisolone (MP) that can be utilized as a therapeutic approach for SCI repair. The polymeric solution of PVA/Gela conjugated with phenolic moieties (PVAPh and GelaPh, respectively) containing HRP and MP, was fed into electrospinning system, using a syringe pump. The fabricated scaffolds were characterized in vitro and were implanted into the compression SCI rat model. After 28 days, the animals were evaluated in terms of remyelination and apoptosis and underwent behavioral test. The mechanical properties, hydrophobicity, and degradation rate of PVAPh/GelaPh fibrous mat were significantly promoted compared to PVAPh samples and could provide the desired structure for sustained release of MP. The seeded cells could adhere and proliferate on composite fibers, which indicates cytocompatibility of resultant PVAPh/GelaPh fibrous mat. The results showed a significant decrease in apoptotic neurons and a remarkable increase in remyelination in SCI + PVAPh/GelaPh + MP group compared to that in other groups. The behavioral test confirmed locomotor hind limb function amelioration after treatment. MP-loaded PVAPh/GelaPh mat developed through long-term release of MP and biocompatible fabricated mat could inhibit axonal demyelination, attenuated apoptosis, and improved functional outcome, which confirmed the potential of PVAPh/GelaPh + MP nanocomposites as a bioactive scaffold for SCI regeneration.

Glioblastoma multiform (GBM) is known as an aggressive glial neoplasm. Recently incorporation of mesenchymal stem cells with anti-tumor drugs have been used due to lack of immunological responses and their easy accessibility. In this study, we have investigated the anti-proliferative and apoptotic activity of atorvastatin (Ator) in combination of mesenchymal stem cells (MSCs) on GBM cells in vitro and in vivo. The MSCs isolated from rats and characterized for their multi-potency features. The anti-proliferative and migration inhibition of Ator and MSCs were evaluated by MTT and scratch migration assays. The annexin/PI percentage and cell cycle arrest of treated C6 cells were evaluated until 72 h incubation. The animal model was established via injection of C6 cells in the brain of rats and subsequent injection of Ator each 3 days and single injection of MSCs until 12 days. The growth rate, migrational phenotype and cell cycle progression of C6 cells decreased and inhibited by the interplay of different factors in the presence of Ator and MSCs. The effect of Ator and MSCs on animal models displayed a significant reduction in tumor size and weight. Furthermore, histopathology evaluation proved low hypercellularity and mitosis index as well as mild invasive tumor cells for perivascular cuffing without pseudopalisading necrosis and small delicate vessels in Ator + MSCs condition. In summary, Ator and MSCs delivery to GBM model provides an effective strategy for targeted therapy of brain tumor.

Current hyaluronic acid-based hydrogels often cause cytotoxicity to encapsulated cells and lack the adhesive property required for effective biomedical and tissue engineering applications. Provision of the cell-adhesive surface is an important requirement to improve its biocompatibility. An aqueous solution of hyaluronic acid possessing phenolic hydroxyl (HA-Ph) moieties is gellable via a horseradish peroxidase (HRP)-catalyzed oxidative cross-linking reaction. This study evaluates the effect of different degrees of cross-linked Ph moieties on cellular adhesiveness and proliferation on the resultant enzymatically cross-linked HA-Ph hydrogels. Mechanical characterization demonstrated that the compression force of engineered hydrogels could be tuned in the range of 0.05-35 N by changing conjugated Ph moieties in the precursor formulation. The water contact angle and water content show hydrophobicity of hydrogels increased with increasing content of cross-linked Ph groups. The seeded mouse embryo fibroblast-like cell line and human cervical cancer cell line, on the HA-Ph hydrogel, proved cell attachment and spreading with a high content of cross-linked Ph groups. The HA-Ph with a higher degree of Ph moieties shows the maximum degree of cell adhesion, spreading, and proliferation which presents this hydrogel as a suitable biomaterial for biomedical and tissue engineering applications.

Cartilage tissue engineering is the interdisciplinary science that will help to improve cartilage afflictions, such as arthrosis, arthritis, or following joints traumatic injuries. In the present work, we developed an injectable hydrogel which derived from decellularized extracellular matrix of sheep cartilage. Successful decellularization was evaluated by measuring the DNA, glycosaminoglycans (GAG), collagen contents, and histological analyses. There was a minor difference in GAG and collagen contents among natural cartilage and decellularized tissue as well as ultimate hydrogel. Rheological analysis showed that the temperature and gelation time of prepared hydrogel were 37°C and between 5 and 7 min, respectively. Mechanical properties evaluation indicated a storage modulus of 20 kPa. The results show that prepared hydrogel possessed cell-friendly microenvironment as confirmed via calcein staining and MTT assay. Also, cells were able to proliferate which observed by H&E and alcian blue staining. Cell attachment and proliferation at the surface of the decellularized hydrogel was apparent by Scanning Electron Microscope (SEM) images and microphotographs. Furthermore, the cells embedded within the hydrogel were able to differentiate into chondrocyte with limited evidence of hypertrophy and osteogenesis in utilized cells which proved by SOX9, CoL2, ACAN, and also CoL1 and CoL10 gene expression levels. In summary, the results suggest that developed novel injectable hydrogel from decellularized cartilage could be utilized as a promising substrate for cartilage tissue engineering applications.

One of the most effective approaches for treatment of cartilage involves the use of porous three-dimensional scaffolds, which are useful for improving not only cellular adhesion but also mechanical properties of the treated tissues. In this study, we manufactured a composite scaffold with optimum properties to imitate nasal cartilage attributes. Cartilage extracellular matrix (ECM) was used in order to improve the cellular properties of the scaffolds; while, chitosan and agarose were main materials that are used to boost the mechanical and rheological properties of the scaffolds. Furthermore, we explored the effect of the various weight ratios of chitosan, agarose, and ECM on the mechanical and biomedical properties of the composite scaffolds using the Taguchi method. The resulting composites display a range of advantages, including good mechanical strength, porous morphology, partial crystallinity, high swelling ratio, controlled biodegradability rate, and rheological characteristics. Additionally, we performed the cytotoxicity tests to confirm the improvement of the structure and better cell attachments on the scaffolds. Our findings illustrate that the presence of the ECM in chitosan/agarose structure improves the biomedical characteristics of the final scaffold. In addition, we were able to control the mechanical properties and microstructure of the scaffolds by optimizing the polymers' concentration and their resulting interactions. These results present a novel scaffold with simultaneously enhanced mechanical and cellular attributes comparing to the scaffolds without ECM for nasal cartilage tissue engineering applications.

Exosomes are endogenous nanoparticles with a lipid bilayer membrane whose natural function as carriers of biological materials has attracted much attention. The ability of exosomes to cross biological barriers, especially the blood-brain barrier, has highlighted them as tools of drug delivery to brain tumors. In a previous study, we isolated and characterized exosomes derived from human endometrial mesenchymal stem cells (hEnMSCs exosomes). In the present study, we used hEnMSCs exosomes as carriers for atorvastatin and investigated its pro-apoptotic and anti-angiogenic effects on U87 glioblastoma spheroids 3D co-cultured with Human Umbilical Vein Endothelial cells (HUVECs). In the study of HUVEC proliferation by using MTT assay, cell treatments with concentrations of 5 and 10 μM of free atorvastatin and atorvastatin-loaded hEnMSCs exosomes (AtoEXOs) showed significant differences in inhibition of proliferation compared to other concentrations. Also, 5 and 10 μM of AtoEXOs inhibited HUVEC migration in both scratch closure and transwell migration assays significantly more than that of free atorvastatin. In addition, in vitro HUVEC capillary tube network formation was inhibited by 5 and 10 μM treatment of AtoEXOs significantly more that of free atorvastatin. Moreover, a significant decrease in VEGF secretion and a significant increase in Bax/Bcl2 expression ratio were observed in U87 spheroids 3D co-cultured with HUVECs, especially for 10 μM AtoEXOs compared to other treated cell groups. Our results showed that hEnMSCs exosomes loaded with atorvastatin not only mimicked the anti-tumor effects of free atorvastatin but also potentiated its anti-tumor effects on glioblastoma cells. The enhanced pro-apoptotic and anti-angiogenic capabilities of atorvastatin loaded in hEnMSCs exosomes offer promising new perspectives for the treatment of glioblastoma.

Cell-laden filament-like hydrogels are advantageous for many applications including drug screening, tissue engineering, and regenerative medicine. However, most of the designed filament vehicles hold weak mechanical properties, which hinder their applications in specific tissue engineering. We present a binary hybrid silk and hyaluronic acid hydrogel microfiber generated through a microfluidic system to encapsulate cells with superior mechanical properties and biocompatibility. Cell-laden hydrogel microfibers were continuously produced through coaxial double orifice microfluidic device and horseradish peroxidase mediated crosslinking, which conjugated introduce phenolic moieties in the backbone of silk fibroin and HA derivatives (Silk-Ph and HA-Ph, respectively). The iterative hybrid Silk-Ph + HA-Ph fibers were fabricated in tunable size distribution between 195 and 680 μm through control of outer flow velocity. Tensile strength and maximum stain of prepared Silk-Ph + HA-Ph sample upregulated more than three times higher than the single HA-Ph sample, which demonstrated significant impacts of synthesized silk derivative in hydrogel fiber composition. The proteolytic degradation of microfibers manipulated by hyaluronidase and collagenase treatment. Encapsulation process and crosslinking did not insert any harmful effect on cell viability (90%) and the cells maintained their growth ability after encapsulation process. Cellular filament-like tissue fabricated from proliferation of cells in Silk-Ph + HA-Ph microfiber.

Despite advances in the treatment of acute myocardial infarction, due to the non-proliferative nature of adult cardiomyocytes, the injured myocardium is mainly replaced by fibrotic tissue, which ultimately causes heart failure. To prevent heart failure, particularly after myocardial infarction, exosome-based therapy has emerged as one of the most promising strategies to regenerate cardiac function. Exosomes can carry microRNAs in support of neovascularization, anti-inflammatory, and endogenous cardiac regeneration. This study demonstrated that animal rat models' combination treatment with microRNA-126 and microRNA-146a mimics in exosomes is desirable for cardiac regeneration after myocardial infarction. The exosomes isolated from stem cells and loaded with microRNAs were characterized their impacts in cell migration, tube formation, and vascular endothelial growth factor degree. In the following, the usefulness of loaded microRNAs in exosomes and their encapsulation within alginate derivative hydrogel was analyzed in myocardial infarction for an animal model. Exosomes isolated and loaded with microRNAs showed the synergetic impact on cell migration, tube formation, and promoted vascular endothelial growth factor folding. Moreover, microRNAs loaded exosomes and encapsulated them in alginate hydrogel could help in reducing infarct size and improving angiogenesis in myocardial infarction. The angiogenesis markers including CD31 and connexion 43 upregulated for myocardial infarction models treated with alginate-based hydrogels loaded with exosomes and microRNAs-exosomes. Histological analysis indicated that myocardial infarction model rats treated with alginate hydrogel loaded with microRNAs-exosomes possessed lower and higher degrees of fibrosis and collagen fiber, respectively. These findings have important therapeutic implications for a myocardial infarction model through angiogenesis and vascular integrity regulation.

BackgroundSilk fibroin (SF) as a natural polymer holds great potential in biomedical research because of its biocompatibility, easy processing, high toughness, and strength. However, slow gelation time has narrowed its applications, specifically in cell-laden microparticles that are versatile structures for tissue engineering due to their unique features. In addition, most crosslinking methods used to decrease gelation time did not occur in a mid-condition. Methods This study aimed to use modified SF with phenol conjugation to accelerate crosslinking mediated via horseradish peroxidase (HRP)/ hydrogen peroxide (H2O2) in a co-flow high-throughput microfluidic device for the ultimate goal of cell-laden silk fibroin-phenol (SF-Ph) microparticles formation. The physical and biochemical properties of fabricated cell-laden SF-Ph were evaluated to reveal its potential for tissue engineering.ResultsThe monodisperse microparticles in shape and size were formed in various diameters changing from 300 to 80 µm by altering oil phase velocity from SF-Ph substrate. More than 90% cell viability and three times cells upregulation of mitochondrial activity of enclosed-cells in microparticles with 150 ± 32 µm diameters revealed that these structures were suitable subcultures produced through a mild process based on morphological and MTT assays. It was noticed that cells approximately cover the microparticles until the 15th day. ConclusionSpherical micro-tissue formation in microparticles, resulting from cell growth promoted by cell-cell and cell-matrix interactions, adds significant weight to this method's applications.

Self-assembled structures mostly arises through enzyme-regulated phenomena in nature under persistent conditions. Enzymatic reactions are one of main biological processes in fabrication and construction of supramolecular hydrogel networks required for biomedical applications. The enzymatic processes provide a unique opportunity to integrate hydrogel formation. In most of cases, structure and substrates of hydrogels are adjusted by enzyme catalysis due to the chemo-, regio- and stereo-selectivity of enzymes. Hydrogels processed by using various enzyme schemes showed remarkable characteristics as dynamic frames for cells, bioactive molecules and drugs in biomedical applications. A novel class of enzyme-mediated crosslinking hydrogels mimics the extracellular matrices by displaying unique physicochemical properties and functionalities like water-retention capacity, drug loading ability, biodegradability, biocompatibility, biostability, bioactivity, optoelectronic properties, self-healing ability, shape memory ability. In recent years, many enzymatic systems investigated hydrogel cross-linking. Results of biocompatible hydrogel products show that these mechanisms of crosslinking can fulfill requirements for variety of biomedical applications including tissue engineering, wound healing and drug delivery.

The main aim of this study was to evaluate the efficacy of cerium oxide nanoparticles (CNPs) encapsulated in fabricated hybrid silk-fibroin (SF)/polycaprolactone (PCL) nanofibers as an artificial neural guidance conduit (NGC) applicable for peripheral nerve regeneration. The NGC was prepared by PCL and SF filled with CNPs. The mechanical properties, contact angle, and cell biocompatibility experiments showed that the optimized concentration of CNPs inside SF and SF/PCL wall of conduits was 1 (wt/wt). The SEM image analysis showed the nanoscale texture of the scaffold in different topologies depend on composition with fiber diameters at about 351 ± 54 nm and 420 ± 73 nm respectively for CNPs + SF and CNPs + SF/PCL fibrous mats. Furthermore, contact angle measurement confirmed the hydrophilic behavior of the membranes, ascribable to the SF content and surface modification through modified methanol treatment. The balance of morphological and biochemical properties of hybrid CNPs 1 (wt/wt) + SF/PCL construct improves cell adhesion and proliferation in comparison with lower concentrations of CNPs in nanofibrous scaffolds. The release of CNPs 1 (wt/wt) from both CNPs + SF and CNPs+ SF/PCL fibrous mats was highly controlled and very slow during the extended time of incubation until 60 days. Fabricated double-layered NGC using CNPs + SF and CNPs + SF/PCL fibers was consistent for application in nervous tissue engineering and regenerative medicine from a structural and biocompatible perspective. © 2021 Wiley Periodicals LLC

Cellular growth of enclosed cells in core-shell microcapsules is a key element for the practical use of the device in tissue engineering and biopharmaceutical fields. We developed alginate derivative microcapsules with a liquid core template by horseradish peroxidase crosslinking using an integrated coaxial microfluidic device by electrospray system. The cells and gelatin solution were extruded from the inner channel of coaxial microfluidic device and alginate possessing phenolic moieties (Alg-Ph) and horseradish peroxidase (HRP) flowed from the outer channel. In open electric filed, concentric drops of the two coaxial fluids broken up into microdrops and sprayed into the gelling bath containing hydrogen peroxide to instantly gel alginate in the shell fluid before the two fluids got mixed or gelatin dispersed in a gelling bath. The core-shell structure of about 350 μm in diameter and gel membrane of 42 μm was developed by optimization of operational parameters including electrical voltage, flow rate and concentration of polymers. The physical properties of microcapsules including swelling and mechanical resistance proved the applicability of fabricated vehicles for cell culture systems in vitro and in vivo. The viability of enclosed fibroblast cells in generated core-shell microcapsule was more than 90% which is sufficiently high compared with it before encapsulation. The growth profile and behavior of cells in microcapsules showed appropriate cell growth and the possibility of fabrication of spherical tissue was confirmed through degradation of hydrogel membrane. These results validate the significant potential of coaxial electrospray system and HRP-mediated hydrogelation in the fabrication of cell-laden core-shell microcapsule for tissue engineering and regenerative medicine.

Hybrid fibrous mat containing cell interactive molecules offers the ability to deliver the cells and drugs in wound bed, which will help to achieve a high therapeutic treatment. In this study, a co-electrospun hybrid of polyvinyl alcohol (PVA), chitosan (Ch) and silk fibrous mat was developed and their wound healing potential by localizing bone marrow mesenchymal stem cells (MSCs)-derived keratinocytes on it was evaluated in vitro and in vivo. It was expected that fabricated hybrid construct could promote wound healing due to its structure, physical, biological specifications. The fabricated fibrous mats were characterized for their structural, mechanical and biochemical properties. The shape uniformity and pore size of fibers showed smooth and homogenous structures of them. Fourier transform infrared spectroscopy (FTIR) verified all typical absorption characteristics of Ch-PVA + Silk polymers as well as Ch-PVA or pure PVA substrates. The contact angle and wettability measurement of fibers showed that mats found moderate hydrophilicity by addition of Ch and silk substrates compared with PVA alone. The mechanical features of Ch-PVA + Silk fibrous mat increase significantly through co-electrospun process as well as hybridization of these synthetic and natural polymers. Higher degrees of cellular attachment and proliferation obtained on Ch-PVA + Silk fibers compared with PVA and Ch-PVA fibers. In terms of the capability of Ch-PVA + Silk fibers and MSC-derived keratinocytes, histological analysis and skin regeneration results showed this novel fibrous construct could be suggested as a skin substitute in the repair of injured skin and regenerative medicine applications.

Exosomes (EXOs) are naturally occurring nanosized lipid bilayers that can be efficiently used as a drug delivery system to carry small pharmaceutical, biological molecules and pass major biological barriers such as the blood-brain barrier. It was hypothesized that EXOs derived from human endometrial stem cells (hEnSCs-EXOs) can be utilized as a drug carrier to enhance tumor-targeting drugs, especially for those have low solubility and limited oral bioactivity. In this study, atorvastatin (Ato) loaded EXOs (AtoEXOs) was prepared and characterized for its physical and biological activities in tumor growth suppression of 3 D glioblastoma model. The AtoEXOs were obtained in different methods to maximize drug encapsulation efficacy. The characterization of AtoEXOs was performed for its size, stability, drug release, and in vitro anti-tumor efficacy evaluated comprising inhibition of proliferation, apoptosis induction of tumor cells. Expression of apoptotic genes by Real time PCR, Annexin V/PI, tunnel assay was studied after 72 h exposing U87 cells where encapsulated in matrigel in different concentrations of AtoEXOs (5, 10 μM). The results showed that the prepared AtoEXOs possessed diameter ranging from 30–150 nm, satisfying stability and sustainable Ato release rate. The AtoEXOs was up taken by U87 and generated significant apoptotic effects while this inhibited tumor growth of U87 cells. Altogether, produced AtoEXOs formulation due to its therapeutic efficacy has the potential to be an adaptable approach to treat glioblastoma brain tumors., Highlights • Isolated exosome for endometrial stem cells show sizes between 30 and 150 nm in spherical-shaped, and atorvastatin could be efficiently loaded into exosome. • Atorvastatin loaded exosome (AtoEXO) were stable during 30 days' incubation and released relatively slowly from exosome. • U87 cells untaken AtoEXO in short time efficiently. • AtoEXO had anti-proliferative impact on U87 cells which proved by attenuated cell density and viable cell as well as decreasing expression of Ki-67 protein. • Efficient activity of AtoEXO to control tumor cell growth proved through up regulation of pro-apoptotic and apoptosis pathways including caspase-3, 8 and 9.

Functional improvement after spinal cord injury remains an unsolved difficulty. Glial scars, a major component of SCI lesions, are very effective in improving the rate of this recovery. Such scars are a result of complex interaction mechanisms involving three major cells, namely, astrocytes, oligodendrocytes, and microglia. In recent years, scientists have identified two subtypes of reactive astrocytes, namely, A1 astrocytes that induce the rapid death of neurons and oligodendrocytes, and A2 astrocytes that promote neuronal survival. Moreover, recent studies have suggested that the macrophage polarization state is more of a continuum between M1 and M2 macrophages. M1 macrophages that encourage the inflammation process kill their surrounding cells and inhibit cellular proliferation. In contrast, M2 macrophages promote cell proliferation, tissue growth, and regeneration. Furthermore, the ability of oligodendrocyte precursor cells to differentiate into adult oligodendrocytes or even neurons has been reviewed. Here, we first scrutinize recent findings on glial cell subtypes and their beneficial or detrimental effects after spinal cord injury. Second, we discuss how we may be able to help the functional recovery process after injury., Highlights • Reactive astrocytes are divided into two subsets: A1 astrocytes and A2 astrocytes. • The transformation of A1 to A2 reactive astrocytes can be a beneficial strategy for better repair. • The NeuroD1 factor can be used to convert most astrocytes into the neurons or the A2 astrocytes. • Differentiation of oligodendrocytes to their progenitor cells after injury may be useful for better response. • Increase the ratio of M2/M1 macrophages at the injury site is a potential therapeutic solution.

One of the most effective approaches for treatment of chronic rhinosinusitis is the use of hydrogel scaffolds with the sustained release of a given required drug. With this in mind, first, we synthesized and characterized poly (lactide-co-glycolide) (PLGA) micro and nano particles loaded with dexamethasone (DEX). We observed a 7-day release of DEX from nanoparticles, while the microparticles showed a 22-day release profile. Due to their slower rate of release, the PLGA microparticles loaded with DEX (PLGADEX microparticles) were specifically chosen for this study. As a second step, chitosan/polyvinylpyrrolidone (PVP) based hydrogels were prepared in various weight ratios and the PLGADEX microparticles were optimized in their structure based on variable gelation times. The morphological studies showed PLGADEX microparticles homogenously dispersed in the hydrogels. Moreover, the effect of weight ratio in the presence and absence of optimum percentage of PLGADEX microparticles was studied. The resultant hydrogels demonstrated a range of advantages, including good mechanical strength, porous morphology, amorphous structure, high swelling ratio, controlled biodegradability rate, and antibacterial activity. Additionally, a cytotoxicity analysis confirmed that the hydrogel scaffolds do not have adverse effects on the cells; our release studies in the hydrogel with the highest PVP content also showed 80 release after 30 days. Based on these results we were able to predict and control some of the mechanical properties, including the microstructure of the scaffolds, as well as the drug release, by optimizing the polymers - microparticle concentration, plus their resulting interactions. This optimized hydrogel can become part of a suitable alternative for treatment of allergic rhinitis and chronic sinusitis. © 2020 Elsevier B.V.

Hydrogels catalyzed by horseradish peroxidase (HRP) serve as an efficient and effective platform for biomedical applications due to their mild reaction conditions for cells, fast and adjustable gelation rate in physiological conditions, and an abundance of substrates as water-soluble biocompatible polymers. In this review, we highlight the tunable characteristics and use of the HRP-catalyzed hydrogels and provide a brief overview of various substrates employed in the HRP system for different biomedical applications of the resultant hydrogels. In addition, we discuss and summarize the biocompatibility, possible functionalization, and biofabrication process. Finally, the future prospective of the HRP crosslinking system is highlighted with biomedical applications.

The hydrogels having the ability to promote migration and morphogenesis of endothelial cells (ECs) are useful for fabricating vascularized dense tissues in vitro. The present study explores the immobilization of low molecular weight hyaluronic acid (LMWHA) derivative within gelatin-based hydrogel to stimulate migration of ECs. The LMWHA derivative possessing phenolic hydroxyl moieties (LMWHA-Ph) was bound to gelatin-based derivative hydrogel through the horseradish peroxidase-catalyzed reaction. The motility of ECs was analyzed by scratch migration assay and microparticle-based cell migration assay. The incorporated LMWHA-Ph molecules within hydrogel was found to be preserved stably through covalent bonds during incubation. The free and immobilized LMWHA-Ph did not lose an inherent stimulatory effect on human umbilical vein endothelial cells (HUVECs). The immobilized LMWHA-Ph within gelatin-based hydrogel induced the high motility of HUVECs, accompanied by robust cytoskeleton extension, and cell subpopulation expressing CD44 cell receptor. In the presence of immobilized LMWHA-Ph, the migration distance and the number of existing HUVECs were demonstrated to be encouraged in dose-dependent and time-dependent manners. Based on the results obtained in this work, it was concluded that the enzymatic immobilization of LMWHA-Ph within gelatin-based hydrogel represents a promising approach to promote ECs' motility and further exploitation for vascular tissue engineering applications.

The hydrogel efficacy of taurine-loaded chitosan nanoparticle/alginate hydrogel was investigated for controlled release of the taurine substrate, which is known as an antioxidative drug. The composition of the fabricated hydrogels was explored by Fourier-transform infrared spectroscopy. The swelling ability and degradation rate of hydrogels were also analyzed in phosphate-buffered saline at the physiological condition and alginate lyase for a period of 21 days. Moreover, morphologies and structure of hydrogels and cells were determined using a scanning electron microscope. The possible cytotoxicity of the fabricated hydrogel was carried out by seeding endometrial stem cells on hydrogels. The results demonstrated that hydrogel of chitosan nanoparticle/alginate hydrogel successfully controlled the release of taurine. We observed that the chitosan nanoparticle/alginate hydrogel possessed adjust swelling ability and degradation rate as compared to neat alginate hydrogel. The results proved that the chitosan nanoparticle/alginate hydrogel is non-cytotoxic and could be utilized as a promising composition in tissue engineering and drug delivery systems.

Hybrid electrospun fiber containing bioactive molecules, which offer the ability to deliver the cells into the wound bed, will help to achieve a high therapeutic effect. In this study, an electrospun polycaperlactone (PCL) and gelatin (Gela) scaffold containing curcumin loaded chitosan nanoparticle (NCs/Cur) was used to evaluate in vivo wound healing ability of the fabricated scaffolds. The electrospun hybrid scaffold seeded with human endometrial stem cells (EnSCs) showed desirable biocompatibility with the host immune system and wound healing ability in a full-thickness excisional animal model. The constructs were characterized for structural, mechanical and biochemical properties. Fourier transform infrared spectroscopy (FTIR) confirmed all typical absorption characteristics of PCL and Gela polymers as well as NCs and Cur. The results showed the perfect contact angle, wettability and degradability of hybrid fiber scaffolds with the good mechanical and structural characteristics including shape uniformity, pore size and porosity. The cell attachment and proliferation on the PCL/Gela/NCs/Cur was higher than PCL and PCL/Gela scaffolds. In term of the capability of hybrid scaffold and EnSCs in histological analysis, this novel tissue-engineered construct could be suggested as a skin substitute to repair injured skin and regenerative medicine application.

Hydrogel microcapsules having the ability to promote cell adhesion and proliferation are a useful tool for fabricating tissue in vitro. The present study explored the effects of two anionic polysaccharide hydrogel membranes which have an impact on the adhesiveness, morphology and growth of cells. Microcapsules were made by coating a cell-laden gelatin microparticle with a hydrogel membrane produced from modified hyaluronic acid or alginate possessing phenolic hydroxyl moieties (HA-Ph or Alg-Ph respectively) via a horseradish peroxidase-catalyzed crosslinking reaction. Some gelatin was retained within the microcapsules to support the attachment and growth of encapsulated cells. The morphological and functional characteristics of encapsulated HeLa and 10T1/2 cells were evaluated. The HA-Ph hydrogel, which exhibited greater retention of gelatin, showed a higher degree of cytocompatibility with respect to cell adhesion, spreading and proliferation compared with the Alg-Ph hydrogel membrane. These findings indicate that HA-Ph microcapsules synthesized around a temporary gelatin microparticles are a promising cell vehicle for tissue engineering applications.

The influence of contents of galactose and phenolic hydroxyl (Ph) groups incorporated into chitosan was investigated on characteristics of the chitosan derivatives and the resultant gels as well as HepG2 cell attachment and growth behaviors. Introduction of galactose groups increased the solubility of the chitosan derivatives. The gelation time decreased with increasing content of Ph groups in the chitosan derivatives. The increase of galactose groups incorporated at a fixed content of Ph groups improved mechanical properties of the resultant gels. In vitro degradation rate of the resultant gels decreased by increasing Ph groups and decreasing galactose groups incorporated into the chitosan derivatives. The HepG2 cells formed dense spheroid cell clusters when the galactose groups were absent or incorporated at high level into chitosan (13.8mol%). However, the cells exhibited spreading morphology with spheroid formation on the gels containing 1.1 and 5.2mol% galactose groups. The albumin secretion level on a cellular basis also increased considerably when the galactose groups increased to 13.8mol%. The results demonstrated the potential of the chitosan derivative hydrogels for liver tissue engineering applications.

Central nervous system trauma can cause loss of functional sensory neurons and motor functions that results in death of neuronal cells and axon degeneration. For this purpose, therapeutic approaches to enhance repair mechanisms spinal cord injury (SCI) is reflected to be a difficult task that requires thoughtful strategy. Hydrogel scaffolds in the presence of stem cells could repair damaged SCI due to their ability to support cellular viability, motility, and differentiation. In this study, we investigated the effectiveness of a biomimetic composite hydrogel of chitosan and gelatin containing endometrial stem cells (EnSCs) as a minimally invasive treatment of SCI in an animal model. Moreover, the simultaneous, effect of intraperitoneal injection of atorvastatin drug was assessed for animal models. From the results, morphological and cell viability studies demonstrated favorable seeding microenvironment and viability in the hydrogel. The transplanted hydrogel containing EnSCs into hemisected SCI rats showed recovery of sensory and motor functions by confirming locomotor activity evaluation using Basso, Beattie, and Bresnahan test. The growth of neuronal cells confirmed with immunohistochemical study. The findings in this study suggested that combination therapy using biomimetic hydrogel seeded with stem cells as well as atorvastatin injection has the potential to heal SCI through neuroinflammation attenuation, improvement of functional recovery, and limit the secondary damages.

The efficacy of Alginate/ Polyvinyl alcohol/ Hydroxyapatite nanopowders (Alg/PVA/HANPs) mats was investigated for increasing periodontal ligament stem cells (PDLSCs) differentiation into the osteoblast cells. The Alg/PVA/HANPs nanofibers were fabricated through electospinning process and following ionic crosslinking of Ca+2 and glutaraldehyde crosslinkers. The physical features of mats and expression of osteoblast cell markers were evaluated. Results showed the diameter of nanofiber increases with decreasing the incorporation of HANPs in Alg/PVA nanofiber. According to MTT assay the viability of cells on scaffolds are better in the all of groups, so the scaffolds were not toxic for the cultured cells. The osteogenic culture media appropriately enhanced osteopontin and osteocalcin markers. Furthermore, in vitro study indicated that the Alg/PVA/HANPs nanofiber provided a suitable three-dimensional structure for osteoblast cells differentiation. Thus, the Alg/PVA/HANPs scaffold can be recommended for in bone repair.

Hydrogel fibers with biodegradable and biocompatible features are useful for the fabrication of filament-like tissues. We developed cell-laden hyaluronic acid (HA)-based hollow hydrogel fibers to create single and bundled filament-like tissues. The cell-laden fibers were fabricated by crosslinking phenolic-substituted hyaluronic acid (HA-Ph) in an aqueous solution containing cells through a horseradish peroxidase (HRP)-catalyzed reaction in the presence of catalase by extruding the solution in ambient flow of an aqueous solution containing H2O2. The encapsulated cells proliferated and grew within the hollow core, and the cells formed filament-like constructs in both single and bundled fibers, which were obtained by collection on a rotating cylindrical tube. Single and bundled filament-like tissues covered with an additional heterogeneous cell layer were obtained by degrading the fiber membrane using hyaluronidase after covering the fiber surface with heterogeneous cells. Cellular viability was preserved during HA-Ph hydrogel fiber fabrication and filament-like tissue formation. These results demonstrate the feasibility of HA-based hollow hydrogel fibers obtained through HRP- and catalase-mediated reactions to engineer filament-like tissues.

The optimization of hyaluronic acid (HA) extraction from eggshell was studied by using a sequential experimentation approach. Using the results of factorial experiments, optimization experiments were carried out by response surface methodology. The experimental data as well as the validation tests showed the optimum performance when the extraction pH and time were bounded in the ranges of 3.4–3.5 and 3–3.4 days, respectively, at 9 °C. The purified HA was characterized by FTIR spectrum, harmonic mean size and average molecular weight. The study depicted the potential of eggshell as an alternative source for HA extraction to use in industry and medicine.

Hydrogel fibers are structurally and biologically useful devices for differentiation of stem cells and fabrication of filament-like tissues. We established cell-laden degradable hydrogel fibers through visible light-initiated crosslinking to differentiate stem cells and fabricate filament-like tissue. Human adipose stem cell (hADSC)-laden fibers were fabricated by cross-linking phenolic-substituted alginate and gelatin (Alg-Ph and Gela-Ph respectively) in an aqueous solution containing cells. The crosslinking of phenolic moieties was mediated by ruthenium(II) tris-bipyridyl dication (Ru(II) bpy and sodium ammonium persulfate (SPS) and irradiating visible light. The hydrogel microfiber fabricated with desirable geometries and dimensions. The encapsulated hADSCs proliferated and grew within hydrogel microfiber, maintained their multipotency ability and formed filament-like constructs. The filament-like tissues covered with an additional heterogeneous cell layer was made by degrading the fiber membrane using alginate-lyase after covering the fiber surface with vascular endothelial cells. Cellular viability is preserved during Alg-Ph and Gela-Ph hydrogel fiber fabrication and filament-like tissue formation. These results demonstrate the feasibility of Alg-based hydrogel fibers obtained through the Ru/SPS-mediated crosslinking system and visible light irradiation for the engineering of filament-like tissues and cell-based therapeutic treatments.

This article describes the preparation of cell-enclosing hyaluronic acid (HA) microparticles with solid core and microcapsules with liquid core through cell-friendly horseradish peroxidase (HRP)-catalyzed hydrogelation. The spherical vehicles were made from HA derivative possessing phenolic hydroxyl moieties (HA-Ph) cross-linkable through the enzymatic reaction by extruding cell-suspending HA-Ph aqueous solution containing HRP from a needle of 180 μm in inner diameter into the ambient coaxial flow of liquid paraffin containing H2O2 in a microtubule of 600 μm in diameter. By altering the flow rate of liquid paraffin, the diameters of gelatin and HA-Ph microparticles were varied in the range of 120–220 μm and 100–300 μm, respectively. The viability of the enclosed human hepatoma HepG2 cells in the HA-Ph microparticles of 180 μm in diameter was 94.2 ± 2.3%. The growth of the enclosed HepG2 cells was enhanced by decreasing the HRP concentration. The microcapsules of 200 μm in diameter were obtained by extruding HA-Ph aqueous solution containing thermally liquefiable cell-enclosing gelatin microparticles of 150 μm in diameter using the same microfluidic system. The enclosed cells grew and filled the cavity within 10 days. Spherical tissues covered with a heterogeneous cell layer were obtained by degrading the microcapsule membrane using hyaluronidase after covering the surface with a heterogeneous cell layer. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016, 133, 43107.