Suppression of SOCS3 expression in macrophage cells: Potential application in diabetic wound healing.

Impaired polarization of M1 to M2 macrophages has been reported in diabetic wounds. We aimed to improve this polarization by down-regulation of expression of the "Suppressor of Cytokine Signaling 3" (SOCS3) gene in macrophages. Two oligodeoxynucleotide (ASO) sequences were designed against SOC3 mRNA and were loaded to mannosylated-polyethyleneimine (Man-PEI). The optimum N/P ratio for Man-PEI-ASO was determined to be 8 based on loading efficiency, particle size, zeta potential, cellular uptake and cytotoxicity assay. pH stability of ASO in Man-PEI-ASO and its protection from DNase I was confirmed. After in vitro treatment of macrophages with Man-PEI-ASO, SOCS3 was downregulated, SOCS1 upregulated, and SOCS1/SOCS3 ratio increased. Also, expressions of macrophage markers of M2 (IL-10, Arg1, CD206) increased and those of M1 (IL-1ß, NOS2, CD68) decreased, and secretion of pro-inflammatory cytokines (TNF-α and IL-1ß) decreased while that of anti-inflammatory cytokine IL-4 increased. All suggested a polarization into M2 phenotype. Finally, the Man-PEI-ASO was loaded in hydrogel and applied to a diabetic wound model in mice. It improved the healing to the level observed in non-diabetic wounds. We show that using antisense sequences against SOC3 mRNA, macrophage polarization could be directed into the M2 phenotype and healing of diabetic wound could be highly improved.

Decellularized Placental Sponge: A Platform for Coculture of Mesenchymal Stem Cells/Macrophages to Assess an M2 Phenotype and Osteogenic Differentiation In Vitro and In Vivo.

Effective communication between immune and bone-forming cells is crucial for the successful healing of bone defects. This study aimed to assess the potential of a decellularized placental sponge (DPS) as a coculture system for inducing M1/M2 polarization in macrophages and promoting osteogenic differentiation in adipose-derived mesenchymal stem cells (AD-MSCs), both in vitro and in vivo. We prepared the DPS and conducted a comprehensive characterization of its biomechanical properties, antibacterial activity, and biocompatibility. In vitro, we examined the influence of the DPS on the polarization of macrophages cocultured with AD-MSCs through nitric oxide assays, cytokine assays, phagocytosis tests, and real-time polymerase chain reaction (PCR). For in vivo assessment, we utilized micro-CT imaging, histological evaluations, and real-time PCR to determine the impact of the DPS seeded with Wharton's jelly mesenchymal stem cells (WJ-MSCs) on bone regeneration in a calvarial bone defect model. The coculture of AD-MSCs and macrophages on the DPS led to increased production of IL-10, upregulation of CD206, Arg1, and YM1 gene expression, and enhanced phagocytic capacity for apoptotic thymocytes. Concurrently, it reduced the secretion of TNF-α and nitric oxide (NO), downregulated the expression of CD86, NOS2, and IRF5 genes, and decreased macrophage phagocytosis of yeast. These results indicated polarization of macrophages toward an M2-like phenotype. In vivo, the presence of the DPS resulted in enhanced bone formation at the defect site. Immunostaining demonstrated that both the DPS and DPS + WJ-MSC constructs induced macrophage polarization toward an M2 phenotype, as compared to the control defect. In conclusion, this immunomodulatory effect, coupled with its biocompatibility and biomechanical properties resembling natural bone, positions the DPS as an attractive candidate for further exploration in the field of bone tissue engineering and regenerative medicine.

Fabrication and characterization of bilayer scaffolds made of decellularized dermis/nanofibrous collagen for healing of full-thickness wounds.

Skin tissue engineering has progressed from simple wound dressings to biocompatible materials with desired physico-chemical properties that can deliver regenerative biomolecules. This study describes using a novel biomimetic hybrid scaffold of decellularized dermis/collagen fibers that can continuously deliver stromal cell-derived factor-1 alpha (SDF-1α) for skin regeneration. In diabetic rat models, the idea that sustained SDF-1α infusion could increase the recruitment of CXCR4-positive cells at the injury site and improve wound regeneration was investigated. The morphology of the scaffold, its biocompatibility, and the kinetics of SDF-1 release were all assessed. SDF-1α was successfully incorporated into collagen nanofibers, resulting in a 200-h continuous release profile. The microscopic observations exhibited that cells are attached and proliferated on proposed scaffolds. As evaluated by in vivo study and histological examination, fabricated scaffold with SDF-1α release capacity exhibited a remarkably more robust ability to accelerate wound regeneration than the control group. Besides, the SDF-1α-loaded scaffold demonstrated functional effects on the proliferation and recruitment of CD31 and CXCR4-positive cells in the wound bed. Additionally, no adverse effects such as hyperplasia or scarring were found during the treatment period. It may be concluded that the fabricated hybrid scaffold based on natural polymer opens up a new option for topical administration of bioactive molecules. We believe the SDF-1α-loaded hybrid scaffold has promise for skin tissue engineering.

Regenerative medicine improve neurodegenerative diseases.

Regenerative medicine is a subdivision of medicine that improves methods to regrow, repair or replace unhealthy cells and tissues to return to normal function. Cell therapy, gene therapy, nanomedicine as choices used to cure neurodegenerative disease. Recently, studies related to the treatment of neurodegenerative disorders have been focused on stem cell therapy and Nano-drugs beyond other than regenerative medicine. Hence, by data from experimental models and clinical trials, we review the impact of stem cell therapy, gene therapy, and nanomedicine on the treatment of Alzheimer's disease (AD), Parkinson's disease (PD), and Amyotrophic lateral sclerosis (ALS). Indeed, improved knowledge and continued research on gene therapy and nanomedicine in treating Alzheimer's disease, Parkinson's disease, and Amyotrophic lateral sclerosis lead to advancements in effective and practical treatments for neurodegenerative diseases.

Circulatory long noncoding RNAs (circulatory-LNC-RNAs) as novel biomarkers and therapeutic targets in cardiovascular diseases: Implications for cardiovascular diseases complications.

Cardiovascular diseases (CVDs) are a group of disorders with major global health consequences. The prevalence of CVDs continues to grow due to population-aging and lifestyle modifications. Non-coding RNAs (ncRNAs) as key regulators of cell signaling pathways have gained attention in the occurrence and development of CVDs. Exosomal-lncRNAs (exos-lncRNAs) are emerging biomarkers due to their high sensitivity and specificity, stability, accuracy and accessibility in the biological fluids. Recently, circulatory and exos-based-lncRNAs are emerging and novel bio-tools in various pathogenic conditions. It is worth mentioning that dysregulation of these molecules has been found in different types of CVDs. In this regard, we aimed to discuss the knowledge gaps and suggest research priorities regarding circulatory and exos-lncRNAs as novel bio-tools and therapeutic targets for CVDs.

Coculture of adipose-derived mesenchymal stem cells/macrophages on decellularized placental sponge promotes differentiation into the osteogenic lineage.

BACKGROUND: Several factors like three-dimensional microstructure, growth factors, cytokines, cell-cell communication, and coculture with functional cells can affect the stem cells behavior and differentiation. The purpose of this study was to investigate the potential of decellularized placental sponge as adipose-derived mesenchymal stem cells (AD-MSCs) and macrophage coculture systems, and guiding the osteogenic differentiation of stem cells. METHODS: The decellularized placental sponge (DPS) was fabricated, and its mechanical characteristics were evaluated using degradation assay, swelling rate, and pore size determination. Its structure was also investigated using hematoxylin and eosin staining and scanning electron microscopy. Mouse peritoneal macrophages and AD-MSCs were isolated and characterized. The differentiation potential of AD-MSCs co-cultured with macrophages was evaluated by RT-qPCR of osteogenic genes on the surface of DPS. The in vivo biocompatibility of DPS was determined by subcutaneous implantation of scaffold and histological evaluations of the implanted site. RESULTS: The DPS had 67% porosity with an average pore size of 238 µm. The in vitro degradation assay showed around 25% weight loss during 30 days in PBS. The swelling rate was around 50% during 72 h. The coculture of AD-MSCs/macrophages on the DPS showed a significant upregulation of four differentiation osteogenic lineage genes in AD-MSCs on days 14 and 21 and a significantly higher mineralization rate than the groups without DPS. Subcutaneous implantation of DPS showed in vivo biocompatibility of scaffold during 28 days follow-up. CONCLUSIONS: Our findings suggest the decellularized placental sponge as an excellent bone substitute providing a naturally derived matrix substrate with biostructure close to the natural bone that guided differentiation of stem cells toward bone cells and a promising coculture substrate for crosstalk of macrophage and mesenchymal stem cells in vitro.

Additive manufacturing of bioactive glass biomaterials.

Tissue engineering (TE) and regenerative medicine have held great promises for the repair and regeneration of damaged tissues and organs. Additive manufacturing has recently appeared as a versatile technology in TE strategies that enables the production of objects through layered printing. By applying 3D printing and bioprinting, it is now possible to make tissue-engineered constructs according to desired thickness, shape, and size that resemble the native structure of lost tissues. Up to now, several organic and inorganic materials were used as raw materials for 3D printing; bioactive glasses (BGs) are among the most hopeful substances regarding their excellent properties (e.g., bioactivity and biocompatibility). In addition, the reported studies have confirmed that BG-reinforced constructs can improve osteogenic, angiogenic, and antibacterial activities. This review aims to provide an up-to-date report on the development of BG-containing raw biomaterials that are currently being employed for the fabrication of 3D printed scaffolds used in tissue regeneration applications with a focus on their advantages and remaining challenges.

Stem cell-mediated angiogenesis in skin tissue engineering and wound healing.

The timely management of skin wounds has been an unmet clinical need for centuries. While there have been several attempts to accelerate wound healing and reduce the cost of hospitalisation and the healthcare burden, there remains a lack of efficient and effective wound healing approaches. In this regard, stem cell-based therapies have garnered an outstanding position for the treatment of both acute and chronic skin wounds. Stem cells of different origins (e.g., embryo-derived stem cells) have been utilised for managing cutaneous lesions; specifically, mesenchymal stem cells (MSCs) isolated from foetal (umbilical cord) and adult (bone marrow) tissues paved the way to more satisfactory outcomes. Since angiogenesis plays a critical role in all four stages of normal wound healing, recent therapeutic approaches have focused on utilising stem cells for inducing neovascularisation. In fact, stem cells can promote angiogenesis via either differentiation into endothelial lineages or secreting pro-angiogenic exosomes. Furthermore, particular conditions (e.g., hypoxic environments) can be applied in order to boost the pro-angiogenic capability of stem cells before transplantation. For tissue engineering and regenerative medicine applications, stem cells can be combined with specific types of pro-angiogenic biocompatible materials (e.g., bioactive glasses) to enhance the neovascularisation process and subsequently accelerate wound healing. As such, this review article summarises such efforts emphasising the bright future that is conceivable when using pro-angiogenic stem cells for treating acute and chronic skin wounds.

Transplantation of decellularised human amniotic membranes seeded with mesenchymal stem cell-educated macrophages into animal models.

The reconstruction of chronic skin wounds remains a public health challenge in dermatology. Precisely controlling and monitoring the wound-healing process should result in enhanced outcomes for the patient. Cell-based therapies have shown great potential in medicine due to their immunomodulatory and healing properties. Herein, we produced activated macrophages by treating circulating monocytes with mesenchymal stem cell (MSC) supernatant. We also demonstrated the critical role of activated macrophages transplantation using amniotic membranes in accelerating wound healing in an animal wound model. The activated macrophages not only exhibited immunomodulatory cytokines like transforming growth factorß (TGFß) and interleukin 10 (and IL10) secretion but also showed attachment and proliferation ability on the amniotic membrane scaffold. Moreover, MSCs supernatant-treated cells also displayed significant ARG1, CD206, and IL 10 genes expression. Inspired by the in vitro results, we examined the in vivo therapeutic efficacy of the activated macrophage transplantation using an acellular amniotic membrane carrier in a full-thickness cutaneous wound model. The wound healing rate was significant in the group treated with macrophages generated via mesenchymal cell therapy seeded human amniotic membrane. There was less scarring in the wound sites after placing cell-scaffold constructs in the wound sites in the animal models. Overall, macrophages stimulated with mesenchymal cells' supernatant exhibited improved healing processes in incisional wounds by decreasing the inflammatory phase, increasing angiogenesis, and reducing scar tissue development.

Animal Kernicterus Models: Progress and Challenges.

Kernicterus is a leading cause of neonatal death throughout the world, especially in low-middle-income countries. It is developed by an unconjugated hyperbilirubinemia in the blood and brain tissue, triggering pathological processes that spawn neurotoxicity and neurodegeneration. However, the biological mechanism (s) of bilirubin-induced neurotoxicity and Kernicterus development remain to be well elucidated. Likewise, a practical therapeutic approach for human Kernicterus has yet to be found. Undoubtedly, animal models of Kernicterus can be helpful in the identification of underlying biological processes of hyperbilirubinemia evolution to Kernicterus, as well as the evaluation of various treatments efficacy in preclinical studies. More importantly, establishing an animal model that can mimic the Kernicterus and its behavioral, neuro-histological, and hematological manifestations is a severe priority in preclinical studies. So far, several Kernicterus animal models have been established that could partially mimic one or more clinical and paraclinical signs of human Kernicterus. The present study aimed to review all methods modeling Kernicterus with a focus on their potentials and shortcomings and subsequently provide the optimal methods for an ideal Kernicterus animal model.

CRISPR-Associated (CAS) Effectors Delivery via Microfluidic Cell-Deformation Chip.

Identifying new and even more precise technologies for modifying and manipulating selectively specific genes has provided a powerful tool for characterizing gene functions in basic research and potential therapeutics for genome regulation. The rapid development of nuclease-based techniques such as CRISPR/Cas systems has revolutionized new genome engineering and medicine possibilities. Additionally, the appropriate delivery procedures regarding CRISPR/Cas systems are critical, and a large number of previous reviews have focused on the CRISPR/Cas9-12 and 13 delivery methods. Still, despite all efforts, the in vivo delivery of the CAS gene systems remains challenging. The transfection of CRISPR components can often be inefficient when applying conventional delivery tools including viral elements and chemical vectors because of the restricted packaging size and incompetency of some cell types. Therefore, physical methods such as microfluidic systems are more applicable for in vitro delivery. This review focuses on the recent advancements of microfluidic systems to deliver CRISPR/Cas systems in clinical and therapy investigations.

Thermo-responsive chitosan hydrogel for healing of full-thickness wounds infected with XDR bacteria isolated from burn patients: In vitro and in vivo animal model.

Treatment of non-healing skin wounds infected with extensively drug-resistant (XDR) bacteria remains as a big challenge. To date, different biomaterials have been applied for treatment of post-wound infections, nevertheless their efficacy for treatment of the wounds infected with XDR isolates has not been determined yet. In this study, the potential of the thermo-responsive chitosan (TCTS) hydrogel for protection of full-thickness wounds XDR bacteria isolated from burn patients was evaluated both in vitro and in vivo in a rat model. Antibacterial activity of the TCTS hydrogel against standard strain and clinical isolates of Acinetobacter baumannii, cytobiocompatibility for Hu02 fibroblast cells, degradation rate and swelling ratio were determined in vitro. MTT assay and disk diffusion test indicated no detectable cytotoxicity and antibacterial activity in vitro, respectively. In vivo study showed significant acceleration of wound healing, re-epithelialization, wound closure, and decreased colony count in the TCTS hydrogel group compared with control. This study suggests TCTS hydrogel as an excellent wound dressing for management of the wounds infected with XDR bacteria, and now promises to proceed with clinical investigations.

Tenocyte-imprinted substrate: a topography-based inducer for tenogenic differentiation in adipose tissue-derived mesenchymal stem cells.

Tendon tissue engineering based on stem cell differentiation has attracted a great deal of attention in recent years. Previous studies have examined the effect of cell-imprinted polydimethylsiloxane (PDMS) substrate on induction differentiation in stem cells. In this study, we used tenocyte morphology as a positive mold to create a tenocyte-imprinted substrate on PDMS. The morphology and topography of this tenocyte replica on PDMS was evaluated with scanning electron microscopy (SEM) and atomic force microscopy. The tenogenic differentiation induction capacity of the tenocyte replica in adipose tissue-derived mesenchymal stem cells (ADSCs) was then investigated and compared with other groups, including tissue replica (which was produced similarly to the tenocyte replica and was evaluated by SEM), decellularized tendon, and bone morphogenic protein (BMP)-12, as other potential inducers. This comparison gives us an estimate of the ability of tenocyte-imprinted PDMS (called cell replica in the present study) to induce differentiation compared to other inducers. For this reason, ADSCs were divided into five groups, including control, cell replica, tissue replica, decellularized tendon and BMP-12. ADSCs were seeded on each group separately and investigated by the real-time reverse transcription polymerase chain reaction (RT-PCR) technique after seven and 14 days. Our results showed that in spite of the higher effect of the growth factor on tenogenic differentiation, the cell replica can also induce tenocyte marker expression (scleraxis and tenomodulin) in ADSCs. Moreover, the tenogenic differentiation induction capacity of the cell replica was greater than tissue replica. Immunocytochemistry analysis revealed that ADSCs seeding on the cell replica for 14 days led to scleraxis and tenomodulin expression at the protein level. In addition, immunohistochemistry indicated that contrary to the promising results in vitro, there was little difference between ADSCs cultured on tenocyte-imprinted PDMS and untreated ADSCs. The results of such studies could lead to the production of inexpensive cell culture plates or biomaterials that can induce differentiation in stem cells without growth factors or other supplements.

Oxygen-Releasing Scaffolds for Accelerated Bone Regeneration.

Hypoxia, the result of disrupted vasculature, can be categorized in the main limiting factors for fracture healing. A lack of oxygen can cause cell apoptosis, tissue necrosis, and late tissue healing. Remedying hypoxia by supplying additional oxygen will majorly accelerate bone healing. In this study, biphasic calcium phosphate (BCP) scaffolds were fabricated by robocasting, an additive manufacturing technique. Then, calcium peroxide (CPO) particles, as an oxygen-releasing agent, were coated on the BCP scaffolds. Segmental radial defects with the size of 15 mm were created in rabbits. Uncoated and CPO-coated BCP scaffolds were implanted in the defects. The empty (control) group received no implantation. Repairing of the bone was investigated via X-ray, histological analysis, and biomechanical tests at 3 and 6 months postoperatively, with immunohistochemical examinations at 6 months after operation. According to the radiological observations, formation of new bone was augmented at the interface between the implant and host bone and internal pores of CPO-coated BCP scaffolds compared to uncoated scaffolds. Histomorphometry analysis represented that the amount of newly formed bone in the CPO-coated scaffold was nearly two times higher than the uncoated one. Immunofluorescence staining revealed that osteogenic markers, osteonectin and octeocalcin, were overexpressed in the defects treated with the coated scaffolds at 6 months of postsurgery, demonstrating higher osteogenic differentiation and bone mineralization compared to the uncoated scaffold group. Furthermore, the coated scaffolds had superior biomechanical properties as in the case of 3 months after surgery, the maximal flexural force of the coated scaffolds reached to 134 N, while it was 92 N for uncoated scaffolds. The results could assure a boosted ability of bone repair for CPO-coated BCP scaffolds implanted in the segmental defect of rabbit radius because of oxygen-releasing coating, and this system of oxygen-generating coating/scaffold might be a potential for accelerated repairing of bone defects.

Additively manufactured small-diameter vascular grafts with improved tissue healing using a novel SNAP impregnation method.

The vascular network has a complex architecture such as branches, curvatures, and bifurcations which is even more complicated in view of individual patients' defect anatomy requiring custom-specifically designed vascular implants. In this work, 3D printing is used to overcome these challenges and a new shorter impregnation method was developed to incorporate S-nitroso-N-acetyl-d-penicillamine (SNAP) as a nitric oxide (NO) donor to printed grafts. The 3D-printed small-diameter vascular grafts (SDVGs) were impregnated with SNAP solution during SNAP synthesis (S1) or with SNAP dissolved in methanol (S2). The advantage of the newly developed S1 impregnation method is the elimination of the synthesis step by direct impregnation inside the S1 solution. Scanning electron microscopy imaging reveals the successful crystal formation in both methods. The results demonstrate that both S1- and S2-impregnated grafts, after covering with polycaprolactone topcoat, can release NO in a controlled manner and in the physiological range (0.5-4.0 × 10-10 mol cm-2 min-1 ) over a 15 days period. The created grafts with a NO-releasing surface have also shown bactericidal effect while the healing properties of the implant were improved by promoting migration and proliferation of endothelial cells (ECs). These results suggest that incorporation of 3D printing technology with the newly developed S1 impregnation of SNAP can optimize and shorten the manufacturing process of the next generation of patient-based antibacterial SDVGs with a higher attraction for ECs.

Decellularization and preservation of human skin: A platform for tissue engineering and reconstructive surgery.

A matrix derived from natural tissue functions as a highly biocompatible and versatile scaffold for tissue engineering applications. It can act as a supportive construct that provides a niche for colonization by host cells. In this work, we describe a cost-effective, reliable and reproducible protocol for decellularization and preservation of human skin as a potential soft tissue replacement. The decellularized human skin is achieved using purely chemical agents without any enzymatic steps. The suitability of the proposed method for the preservation of the extracellular matrix (ECM) structure and its main components and integrity were evaluated using histological and immunohistochemical analysis. Cryopreservation and final sterility were conducted using programmable freeze-drying and gamma irradiation. The architecture, basement membrane and 3D structure of ECM can be successfully preserved after decellularization. Our protocol was found to be appropriate to maintain key proteins such as collagen type I, III, IV and laminin in the structure of final scaffold. This protocol offers a novel platform for the preparation of a dermal substitute for potential clinical applications. STATEMENT OF SIGNIFICANCE: Clinical application of naturally-based scaffolds for verity of health problems obliges development of a reproducible and effective technology that does not change structural and compositional material properties during scaffold preparation and preservation. Lack of an effective protocol for the production of biological products using decellularization method is still remaining. This effort is directing to solve this challenge in order to accomplish the off-the -shelf availability of decellularized dermal scaffold in market for clinical application.

Nitric oxide-releasing vascular grafts: A therapeutic strategy to promote angiogenic activity and endothelium regeneration.

Small-diameter vascular grafts (SDVGs) are associated with a high incidence of failure due to infection and obstruction. Although several vascular grafts are commercially available, specific anatomical differences of defect sites require patient-based design and fabrication. Design and fabrication of such custom-tailored grafts are possible with 3d-printing technology. The aim of this study is to develop 3d-printed SDVGs with a nitric oxide (NO)-releasing coating to improve the success rate of implantation. The SDVGs were printed from polylactic acid and coated with blending of 10 wt% S-nitroso-N-acetyl-D-penicillamine into the polymeric substrate consisting of poly (ethylene glycol) and polycaprolactone. Our results show that NO is released in the physiological range (0.5-4 × 10-10 mol·cm-2·min-1) for 14 days and NO-releasing coating showed significant antibacterial potential against Gram-positive and Gram-negative strains. It was shown that both NO-releasing and control grafts are biocompatible in-vitro and in-vivo. Interestingly, the NO-releasing SDVGs dramatically enhanced ECs proliferation and significantly enhanced ECs migration in-vitro compared to control grafts. In addition, the NO-releasing SDVGs showed angiogenic potential in-vivo which can further prove the results of our in-vitro study. These findings are expected to facilitate tissue regeneration and integration of custom-made vascular implants with enhanced clinical success. STATEMENT OF SIGNIFICANCE: A series of 3d-printed small-diameter vascular grafts (SDVGs, <6 mm) with controlled release of nitric oxide (NO) were prepared to combine the advantages of 3D printing technology and NO-releasing systems. The resulting NO-releasing grafts were promisingly showing sustained NO release in the physiological range over a two weeks period. In addition to the evaluation of endothelial cell migration in-vitro, we implanted for the first time the NO-releasing vascular grafts in a chick chorioallantoic membrane (CAM) to investigate the effect of the prepared grafts on the angiogenesis in-vivo. The fabricated grafts also exhibited bactericidal properties which prevent the formation of a biofilm layer and can thereby enhance the chance of endothelialization on the surface. Taken together, the innovative combination of rapid and highly accurate 3d-printing technology as a patient-specific fabrication method with NO-releasing coating represents a promising approach to develop bactericidal SDVGs with improved endothelialization.

A Novel Intervention Technology for Cerebral Palsy: Brain Stimulation.

A common pediatric disorder with posture and motor dysfunction in neurological diseases is known as cerebral palsy (CP). Recently, a series of effective techniques have been developed for treatment of CP. These promising methods need high-tech equipment for brain stimulation and mainly classified into invasive and no-invasive approaches. This study aimed to introduce these techniques for treatment of patients who suffer from CP. The potential and performance of currently available brain stimulation techniques have been mentioned in detail. Moreover, the clinical application, safety, efficacy and challenges of these methods have been discussed. Here we review the recent advances in the CP treatment with an emphasis on brain stimulation techniques.

3D Protein-Based Bilayer Artificial Skin for the Guided Scarless Healing of Third-Degree Burn Wounds in Vivo.

Severe burn injuries can lead to delays in healing and devastating scar formation. Attempts have been made to develop a suitable skin substitute for the scarless healing of such skin wounds. Currently, there is no effective strategy for completely scarless healing after the thermal injuries. In our recent work, we fabricated and evaluated a 3D protein-based artificial skin made from decellularized human amniotic membrane (AM) and electrospun nanofibrous silk fibroin (ESF) in vitro. We also characterized both biophysical and cell culture investigation to establish in vitro performance of the developed bilayer scaffolds. In this report, we evaluate the appropriate utility of this fabricated bilayered artificial skin in vivo with particular emphasis on healing and scar formation due to the biochemical and biomechanical complexity of the skin. For this work, AM and AM/ESF membranes alone or seeded with adipose-tissue-derived mesenchymal stem cells (AT-MSCs) are implanted on full-thickness burn wounds in mice. The healing efficacy and scar formation are evaluated at 7, 14, and 28 days post-implantation in vivo. Our data reveal that ESF accelerates the wound-healing process through the early recruitment of inflammatory cells such as macrophages into the defective site as well as the up-regulation of angiogenic factors from the AT-MSCs and the facilitation of the remodeling phase. In vivo application of the prepared AM/ESF membrane seeded with the AT-MSCs reduces significantly the post-burn scars. The in vivo data suggest that the potential applications of the AM/ESF bilayered artificial skin may be considered a clinical translational product with stem cells to guide the scarless healing of severe burn injuries.