Skin Defects In Mice Research Articles

IPSC-derived megakaryocytes and platelets accelerate wound healing and angiogenesis

BackgroundPlatelet-rich plasma (PRP), which is prepared by concentrating platelets in autologous blood, shows efficacy in chronic skin wounds via multiple growth factors. However, it exhibits heterogeneity across patients, leading to unstable therapeutic efficacy. Human induced pluripotent stem cell (iPSC)-derived megakaryocytes and platelets (iMPs) are capable of providing a stable supply, holding promise as materials for novel platelet concentrate-based therapies. In this context, we evaluated the effect of iMPs on wound healing and validated lyophilization for clinical applications.MethodsThe growth factors released by activated iMPs were measured. The effect of the administration of iMPs on human fibroblasts and human umbilical vein endothelial cells (HUVECs) was investigated in vitro. iMPs were applied to dorsal skin defects of diabetic mice to assess the wound closure rate and quantify collagen deposition and angiogenesis. Following the storage of freeze-dried iMPs (FD-iMPs) for three months, the stability of growth factors and their efficacy in animal models were determined.ResultMultiple growth factors that promote wound healing were detected in activated iMPs. iMPs specifically released FGF2 and exhibited a superior enhancement of HUVEC proliferation compared to PRP. Moreover, an RNA-seq analysis revealed that iMPs induce polarization to stalk cells and enhance ANGPTL4 gene expression in HUVECs. Animal studies demonstrated that iMPs promoted wound closure and angiogenesis in chronic wounds caused by diabetes. We also confirmed the long-term stability of growth factors in FD-iMPs and their comparable effects to those of original iMPs in the animal model.ConclusionOur study demonstrates that iMPs promote angiogenesis and wound healing through the activation of vascular endothelial cells. iMPs exhibited more effectiveness than PRP, an effect attributed to the exclusive presence of specific factors including FGF2. Lyophilization enabled the long-term maintenance of the composition of the growth factors and efficacy of the iMPs, therefore contributing to stable supply for clinical application. These findings suggest that iMPs provide a novel treatment for chronic wounds.

Potential of gelatin hydrogel nonwoven fabrics (Genocel) as a skin substitute in a diabetic mouse skin defect model

BackgroundGelatin hydrogel nonwoven fabrics (Genocel) are three-dimensional gelatin scaffolds that provide cells with space for proliferation, migration, and differentiation. They are expected to be an effective wound healing modality to treat intractable wounds, such as diabetic foot ulcers, because they enhance early neovascularization when used as a skin substitute. In this study, we explored the healing process of Genocel applied to skin defects in diabetic mice and compared it with that of a conventional skin substitute, Pelnac. MethodsGenocel and Pelnac sheets were used to treat skin defects on the backs of diabetic mice. On days 7 and 14, the remaining wound area was evaluated and specimens were harvested for HE, Azan, anti-CD31, CD68, and CD163 staining to assess neoepithelialization, granulation tissue formation, capillary formation, and macrophage infiltration. ResultsWounds treated with Genocel showed a wound healing process comparable to that of wounds treated with Pelnac. No significant differences were observed in the remaining wound area, neoepithelial length, granulation formation, number of pan-macrophages, or M2 ratio on days 7 and 14. The only significant difference was the number of induced M2 macrophages, which was higher in Pelnac group than in the Genocel group on day 7 (p < 0.05). ConclusionsGenocel showed similar healing effects in diabetic wounds as Pelnac and is considered an effective wound management modality for diabetic ulcers.

Effects of applying human umbilical cord mesenchymal stem cell exosomes through different pathways to treat full-thickness skin defect wounds in mice

Objective: To investigate the effects of human umbilical cord mesenchymal stem cell (hUCMSC) exosomes in the treatment of full-thickness skin defect wounds in mice through local wound application, subcutaneous injection at the wound margin, and tail vein injection, and to explore the optimal administration route of hUCMSC exosomes for wound treatment. Methods: This study was an experimental study. hUCMSC exosomes were extracted from the discarded umbilical cord tissue of three normal delivery women aged 25-35 years in the Department of Obstetrics and Gynecology of Baogang Hospital of Inner Mongolia and successfully identified. Totally 120 male BALB/c mice aged 6-8 weeks were selected, and full-thickness skin defect wounds were prepared on the back of them. According to the random number table, the injured mice were divided into control group (without drug administration), local wound application group, wound margin subcutaneous injection group, and tail vein injection group (with 30 mice in each group). Mice in the latter three groups were given 0.2 mL phosphate buffer solution containing 200 μg hUCMSC exosomes by local wound application, subcutaneous injection at the wound margin, and tail vein injection, respectively. On post injury day (PID) 7, 14, and 21, the general condition of the wound was observed, and the wound healing rate was calculated; the wound tissue was collected, the pathological changes and collagen fibers were observed respectively by hematoxylin-eosin staining and Masson staining, the number of new microvessels was observed by CD31 immunohistochemical staining, and the content of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) was detected by enzyme-linked immunosorbent assay. The sample number was 10 in each group at each time point. Results: On PID 7, 14, and 21, the wounds of mice in the 4 groups all healed gradually, and the wound healing of the mice in wound margin subcutaneous injection group was the best; the wound healing rates of mice in the three administration groups were significantly higher than those in control group (P<0.05), the wound healing rates of mice in wound margin subcutaneous injection group and tail vein injection group were significantly higher than those in local wound application group (P<0.05), and the wound healing rates of mice in wound margin subcutaneous injection group were significantly higher than those in tail vein injection group (P<0.05). On PID 7, 14, and 21, the growth and epithelialization speed of the wound tissue of mice in the three administration groups were significantly accelerated, and the collagen fibers in the wounds of mice in the three administration groups were larger in number and more neatly arranged in comparison with the control group. On PID 7, 14, and 21, under every 200-fold visual field, the number of new microvessels in the wound tissue of mice in local wound application group was 24.1±2.5, 50.7±4.1, and 44.2±2.3, respectively, the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was 32.2±2.9, 67.5±4.9, and 53.6±3.7, respectively, and the number of new microvessels in the wound tissue of mice in tail vein injection group was 27.8±2.4, 59.1±3.7, and 49.6±2.6, respectively, which was significantly more than 20.6±1.7, 46.7±3.4, and 40.9±2.8 in control group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly more than that in local wound application group (P<0.05); the number of new microvessels in the wound tissue of mice in wound margin subcutaneous injection group was significantly more than that in tail vein injection group (P<0.05). On PID 7, 14, and 21, the content of TNF-α and IL-6 in the wound tissue of mice in the three administration groups was significantly less than that in control group (P<0.05), the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group and tail vein injection group was significantly less than that in local wound application group (P<0.05), and the content of TNF-α and IL-6 in the wound tissue of mice in wound margin subcutaneous injection group was significantly less than that in tail vein injection group (P<0.05). Conclusions: Local wound application, subcutaneous injection at the wound margin, and tail vein injection of hUCMSC exosomes can all promote the wound healing of full-thickness skin defects in mice through alleviating excessive inflammatory response and promoting angiogenesis. Among them, subcutaneous injection at the wound margin has a better therapeutic effect, indicating subcutaneous injection at the wound margin is the optimal administration route for hUCMSC exosomes in wound treatment.

Framework Nucleic Acids-Based VEGF Signaling Activating System for Angiogenesis: A Dual Stimulation Strategy.

Angiogenesis is crucial for tissue engineering, wound healing, and regenerative medicine. Nanomaterials constructed based on specific goals can be employed to activate endogenous growth factor-related signaling. In this study, based on the conventional single-stranded DNA self-assembly into tetrahedral framework nucleic acids (tFNAs), the Apt02 nucleic acid aptamer and dimethyloxallyl glycine (DMOG) small molecule are integrated into a complex via a template-based click chemistry reaction and toehold-mediated strand displacement reaction. Thus, being able to simulate the VEGF (vascular endothelial growth factor) function and stabilize HIF (hypoxia-inducible factor), a functional whole is constructed and applied to angiogenesis. Cellular studies demonstrate that the tFNAs-Apt02 complex (TAC) has a conspicuous affinity to human umbilical vein endothelial cells (HUVECs). Further incubation with DMOG yields the tFNAs-Apt02-DMOG complex (TACD), which promotes VEGF secretion, in vitro blood vessel formation, sprouting, and migration of HUVECs. Additionally, TACD enhances angiogenesis by upregulating the VEGF/VEGFR and HIF signaling pathways. Moreover, in a diabetic mouse skin defect repair process, TACD increases blood vessel formation and collagen deposition, therefore accelerating wound healing. The novel strategy simulating VEGF and stabilizing HIF promotes blood-vessel formation in vivo and in vitro and has the potential for broad applications in the vascularization field.

Modulation of macrophages by a phillyrin-loaded thermosensitive hydrogel promotes skin wound healing in mice

BackgroundImpaired wound healing in traumatic skin injuries remains a severe clinical challenge due to impaired re-vascularization, harmful bacteria infection, and inflammation dysregulation. Macrophages are recognized as prominent immune cells in tissue regeneration and wound healing. Consequently, the modulation of macrophages provides a promising therapeutic target for wound healing disorders. Here, we aimed to explore whether a novel constructed combination of thermosensitive hydrogel Pluronic F-127 (PF-127) and phillyrin (PH, the main active compound of forsythia suspensa) could improve skin wound healing. MethodsFirstly, the biological effects of pH on the phenotype and inflammation of macrophages were assessed by flow cytometry and ELISA. The biocompatibility of the PF-127 plus PH combination was investigated on keratinocytes and red blood cells. The biological effect of PF-127/PH hydrogel on the migratory ability of keratinocytes in vitro was evaluated using the scratch and transwell migration assays. In addition,S. aureusandE. coliwere employed to test the antibacterial properties of the PF-127 plus PH combination. Finally, PF-127 plus PH scaffold was appliedto the full-thickness skin defect in mice. Histomorphological evaluation and immunochemistry were performed to explore the wound-healing activity of PF-127/PH hydrogel. ResultsPH can promote the polarization of macrophages from the M1 (pro-inflammatory) phenotype to the M2 (anti-inflammatory) phenotype. The PF-127/PH hydrogel was highly biocompatible and showed a potent stimulative effect on the migration of keratinocytesin vitro. The combination of PF-127 and PH exerted a pronounced antibacterial activity onS. aureusandE. coli in vitro.PF-127/PH hydrogel potently accelerates the healing of full-thickness skin defects by promoting skin cell proliferation, accelerating angiogenesis, and inhibiting inflammation. ConclusionsOur study suggests that PF-127/PH hydrogel has excellent potential for treating traumatic skin defects.

Manipulation of Surface Electrical Charge on Nanocomposite Membranes Confers Wide Spectrum Bactericidal Effects and Promotes Tissue Regeneration

AbstractUtilization of electro‐responsive biomaterials with antibacterial properties is advantageous for facilitating septic wound healing and tissue regeneration. However, the dose‐response effects of electrical stimuli from these materials against bacteria are not rigorously characterized, and achieving synergy of bactericidal and pro‐regenerative effects of biomaterials remains a major challenge. Here, a graded series of flexible BaTiO3/P(VDF‐TrFE) electroactive nanocomposite membranes (EMs) are developed with varying surface charge intensities, to serve as antibacterial dressing for septic wound healing. EMs display broad‐spectrum antibacterial effects against both Gram‐positive and Gram‐negative bacteria in a dose‐dependent manner, depending on the magnitude of their surface electrical potential. Mechanistically, the surface charge of EMs increase intracellular levels of reactive oxygen species within bacteria cells, which in turn caused oxidative damage to the bacterial membrane, thereby suppressing bacterial activity and biofilm formation. Moreover, in vivo studies demonstrated that EMs effectively inhibited S. aureus infection and accelerated wound healing in a mouse skin defect model, as well as ameliorated P. gingivalis‐mediated periodontal inflammation in a mouse periodontitis model. Hence, this study optimizes the antibacterial properties of electroactive materials and characterizes the dose‐response effects of surface electrical charge against bacteria, thus validating the therapeutic applications of electroactive biomaterials in combating bacterial infection.

Development of a Self-Assembled Dermal Substitute from Human Fibroblasts Using Long-term Three-Dimensional Culture.

Skin substitutes have emerged as an alternative to autografts for the treatment of skin defects. Among them, scaffold-based dermal substitutes have been extensively studied; however, they have certain limitations, such as delayed vascularization, limited elasticity, and the inability to achieve permanent engraftment. Self-assembled, cell-based dermal substitutes are a promising alternative that may overcome these shortcomings but have not yet been developed. In this study, we successfully developed a cell-based dermal substitute (cultured dermis) through the long-term culture of human dermal fibroblasts using the net-mold method, which enables three-dimensional cell culture without the use of a scaffold. Spheroids prepared from human dermal fibroblasts were poured into a net-shaped mold and cultured for 2, 4, or 6 months. The dry weight, tensile strength, collagen and glycosaminoglycan levels, and cell proliferation capacity were assessed and compared among the 2-, 4-, and 6-month culture periods. We found that collagen and glycosaminoglycan levels decreased over time, while the dry weight remained unchanged. Tensile strength increased at 4 months, suggesting that remodeling had progressed. In addition, the cell proliferation capacity was maintained, even after a 6-month culture period. Unexpectedly, the internal part of the cultured dermis became fragile, resulting in the division of the cultured dermis into two collagen-rich tissues, each of which had a thickness of 400 μm and sufficient strength to be sutured during in vivo analysis. The divided 4-month cultured dermis was transplanted to skin defects of immunocompromised mice and its wound healing effects were compared to those of a clinically available collagen-based artificial dermis. The cultured dermis promoted epithelialization and angiogenesis more effectively than the collagen-based artificial dermis. Although further improvements are needed, such as the shortening of the culture period and increasing the size of the cultured dermis, we believe that the cultured dermis presented in this study has the potential to be an innovative material for permanent skin coverage.

Silk fibroin-gelatin photo-crosslinked 3D-bioprinted hydrogel with MOF-methylene blue nanoparticles for infected wound healing.

Photo-crosslinked hydrogel (PH) is an outstanding candidate for three-dimensional (3D) printing as a wound dressing because of its high efficiency in crosslinking and injectability. In this study, methylene blue (MB)-loaded UiO-66(Ce) nanoparticles (NPs) were synthesized to prevent drug self-aggregation and achieve the photodynamic therapy (PDT) effect for efficient antibacterial action. Then, a composite photocrosslinked silk fibroin (SF)/gelatin hydrogel loaded with MB@UiO-66(Ce) NPs (MB@UiO-66(Ce)/PH) was fabricated. The printability and the improvement of the mechanical properties of the hydrogel by the NPs were clarified. The hydrogel exhibited good biocompatibility and promoted the migration and proliferation of fibroblasts. With the PDT effect of MB@UiO-66(Ce) NPs, the hydrogel showed an excellent antibacterial effect, which became more pronounced as the concentration increased. In vivo study showed that the MB@UiO-66(Ce)/PH could fill the defects without gaps and accelerate the repair rate of full-thickness skin defects in mice. The MB@UiO-66(Ce)/PH with antibacterial properties and tissue healing-promoting ability provides a new strategy involving 3D bioprinting for preparing wound dressings.

Effects of tumor necrosis factor-alpha/extracellular signal-regulated kinase pathway on migration ability of HaCaT cells and full-thickness skin defects in mice

Effects of tumor necrosis factor-alpha/extracellular signal-regulated kinase pathway on migration ability of HaCaT cells and full-thickness skin defects in mice

Effects of interleukin-4-modified gold nanozymes on the full-thickness skin defects in diabetic mice

Effects of interleukin-4-modified gold nanozymes on the full-thickness skin defects in diabetic mice

Effects of exosomes from hepatocyte growth factor-modified human adipose mesenchymal stem cells on full-thickness skin defect in diabetic mice

Effects of exosomes from hepatocyte growth factor-modified human adipose mesenchymal stem cells on full-thickness skin defect in diabetic mice

Comparison of Wound Healing Effect of Skin Micrograft Impregnated into Two Kinds of Artificial Dermis in a Murine Wound Model.

Skin MG was prepared from the skin of C57BL/6JJcl mice using Rigeneracons and administered onto Pelnac Gplus and Integra sheets. The amount of MG suspension impregnated in Pelnac Gplus and Integra was evaluated. Pelnac Gplus and Integra sheets combined with MG were applied to murine defects, and wound area, neoepithelium length, granulation tissue formation, and newly formed capillaries were compared with the control groups on days 7 and 14. The weight percentage of the MG absorbed by Pelnac Gplus and Integra was 88.8% ± 3.5% and 28.2% ± 7.0%, respectively (P < 0.05). In the in vivo experiment, the area of newly formed granulation tissue and both the number and area of newly formed capillaries in the PelnacG + MG group were significantly larger than those in the control group at 14 days after implantation (P < 0.05). Skin MG was successfully impregnated into Pelnac Gplus by simple administration but not into Integra. Administration of skin MG into the Pelnac Gplus promoted granulation formation and angiogenesis. Pelnac Gplus was more suitable than Integra in the combination therapy.

Application of collagen-chondroitin sulfate scaffolds with different pore sizes combined with acidic fibroblast growth factor in repairing full thickness skin defects in nude mice

Wound healing of skin defects is complex. For the treatment of large and deep wounds, it is a good alternative to accept artificial dermis grafting at the first stage surgery, and autologous split-thickness skin grafting 2–3 weeks later at the second stage surgery. In addition, the effectiveness of numerous cytokines such as fibroblast growth factor (FGF) on wounds healing has been widely researched. The traditional view is that direct external application or in vivo injection of exogenous FGFs may not achieve the desired therapeutic effect as the effective concentration cannot be maintained for a long time. Therefore, some researchers have tried to integrate various cytokines into skin substitutes for combined application. However, we believe that considering the current situation, it is still difficult to achieve mass production of these types of artificial dermis. Here, we manufactured a collagen-chondroitin sulfate scaffold material by imitating the marketed artificial dermis materials. Then, we combined it with recombinant human acidic FGF in a single full dose during the first-stage artificial dermis transplantation, which is simple and completely feasible but always controversial in the current clinical work, to explore whether this combinatorial therapy could serve as an efficient way wound healing in the Balb/c-nu mice full-thickness skin defect model.

Injectable protocatechuic acid based composite hydrogel with hemostatic and antioxidant properties for skin regeneration

Acute skin injuries require timely and effective treatment. Injectable self-healing hydrogels are a very promising approach, but they face many challenges because of the complex microenvironments of skin wound. Although hydrogels have been extensively studied for multifunction, injectable self-healing hydrogels with both hemostatic and antioxidant properties are rare. In this paper, gelatin grafted protocatechuic acid (GT-PCA) and oxidized-dextran (Odex) were used to engineer GT-PCA/Odex composite hydrogels via Schiff base reaction. The in vitro experimental results show that the hydrogels not only are injectable, self-healing, hemostatic, antioxidant, but also have good degradability, adhesion and biocompatibility. Further, the hydrogels can promote wound healing in full-thickness mouse skin defect models by reducing inflammation and increasing angiogenesis, collagen deposition and granulation tissue formation. Notably, the preparation method of GT-PCA/Odex hydrogels is gentle and simple, the hydrogel system has bright prospects in the treatment of acute skin injury and is expected to be applied in clinical trials.

Effects of in situ cross-linked graphene oxide-containing gelatin methacrylate anhydride hydrogel on wound vascularization of full-thickness skin defect in mice

Effects of in situ cross-linked graphene oxide-containing gelatin methacrylate anhydride hydrogel on wound vascularization of full-thickness skin defect in mice

Role and mechanism of Vγ4 T cells in impaired wound healing of rapamycin-induced full-thickness skin defects in mice

Role and mechanism of Vγ4 T cells in impaired wound healing of rapamycin-induced full-thickness skin defects in mice

Effects of exosomes from human adipose-derived mesenchymal stem cells on inflammatory response of mouse RAW264.7 cells and wound healing of full-thickness skin defects in mice

Effects of exosomes from human adipose-derived mesenchymal stem cells on inflammatory response of mouse RAW264.7 cells and wound healing of full-thickness skin defects in mice

Injectable multifunctional CMC/HA-DA hydrogel for repairing skin injury.

Injectable Hydrogels with adhesive, antioxidant and hemostatic properties are highly desired for promoting skin injury repair. In this study, we prepared a multi-functional carboxymethyl chitosan/hyaluronic acid-dopamine (CMC/HA-DA) hydrogel, which can be crosslinked by horseradish peroxidase and hydrogen peroxide. The antioxidation, gelation time, degradability, rheology and antihemorrhagic properties of hydrogels can be finely tuned by varying composition ratio. The cytocompatibility test and hemolysis test confirmed that the designed hydrogel holds good biocompatibility. More importantly, the repair effect of the hydrogel on full-thickness skin injury model in mice was studied. The results of wound healing, collagen deposition, immunohistochemistry and immunofluorescence showed that CMC/HA-DA hydrogel could significantly promote angiogenesis and cell proliferation at the injured site. Notably, the inflammatory response can also be regulated to promote the repair of full-thickness skin defect in mice. Results indicate that this injectable CMC/HA-DA hydrogel holds high application prospect for promising wound healing.

Dried human cultured epidermis accelerates wound healing in diabetic mouse skin defect wounds

Cryopreserved allogeneic cultured epidermis (CE) is used for treating second-degree burn wounds and diabetic foot ulcers; however, the need for cryopreservation limits its use. We have previously reported that CE accelerates wound healing irrespective of its viability and hypothesized that dehydrated CEs lacking living cells may act as an effective wound dressing. We prepared dried CE and investigated its morphological and physical properties and wound-healing effects and compared them with those of cryopreserved CE. Hematoxylin–eosin staining, immunostaining for basement membrane, and electron microscopy revealed that the morphologies of dried CE and cryopreserved CE were comparable and that the membrane structure was not damaged. The breaking strength, modulus of elasticity, and water permeability of dried CE were comparable with those of the cryopreserved CE. Furthermore, the levels of various active cytokines and chemokines in dried CE were comparable with those in cryopreserved CE. Dried CE applied to skin defect in diabetic mice significantly reduced the wound area and increased the new epithelium length 4 and 7 days after implantation, similar to that observed for cryopreserved CE. Consequently, dried CE had similar morphological and physical properties and wound-healing effects compared with those of cryopreserved CE and can be a physiological and versatile wound-dressing.

Effects of porcine acellular dermal matrix combined with human epidermal stem cells on wound healing of full-thickness skin defect in nude mice

Effects of porcine acellular dermal matrix combined with human epidermal stem cells on wound healing of full-thickness skin defect in nude mice