Effects of human amniotic membrane grafts combined with marrow mesenchymal stem cells on healing of full-thickness skin defects in rabbits

We have investigated the wound-healing effects of mesenchymal stem cells (MSCs) in combination with human amniotic membrane (HAM) when grafted into full-thickness skin defects of rabbits. Five defects in each of four groups were respectively treated with HAM loaded with autologous MSCs (group A), HAM loaded with allologous MSCs (group B), HAM with injected autologous MSCs (group C), and HAM with injected allologous MSCs (group D). The size of the wounds was calculated for each group at 7, 12, and 15 days after grafting. The wounds were subsequently harvested at 25 days after grafting. Sections stained with hematoxylin and eosin were used to determine the quality of wound healing, as based on the characteristics and amount of granulated tissue in the epidermal and dermal layers. Groups A and B showed the most pronounced effect on wound closure, with statistically significant improvement in wound healing being seen on post-operative days 7, 12, and 15. Although a slight trend toward improved wound healing was seen in group A compared with group B, no statistically significant difference was found at any time point between the two groups. Histological examination of healed wounds from groups A and B showed a thin epidermis with mature differentiation and collagen bundle deposition plus recovered skin appendages in the dermal layer. In contrast, groups C and D showed thickened epidermis with immature epithelial cells and increased fibroblast proliferation with only partially recovered skin appendages in the dermal layer. Thus, the graft of HAM loaded with MSCs played an effective role during the healing of skin defects in rabbits, with no significant difference being observed in wound healing between autologous and allologous MSC transplantation. This study was supported by research funds from Dong-A University.     

Adult bone-marrow-derived mesenchymal stem cells contribute to wound healing of skin appendages

Adult bone-marrow-derived mesenchymal stem cells (MSCs) are well-established as having the capacity to differentiate into cells with mesodermal, ectodermal, and endodermal characteristics and can leave their niche to home toward and engraft within foreign tissues. To investigate whether adult MSCs contribute to the repair of skin appendages after injury, BrdU-labeled MSCs were co-cultured with heat-shocked confluent sweat gland cells (SGCs) in vitro and later intravenously injected into full-thickness skin wounds in rats. When adult MSCs were co-cultured with heat-shocked SGCs, a subset of adult MSCs differentiated into SGCs, the percentage of differentiation being enhanced by epidermal growth factor and the injured microenviroment, but weakened by PD98059. The ERK (extracellular signal-regulated kinase) pathway, especially pERK, was involved in the phenotype conversion of human MSCs into human SGC. Labeled MSCs were noted in hair follicles, sebaceous glands, blood vessels, and dermis in full-thickness wounds, and the incorporated cells in hair follicles and sebaceous glands were also positive for pan-cytokeratin. After wound healing, some labeled MSCs returned to the bone marrow, whereas other were retained in the dermis. We conclude that adult MSCs have the capacity to dock at specific sites, to contribute to wound healing of skin appendages, and to home toward marrow, and that engraftment of bone-marrow-derived cells is a functional event.This work was supported in part by the National Basic Science and Development Program (973 Program and 2005CB522603) and the National Natural Science Foundation of China (30230370 and 30500194).     

Fifteen years of preclinical and clinical experiences about biotherapy treatment of lesions induced by accidental irradiation and radiotherapy

High dose radiation exposures involving medical treatments or accidental irradiation may lead to extended damage to the irradiated tissue. Alleviation or even eradication of irradiation induced adverse events is therefore crucial. Because developments in cell therapy have brought some hope for the treatment of tissues damages induced by irradiation, the Institute for Radiation and Nuclear Safety contributed to establish the clinical guidelines for the management of accidentally irradiated victims and to provide the best supportive care to patients all over the world. In the past 15 years, we contributed to develop and test cell therapy for protection against radiation side effects in several animal models, and we proposed mechanisms to explain the benefit brought by this new therapeutic approach. We established the proof of concept that mesenchymal stem cells (MSCs) migrate to damaged tissues in the nonobese diabetic/severe combined immunodeficiency immunotolerant mice model and in non-human primate after radiation exposure. We showed that the intravenous injection of MSCs sustains hematopoiesis after total body irradiation, improves wound healing after radiodermatitis and protects gut function from irradiation damages. Thanks to a tight collaboration with clinicians from several French hospitals, we report successful treatments of therapeutic/accidental radiation damages in several victims with MSC infusions for hematopoiesis correction, radio-induced burns, gastrointestinal disorders and protection homeostatic functions of gut management after radio-therapy.     

Mesenchymal stem cells: Potential role in corneal wound repair and transplantation

Corneal diseases are a major cause of blindness in the world. Although great progress has been achieved in the treatment of corneal diseases, wound healing after severe corneal damage and immunosuppressive therapy after corneal transplantation remain prob-lematic. Mesenchymal stem cells(MSCs) derived from bone marrow or other adult tissues can differentiate into various types of mesenchymal lineages, such as osteocytes, adipocytes, and chondrocytes, both in vivo and in vitro. These cells can further differentiate into specific cell types under specific conditions. MSCs migrate to injury sites and promote wound healing by secreting anti-inflammatory and growth factors. In ad-dition, MSCs interact with innate and acquired immune cells and modulate the immune response through their powerful paracrine function. Over the last decade, MSCs have drawn considerable attention because of their beneficial properties and promising therapeutic prospective. Furthermore, MSCs have been applied to various studies related to wound healing, autoim-mune diseases, and organ transplantation. This review discusses the potential functions of MSCs in protecting corneal tissue and their possible mechanisms in corneal wound healing and corneal transplantation.     

Assay of radiation effects in mouse skin as expressed in wound healing

The effect of 150 kVp X irradiation on the healing of full depth surgical wounds in the lower dorsal skin of the mouse was assayed by measuring the wound strength of seven 2-mm-wide segments along each wound. The strength of unirradiated wounds increased with time in two phases: during the first 2 weeks it reached nearly half of the values recorded from unwounded skin, after which the rate of increase slowed for at least 2 weeks before beginning a second increase. By 150 days, the breaking strength of the wound was about 80% of that of unwounded skin. A single dose of 18 Gy prior to wounding reduced the strength of the wounds to about one-third to one-half that of an unirradiated wounds within the 3 months of follow-up. The effect of irradiation on wound strength did not change as the interval between exposure and wounding was increased to 2 months but decreased slightly when this interval was extended to 3 months. When the healing wound was irradiated within 5 days of surgery, the effect on healing was about the same as with preirradiation; if irradiation was delayed for 12 days after wounding the second phase of healing was only postponed and the wound strength ultimately approached the values recorded from unirradiated wounds. The wound strength of skin preirradiated by X rays and assayed 14 days after wounding showed a clear sigmoid dose response with a threshold between 8 and 10 Gy and a plateau at the maximum effect above 20 Gy. The persistence for at least 3 months of the effect of radiation on wound healing suggests that the tissues involved in the healing process are normally proliferating slowly. The accelerated expression of radiation injury through surgical wounding permits the early quantification of the radiation response of tissues that would normally be delayed in their expression of radiation damage.     

Mesenchymal stromal cells in wound healing applications: role of the secretome,targeted delivery and impact on recessive dystrophic epidermolysis bullosa treatment

Mesenchymal stromal cells (MSCs) are multi-potent stromal-derived cells capable of self-renewal that possess several advantageous properties for wound healing, making them of interest to the field of dermatology. Research has focused on characterizing the unique properties of MSCs, which broadly revolve around their regenerative and more recently discovered immunomodulatory capacities. Because of ease of harvesting and expansion, differentiation potential and low immunogenicity, MSCs have been leading candidates for tissue engineering and regenerative medicine applications for wound healing, yet results from clinical studies have been variable, and promising pre-clinical work has been difficult to reproduce. Therefore, the specific mechanisms of how MSCs influence the local microenvironment in distinct wound etiologies warrant further research. Of specific interest in MSC-mediated healing is harnessing the secretome, which is composed of components known to positively influence wound healing. Molecules released by the MSC secretome can promote re-epithelialization and angiogenesis while inhibiting fibrosis and microbial invasion. This review focuses on the therapeutic interest in MSCs with regard to wound healing applications, including burns and diabetic ulcers, with specific attention to the genetic skin disease recessive dystrophic epidermolysis bullosa. This review also compares various delivery methods to support skin regeneration in the hopes of combating the poor engraftment of MSCs after delivery, which is one of the major pitfalls in clinical studies utilizing MSCs.     

Dopamine regulates mobilization of mesenchymal stem cells during wound angiogenesis

Angiogenesis is an important step in the complex biological and molecular events leading to successful healing of dermal wounds. Among the different cellular effectors of wound angiogenesis, the role of mesenchymal stem cells (MSCs) is of current interest due to their transdifferentiation and proangiogenic potentials. Skin is richly innervated by sympathetic nerves which secrete dopamine (DA) and we have recently shown that concentration of DA present in synaptic cleft can significantly inhibit wound tissue neovascularization. As recent reports indicate that MSCs by mobilizing into wound bed play an important role in promoting wound angiogenesis, we therefore investigated the effect of DA on the migration of MSCs in wound tissues. DA acted through its D(2) receptors present in the MSCs to inhibit their mobilization to the wound beds by suppressing Akt phosphorylation and actin polymerization. In contrast, this inhibitory effect of DA was reversed after treatment with specific DA D(2) receptor antagonist. Increased mobilization of MSCs was demonstrated in the wound site following blockade of DA D(2) receptor mediated actions, and this in turn was associated with significantly more angiogenesis in wound tissues. This study is of translational value and indicates use of DA D(2) receptor antagonists to stimulate mobilization of these stem cells for faster regeneration of damaged tissues.     

Mesenchymal stem cells and skin wound repair and regeneration: possibilities and questions

Adult mesenchymal stem cells (MSCs) have the capacity for self-renewal and for differentiating into a variety of cells and tissues. They may leave their niche to migrate to remote tissues and play a critical role in wound repair and tissue regeneration. Because of their multipotency, easy isolation and culture, highly expansive potential, and immunosuppression properties, these cells may be an attractive therapeutic tool for regenerative medicine and tissue engineering. Several studies have indicated a contribution of MSCs to reconstituting skin in cutaneous wounds, but problems still need resolution before MSCs can be widely used clinically. This review focuses mainly on the benefits of MSCs in skin wound healing and tissue regeneration and on the questions that remain to be answered before MSCs can be used in clinical practice. This study was supported in part by the National Natural Science Foundation of China (30730090, 30672176, 30500194) and by State Key Development Program of Basic Research of China (973 Program, 2005CB522603).     

The Effect of Mesenchymal Stem Cells and Chitosan Gel on Full Thickness Skin Wound Healing in Albino Rats: Histological,Immunohistochemical and Fluorescent Study

Background

Wound healing involves the integration of complex biological processes. Several studies examined numerous approaches to enhance wound healing and to minimize its related morbidity. Both chitosan and mesenchymal stem cells (MSCs) were used in treating skin wounds. The aim of the current work was to compare MSCs versus chitosan in wound healing, evaluate the most efficient route of administration of MSCs, either intradermal or systemic injection, and elicit the mechanisms inducing epidermal and dermal cell regeneration using histological, immunohistochemical and fluorescent techniques.

Material and Methods

Forty adult male Sprague Dawley albino rats were divided into four equal groups (ten rats in each group): control group (Group I); full thickness surgical skin wound model, Group II: Wound and chitosan gel. Group III: Wound treated with systemic injection of MSCs and Group IV: Wound treated with intradermal injection of MSCs. The healing ulcer was examined on day 3, 5, 10 and 15 for gross morphological evaluation and on day 10 and 15 for histological, immunohistochemical and fluorescent studies.

Results

Chitosan was proved to promote wound healing more than the control group but none of their wound reached complete closure. Better and faster healing of wounds in MSCs treated groups were manifested more than the control or chitosan treated groups. It was found that the intradermal route of administration of stem cells enhanced the rate of healing of skin wounds better than the systemic administration to the extent that, by the end of the fifteenth day of the experiment, the wounds were completely healed in all rats of this group. Histologically, the wound areas of group IV were hardly demarcated from the adjacent normal skin and showed complete regeneration of the epidermis, dermis, hypodermis and underlying muscle fibers. Collagen fibers were arranged in many directions, with significant increase in their area percent, surrounding fully regenerated hair follicles and sebaceous glands in the dermis of the healed areas more than in other groups.

Conclusion

MSCs enhanced the healing process of wound closure more than chitosan gel treatment. Furthermore, MSCs injected intradermally, were more efficient in accelerating wound healing than any other mode of treatment.     

Mesenchymal stromal cells from dermal and adipose tissues induce macrophage polarization to a pro-repair phenotype and improve skin wound healing

The process of wound healing restores skin homeostasis but not full functionality; thus, novel therapeutic strategies are needed to accelerate wound closure and improve the quality of healing. In this context, tissue engineering and cellular therapies are promising approaches. Although sharing essential characteristics, mesenchymal stromal cells (MSCs) isolated from different tissues might have distinct properties. Therefore, the aim of this study was to comparatively investigate, by a mouse model in vivo assay, the potential use of dermal-derived MSCs (DSCs) and adipose tissue–derived MSCs (ASCs) in improving skin wound healing. Human DSCs and ASCs were delivered to full-thickness mouse wounds by a collagen-based scaffold (Integra Matrix). We found that the association of both DSCs and ASCs with the Integra accelerated wound closure in mice compared with the biomaterial only (control). Both types of MSCs stimulated angiogenesis and extracellular matrix remodeling, leading to better quality scars. However, the DSCs showed smaller scar size,superior extracellular matrix deposition, and greater number of cutaneous appendages. Besides, DSCs and ASCs reduced inflammation by induction of macrophage polarization from a pro-inflammatory (M1) to a pro-repair (M2) phenotype. In conclusion, both DSCs and ASCs were able to accelerate the healing of mice skin wounds and promote repair with scars of better quality and more similar to healthy skin than the empty scaffold. DSCs associated with Integra induced superior overall results than the Integra alone, whereas scaffolds with ASCs showed an intermediate effect, often not significantly better than the empty biomaterial.     

MSC-exosome: A novel cell-free therapy for cutaneous regeneration

Cutaneous regeneration is a dynamic and complex process that requires a series of coordinated interactions involving epidermal cells, dermal cells, growth factors, the extracellular matrix (ECM), nerves and blood vessels at a damaged site. Mesenchymal stromal cells (MSCs) have been reported to participate in all afore-mentioned stages. Exosomes are one of the key secretory products of MSCs, resembling the effect of parental MSCs. They can shuttle various proteins, messenger RNA (mRNA) and microRNAs (miRNAs) to modulate the activity of recipient cells, and play important roles in cutaneous wound healing. Compared with MSCs, exosomes are more convenient to store and transport. Moreover, they avoid many risks associated with cell transplantation. Therefore, MSC-exosome–mediated therapy may be more safe and efficient. In this review, we summarize the latest studies and observations on the role of MSC-exosome in the acute and chronic wound model and provide a comprehensive understanding of the role of exosomes in wound healing. This review can assist investigators in exploring new therapeutic strategies for enhancing the efficacy of MSC-exosome for cutaneous repair and regeneration.     

Nacre-driven water-soluble factors promote wound healing of the deep burn porcine skin by recovering angiogenesis and fibroblast function

To assess the recovery effect of water-soluble components of nacre on wound healing of burns, water-soluble nacre (WSN) was obtained from powdered nacre. Alterations to WSN-mediated wound healing characteristics were examined in porcine skin with deep second-degree burns; porcine skin was used as a proxy for human. When WSN was applied to a burned area, the burn-induced granulation sites were rapidly filled with collagen, and the damaged dermis and epidermis were restored to the appearance of normal skin. WSN enhanced wound healing recovery properties for burn-induced apoptotic and necrotic cellular damage and spurred angiogenesis. Additionally, WSN-treated murine fibroblast NIH3T3 cells showed increased proliferation and collagen synthesis. Collectively, the findings indicate that WSN improves the process of wound healing in burns by expeditiously restoring angiogenesis and fibroblast activity. WSN may be useful as a therapeutic agent, with superior biocompatibility to powdered nacre, and evoking less discomfort when applied to a wounded area.     

A Modeling Approach to Study the Effect of Cell Polarization on Keratinocyte Migration

The skin forms an efficient barrier against the environment, and rapid cutaneous wound healing after injury is therefore essential. Healing of the uppermost layer of the skin, the epidermis, involves collective migration of keratinocytes, which requires coordinated polarization of the cells. To study this process, we developed a model that allows analysis of live-cell images of migrating keratinocytes in culture based on a small number of parameters, including the radius of the cells, their mass and their polarization. This computational approach allowed the analysis of cell migration at the front of the wound and a reliable identification and quantification of the impaired polarization and migration of keratinocytes from mice lacking fibroblast growth factors 1 and 2 – an established model of impaired healing. Therefore, our modeling approach is suitable for large-scale analysis of migration phenotypes of cells with specific genetic defects or upon treatment with different pharmacological agents.     

Notch与相关调控信号的相互作用在伤口愈合中的调控

皮肤是哺乳动物最重要的组织之一.当皮肤受损时,受损组织通过系列伤口愈合反应的生理和心理作用被修复,实现组织再生.再生反应主要发生在胚胎发育早期,伤口自愈能力随着机体的成熟而减弱;并且哺乳动物的组织重塑过程较为复杂,在不正确的信号引导下,可能引起并发症而导致创面愈合异常.研究表明,伤口微环境的稳态和信号分子的辅助作用是愈...     

Effects of genistein on early-stage cutaneous wound healing

Wound healing occurs in three sequential phases: hemostasis and inflammation, proliferation, and remodeling. Inflammation, the earliest phase, is considered a critical period for wound healing because immune cells remove damaged tissues, foreign debris, and remaining dead tissue. Wound healing would be delayed without inflammation, and this phase is affected by antioxidation capacity. Therefore, we hypothesized that genistein, which has an antioxidant effect, might modulate the wound healing process by altering the inflammatory response. After three days of acclimation, mice were divided into three groups: control, 0.025% genistein, and 0.1% genistein. After two weeks of an experimental diet, skin wounds were induced. Wounded skin areas were imaged, and the healing rate calculated. To measure lipid peroxidation, antioxidant enzyme expression and activity, and pro-inflammatory cytokine expression, skin and liver tissues were harvested at 12, 24, 48, and 72 h. Genistein did not affect body weight. The rate of wound closure in mice fed genistein was significantly faster than in the control group during the early stage of wound healing, especially in first three days. Cu, Zn-SOD and Mn-SOD expression in wound skin tissue in the 0.1% genistein group was lower than in the control group. However, CAT expression did not differ among groups. We also found that genistein modulated NF-κB and TNF-α expression during the early stage of wound healing. The genistein group had significantly lower hepatic lipid peroxidation and higher SOD, CAT, and GPx activities than the control group. These results suggest that genistein supplementation reduces oxidative stress by increasing antioxidant capacity and modulating proinflammatory cytokine expression during the early stage of wound healing.     

c-Met is essential for wound healing in the skin

Wound healing of the skin is a crucial regenerative process in adult mammals. We examined wound healing in conditional mutant mice, in which the c-Met gene that encodes the receptor of hepatocyte growth factor/scatter factor was mutated in the epidermis by cre recombinase. c-Met-deficient keratinocytes were unable to contribute to the reepithelialization of skin wounds. In conditional c-Met mutant mice, wound closure was slightly attenuated, but occurred exclusively by a few (5%) keratinocytes that had escaped recombination. This demonstrates that the wound process selected and amplified residual cells that express a functional c-Met receptor. We also cultured primary keratinocytes from the skin of conditional c-Met mutant mice and examined them in scratch wound assays. Again, closure of scratch wounds occurred by the few remaining c-Met-positive cells. Our data show that c-Met signaling not only controls cell growth and migration during embryogenesis but is also essential for the generation of the hyperproliferative epithelium in skin wounds, and thus for a fundamental regenerative process in the adult.     

Mesenchymal stem cell-derived exosomes: A novel and potential remedy for cutaneous wound healing and regeneration

Poor healing of cutaneous wounds is a common medical problem in the field of traumatology. Due to the intricate pathophysiological processes of wound healing, the use of conventional treatment methods, such as chemical molecule drugs and traditional dressings, have been unable to achieve satisfactory outcomes. Within recent years, explicit evidence suggests that mesenchymal stem cells (MSCs) have great therapeutic potentials on skin wound healing and regeneration. However, the direct application of MSCs still faces many challenges and difficulties. Intriguingly, exosomes as cell-secreted granular vesicles with a lipid bilayer membrane structure and containing specific components from the source cells may emerge to be excellent substitutes for MSCs. Exosomes derived from MSCs (MSC-exosomes) have been demonstrated to be beneficial for cutaneous wound healing and accelerate the process through a variety of mechanisms. These mechanisms include alleviating inflammation, promoting vascularization, and promoting proliferation and migration of epithelial cells and fibroblasts. Therefore, the application of MSC-exosomes may be a promising alternative to cell therapy in the treatment of cutaneous wounds and could promote wound healing through multiple mechanisms simultaneously. This review will provide an overview of the role and the mechanisms of MSC-derived exosomes in cutaneous wound healing, and elaborate the potentials and future perspectives of MSC-exosomes application in clinical practice.     

Mice lacking desmocollin 1 show epidermal fragility accompanied by barrier defects and abnormal differentiation.

The desmosomal cadherin desmocollin (Dsc)1 is expressed in upper epidermis where strong adhesion is required. To investigate its role in vivo, we have genetically engineered mice with a targeted disruption in the Dsc1 gene. Soon after birth, null mice exhibit flaky skin and a striking punctate epidermal barrier defect. The epidermis is fragile, and acantholysis in the granular layer generates localized lesions, compromising skin barrier function. Neutrophils accumulate in the lesions and further degrade the tissue, causing sloughing (flaking) of lesional epidermis, but rapid wound healing prevents the formation of overt lesions. Null epidermis is hyperproliferative and overexpresses keratins 6 and 16, indicating abnormal differentiation. From 6 wk, null mice develop ulcerating lesions resembling chronic dermatitis. We speculate that ulceration occurs after acantholysis in the fragile epidermis because environmental insults are more stringent and wound healing is less rapid than in neonatal mice. This dermatitis is accompanied by localized hair loss associated with formation of utriculi and dermal cysts, denoting hair follicle degeneration. Possible resemblance of the lesions to human blistering diseases is discussed. These results show that Dsc1 is required for strong adhesion and barrier maintenance in epidermis and contributes to epidermal differentiation.     

Matrix metalloproteinase 9 (MMP-9) is upregulated during scarless wound healing in athymic nude mice

Cutaneous wound healing is associated with migratory and remodeling events that require the action of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). Differences in their expressions were observed during scar-forming and scar-free skin wound healing. We previously found that athymic nude mice are exceptional among mature mammals in their ability to heal injured skin scarlessly. The present study was undertaken to determine whether the modulation of MMP-2 and MMP-9 expression during scarless healing in nude mice was different from scar-forming animals. Full thickness skin wounds were made into the back of nude, wild-type controls (C57BL/6J), immunodeficient SCID and Rag, thymectomized neonates and adults, and cyclosporin A treated mice. Post-injured skin tissues were harvested at Day 7 and 24 after injury. Quantitative RT-PCR, Western blot, gelatin zymography and immunohistochemical assays were performed. Our results show that MMP-2 protein was high but similarly expressed in all post-injured animals on Day 7 after injury. Late phase (Day 24) of wound repair was characterized by a decrease in mRNA and protein expression and a decrease in gelatinolytic activity of MMP-2 in all post-injured samples. On the contrary, high (p < 0.001) levels of mRNA expression, prominent pro-and active forms of MMP-9 and cells immunopositive for MMP-9 were present exclusively in the post-injured tissues from nude mice on Day 24 after wounding. This data suggest that MMP-9 expression in the remodeling phase of wound healing in nude mice could be a major component of their ability for scar-free healing.