Concise Review: Bone Marrow-Derived Stem/Progenitor Cells in Cutaneous Repair and Regeneration

Our understanding of the role of bone marrow (BM)-derived cells in cutaneous homeostasis and wound healing had long been limited to the contribution of inflammatory cells. Recent studies, however, suggest that the BM contributes a significant proportion of noninflammatory cells to the skin, which are present primarily in the dermis in fibroblast-like morphology and in the epidermis in a keratinocyte phenotype; and the number of these BM-derived cells increases markedly after wounding. More recently, several studies indicate that mesenchymal stem cells derived from the BM could significantly impact wound healing in diabetic and nondiabetic animals, through cell differentiation and the release of paracrine factors, implying a profound therapeutic potential. This review discusses the most recent understanding of the contribution of BM-derived noninflammatory cells to cutaneous homeostasis and wound healing

Anti-inflammatory activity of nanocrystalline silver-derived solutions in porcine contact dermatitis

<p>Abstract</p> <p>Background</p> <p>Nanocrystalline silver dressings have anti-inflammatory activity, unlike solutions containing Ag⁺only, which may be due to dissolution of multiple silver species. These dressings can only be used to treat surfaces. Thus, silver-containing solutions with nanocrystalline silver properties could be valuable for treating hard-to-dress surfaces and inflammatory conditions of the lungs and bowels. This study tested nanocrystalline silver-derived solutions for anti-inflammatory activity.</p> <p>Methods</p> <p>Inflammation was induced on porcine backs using dinitrochlorobenzene. Negative and positive controls were treated with distilled water. Experimental groups were treated with solutions generated by dissolving nanocrystalline silver in distilled water adjusted to starting pHs of 4 (using CO₂), 5.6 (as is), 7, and 9 (using Ca(OH)₂). Solution samples were analyzed for total silver. Daily imaging, biopsying, erythema and oedema scoring, and treatments were performed for three days. Biopsies were processed for histology, immunohistochemistry (for IL-4, IL-8, IL-10, TNF-α, EGF, KGF, KGF-2, and apoptotic cells), and zymography (MMP-2 and -9). One-way ANOVAs with Tukey-Kramer post tests were used for statistical analyses.</p> <p>Results</p> <p>Animals treated with pH 7 and 9 solutions showed clear visual improvements. pH 9 solutions resulted in the most significant reductions in erythema and oedema scores. pH 4 and 7 solutions also reduced oedema scores. Histologically, all treatment groups demonstrated enhanced re-epithelialisation, with decreased inflammation. At 24 h, pMMP-2 expression was significantly lowered with pH 5.6 and 9 treatments, as was aMMP-2 expression with pH 9 treatments. In general, treatment with silver-containing solutions resulted in decreased TNF-α and IL-8 expression, with increased IL-4, EGF, KGF, and KGF-2 expression. At 24 h, apoptotic cells were detected mostly in the dermis with pH 4 and 9 treatments, nowhere with pH 5.6, and in both the epidermis and dermis with pH 7. Solution anti-inflammatory activity did not correlate with total silver content, as pH 4 solutions contained significantly more silver than all others.</p> <p>Conclusions</p> <p>Nanocrystalline silver-derived solutions appear to have anti-inflammatory/pro-healing activity, particularly with a starting pH of 9. Solutions generated differently may have varying concentrations of different silver species, only some of which are anti-inflammatory. Nanocrystalline silver-derived solutions show promise for a variety of anti-inflammatory treatment applications.</p

Paracrine Factors of Mesenchymal Stem Cells Recruit Macrophages and Endothelial Lineage Cells and Enhance Wound Healing

Bone marrow derived mesenchymal stem cells (BM-MSCs) have been shown to enhance wound healing; however, the mechanisms involved are barely understood. In this study, we examined paracrine factors released by BM-MSCs and their effects on the cells participating in wound healing compared to those released by dermal fibroblasts. Analyses of BM-MSCs with Real-Time PCR and of BM-MSC-conditioned medium by antibody-based protein array and ELISA indicated that BM-MSCs secreted distinctively different cytokines and chemokines, such as greater amounts of VEGF-α, IGF-1, EGF, keratinocyte growth factor, angiopoietin-1, stromal derived factor-1, macrophage inflammatory protein-1alpha and beta and erythropoietin, compared to dermal fibroblasts. These molecules are known to be important in normal wound healing. BM-MSC-conditioned medium significantly enhanced migration of macrophages, keratinocytes and endothelial cells and proliferation of keratinocytes and endothelial cells compared to fibroblast-conditioned medium. Moreover, in a mouse model of excisional wound healing, where concentrated BM-MSC-conditioned medium was applied, accelerated wound healing occurred compared to administration of pre-conditioned or fibroblast-conditioned medium. Analysis of cell suspensions derived from the wound by FACS showed that wounds treated with BM-MSC-conditioned medium had increased proportions of CD4/80-postive macrophages and Flk-1-, CD34- or c-kit-positive endothelial (progenitor) cells compared to wounds treated with pre-conditioned medium or fibroblast-conditioned medium. Consistent with the above findings, immunohistochemical analysis of wound sections showed that wounds treated with BM-MSC-conditioned medium had increased abundance of macrophages. Our results suggest that factors released by BM-MSCs recruit macrophages and endothelial lineage cells into the wound thus enhancing wound healing

Genetically Modified Dermal Keratinocytes Express High Levels of Transforming Growth Factor-β1

In an attempt to genetically modify cultured keratinocytes with transforming growth factor-β1 (TGF-β1), which has been proven to be one of the most important cytokines involved in wound healing, two constructs were made. One, designated pG3Z:K14-TGF-β1, is a plasmid in which the expression of TGF-β1 is driven by the keratin 14 promoter. The other, designated pLin-TGF-β1, is a retroviral vector in which the retroviral 5' long-terminal repeat promoter drives expression. In both constructs, the deletion of a small fragment of the noncoding region of the TGF-β1 gene was made to differentiate the transcript from that for endogenously expressed TGF-β1. Different types of cells were transfected with the pG3Z:K14-TGF-β1 construct using the calcium phosphate method. The pLin-TGF-β1 construct was propagated in a retroviral packaging cell line and conditioned medium that contained high titers of the virus was used to transduce keratinocytes or other types of cells grown in standard culture. Northern analysis, used to evaluate the expression of TGF-β1 mRNA in the pG3Z:K14-TGF-β1 transfected keratinocyte Cl-177 cell line, showed a smaller TGF-β1 transcript compared with that endogenously expressed by dermal fibroblasts. The level of TGF-β1 protein evaluated by enzyme-linked immunosorbent assay was significantly higher in medium conditioned by either the K14-TGF-β1 transfected or the pLin-TGF-β1 transduced keratinocytes, compared with that obtained from control cells; however, the level of TGF-β1 protein was unchanged in cultures of pG3Z:K14-TGF-β1 transfected nonkeratinocyte cells such as fetal and adult fibroblasts. Using the mink lung epithelial cell growth inhibition assay, we found an increase in TGF-β1 activity in conditioned medium from the pG3Z:K14-TGF-β1 transfected cells. To evaluate possible paracrine effects of the keratinocyte derived TGF-β1, a coculture system was established with pLin-TGF-β1 transduced keratinocytes grown in the upper chamber and dermal fibroblasts in the lower chamber. The results showed that TGF-β1 released from keratinocytes diffused to the lower chamber where it stimulated collagen production by dermal fibroblasts. In summary, we demonstrate here that primary cultured keratinocytes can be genetically modified to express high levels of TGF-β1 and suggest that this offers a potential approach for the therapy of dermal lesions such as nonhealing wounds

Analysis of Allogenicity of Mesenchymal Stem Cells in Engraftment and Wound Healing in Mice

Studies have shown that allogeneic (allo-) bone marrow derived mesenchymal stem cells (BM-MSCs) may enhance tissue repair/regeneration. However, recent studies suggest that immune rejection may occur to allo-MSCs leading to reduced engraftment. In this study, we compared allo-BM-MSCs with syngeneic BM-MSCs or allo-fibroblasts in engraftment and effect in wound healing. Equal numbers of GFP-expressing allo-BM-MSCs, syngeneic BM-MSCs or allo-fibroblasts were implanted into excisional wounds in GFP-negative mice. Quantification of GFP-expressing cells in wounds at 7, 14 and 28 days indicated similar amounts of allogeneic or syngeneic BM-MSCs but significantly reduced amounts of allo-fibroblasts. With healing progression, decreasing amounts of allogeneic and syngeneic BM-MSCs were found in the wound; however, the reduction was more evident (2 fold) in allo-fibroblasts. Similar effects in enhancing wound closure were found in allogeneic and syngeneic BM-MSCs but not in allo-fibroblasts. Histological analysis showed that allo-fibroblasts were largely confined to the injection sites while allo-BM-MSCs had migrated into the entire wound. Quantification of inflammatory cells in wounds showed that allo-fibroblast- but not allo-BM-MSC-treated wounds had significantly increased CD45+ leukocytes, CD3+ lymphocytes and CD8+ T cells. Our study suggests that allogeneic BM-MSCs exhibit ignorable immunogenicity and are equally efficient as syngeneic BM-MSCs in engraftment and in enhancing wound healing

Advances in Skin Substitutes—Potential of Tissue Engineered Skin for Facilitating Anti-Fibrotic Healing

Skin protects the body from exogenous substances and functions as a barrier to fluid loss and trauma. The skin comprises of epidermal, dermal and hypodermal layers, which mainly contain keratinocytes, fibroblasts and adipocytes, respectively, typically embedded on extracellular matrix made up of glycosaminoglycans and fibrous proteins. When the integrity of skin is compromised due to injury as in burns the coverage of skin has to be restored to facilitate repair and regeneration. Skin substitutes are preferred for wound coverage when the loss of skin is extensive especially in the case of second or third degree burns. Different kinds of skin substitutes with different features are commercially available; they can be classified into acellular skin substitutes, those with cultured epidermal cells and no dermal components, those with only dermal components, and tissue engineered substitutes that contain both epidermal and dermal components. Typically, adult wounds heal by fibrosis. Most organs are affected by fibrosis, with chronic fibrotic diseases estimated to be a leading cause of morbidity and mortality. In the skin, fibroproliferative disorders such as hypertrophic scars and keloid formation cause cosmetic and functional problems. Dermal fibroblasts are understood to be heterogeneous; this may have implications on post-burn wound healing since studies have shown that superficial and deep dermal fibroblasts are anti-fibrotic and pro-fibrotic, respectively. Selective use of superficial dermal fibroblasts rather than the conventional heterogeneous dermal fibroblasts may prove beneficial for post-burn wound healing

Prevention and management of scarring after thermal injury

Survival from burn injury has improved considerably over the past two decades such that the quality of life of the victim of thermal injuries has become a major concern. Severe proliferative scarring or hypertrophic scarring (HTS) is an all too frequent complication of burn wound healing that severely compromises quality of life for surviving burn victims. Prevention of such scarring in burn patients involves better understanding of the pathophysiology of scar formation, development of newer methods for determining depth of burn injury and earlier and advanced surgical interventions. Many established and evolving novel treatments for HTS in patients after thermal injury exist and include antifibrotic pharmaceuticals and cellular-based therapies as reviewed herein

Free Tissue Transfer in the Reconstruction of Neck Contractures after Burn Injury: A Case Series

Background: Recent advances in burn care have significantly improved the survival rate of patients with extensive burn injuries, placing greater emphasis on reconstruction to improve the long-term outcomes of scar deformities. Anterior and lateral neck contractures are common after burn injuries; they limit range of motion, complicate airway management and create significant cosmetic deformities. Traditional methods have been used to release contractures and improve function. However, they are subject to variable results, residual neck tightness, recurrence and suboptimal cosmetic appearance. Microvascular free tissue transfer is a more technically challenging and time-consuming method, but it offers the potential to overcome the long-term limitations of simpler options. In this paper, we present our experience with microvascular free flaps for the release of burn scar contractures of the neck as a potential high-quality permanent solution. Methods: Over a 10-year period, nine free flaps were performed on burn patients with total body surface area (TBSA) burns between 20 and 70%, who developed moderate to severe neck contractures. Four anterolateral thigh (ALT) flaps, four radial forearm free flaps (RFFFs) and one ulnar forearm flap were used to release neck contractures. Results: All nine flaps were completed successfully with significant improvement in the neck’s range of motion. Good aesthetic results were achieved with smooth contour and thin coverage. Overall, the patients were satisfied. However, five out of nine cases required at least one secondary procedure for flap defatting to reach optimal results. Conclusion: Post-burn scar contractures of the cervical region compromise the cosmetic appearance and airway security of recovering burn patients, imposing a significant impact on their psychological and functional quality of life. Consequently, cervical contractures can be prioritized when planning reconstruction for burn patients. Free flaps can be considered an important and reliable method of reconstruction for neck contracture deformity following burn injuries