[Targeted therapies in Pyoderma gangrenosum: deciphering pathophysiology and improving disease management]. / Pyoderma gangraenosum: durch zielgerichtete Therapieansätze besser verstehen und behandeln können.

The immune response is a central process during wound healing. Malfunctions often lead to chronic inflammation, barrier disorders, and ulcerations of the skin. The underlying pathomechanisms are complex and the subject of current dermatological research. The care of wound healing disorders is still inadequate and urgently needs improved therapy concepts. For several years now, the development of modern immunomodulators has enabled the targeted regulation of specific signaling cascades, and their effectiveness in the treatment of wound healing disorders has been proven in numerous case studies. Thus, their use not only leads to more efficient therapeutic approaches, but also provides deeper insight into the pathomechanistic importance of specific signaling pathways in inflammatory and degenerative diseases of the skin, which are poorly understood so far. Pyoderma gangrenosum, an autoinflammatory disease, provides a good example to illustrate the progress in therapy and pathomechanistic understanding through the use of new immunomodulators and is explained in more detail in the following article.

[Current pathophysiological developments in fibrosing diseases: insights into novel treatment concepts]. / Aktuelle pathophysiologische Entwicklungen bei fibrosierenden Erkrankungen: Ansatzpunkte für neue Konzepte in der Therapie.

Fibrosis is a common symptom of a variety of skin disorders of diverse entity. The cellular and molecular pathophysiology of fibrosis development is unresolved and current treatment options are not sufficient. Tissue fibrosis leads to increased tissue stiffness, impaired organ function, decline of quality of life and ultimately increased morbidity and mortality. Epidemiologic studies indicate that nearly 45% of all deaths in the western world are associated with tissue fibrosis in diverse organs. Only few recently approved treatment options specifically target the process of fibrogenesis. The development of novel and efficient therapies is urgently needed, and at the same time provides a major challenge but also an opportunity to contribute to the advancement of this unresolved medical problem. This article highlights recent insights in the developments on tissue fibrosis with a focus on immunoregulatory mechanisms.

[Wound healing in the elderly]. / Wundheilung im Alter.

Restoration of tissue integrity is essential for host defense and protection of the organism. The efficacy and quality of skin repair varies significantly over a person's lifetime. Whereas prenatal wound healing is characterized by regeneration and scarless healing, scarring, fibrosis, and loss of function are features of postnatal repair. In fact, aging is the prominent risk factor for chronic wounds, skin fragility, infections, comorbidities, and decreased quality of life. Current strategies for restoration of tissue integrity and wound therapy are not sufficient and require further investigation of the underlying pathomechanisms and the development of causal-based concepts.

Protease-modulating polyacrylate-based hydrogel stimulates wound bed preparation in venous leg ulcers--a randomized controlled trial.

BACKGROUND: Stringent control of proteolytic activity represents a major therapeutic approach for wound-bed preparation. OBJECTIVES: We tested whether a protease-modulating polyacrylate- (PA-) containing hydrogel resulted in a more efficient wound-bed preparation of venous leg ulcers when compared to an amorphous hydrogel without known protease-modulating properties. METHODS: Patients were randomized to the polyacrylate-based hydrogel (n = 34) or to an amorphous hydrogel (n = 41). Wound beds were evaluated by three blinded experts using photographs taken on days 0, 7 and 14. RESULTS: After 14 days of treatment there was an absolute decrease in fibrin and necrotic tissue of 37.6 ± 29.9 percentage points in the PA-based hydrogel group and by 16.8 ± 23.0 percentage points in the amorphous hydrogel group. The absolute increase in the proportion of ulcer area covered by granulation tissue was 36.0 ± 27.4 percentage points in the PA-based hydrogel group and 14.5 ± 22.0 percentage points in the control group. The differences between the groups were significant (decrease in fibrin and necrotic tissue P = 0.004 and increase in granulation tissue P = 0.0005, respectively). CONCLUSION: In particular, long-standing wounds profited from the treatment with the PA-based hydrogel. These data suggest that PA-based hydrogel dressings can stimulate normalization of the wound environment, particularly in hard-to-heal ulcers.

[Parallels between wound healing, chronic inflammatory skin diseases and neoplasia: clinical aspects]. / Parallelen zwischen Wundheilung, chronisch entzündlichen Dermatosen und Neoplasien: Wissenswertes für die Praxis.

Chronic wounds, scars, burns and recalcitrant chronic inflammatory skin lesions can give rise to malignancy. These neoplasias are usually squamous cell carcinomas but basal cell carcinomas can also develop. Tumorigenesis is a severe complication of chronic ulcers as well as certain inflammatory skin diseases; early diagnosis is critical for prognosis. This article describes parallels between wound healing, chronic inflammatory skin diseases and carcinogenesis and provides advice on practical aspects of diagnosis and therapy.

[Delayed wound healing during therapy of cutaneous graft-versus-host disease with everolimus]. / Wundheilungsstörung bei kutaner Graft-versus-Host-Disease unter Therapie mit Everolimus.

Graft-versus-host-disease (GvHD) is despite improvement in transplantation medicine the major cause for morbidity and mortality after allogeneic stem cell transplantation. We describe a patient with chronic cutaneous GvHD who developed massive skin ulcerations after changing the immunosuppressive therapy to a mammalian target of rapamycin (mTOR)-inhibitor.

[Autoinflammatory diseases as cause of wound healing defects]. / Autoinflammatorische Erkrankungen als Ursache von Wundheilungsstörungen.

Ulcerations of the skin and mucosal membranes are a common feature of autoinflammatory diseases. They can give raise to chronic wound healing defects and should be considered in the differential diagnosis of chronic skin ulcers. The increased activation of the innate immune system in the absence of an apparent provocation for inflammation is a hallmark of autoinflammatory diseases. Mutations and alterations of signaling pathways regulating the innate immune response to physical trauma/tissue damage result into an unrestrained activation of the inflammasome, which leads to increased activation of Interleukin-1. Uncontrolled recruitment and activation of myeloid effector cells within the wound site lead to the release of potent proteases that cause the degradation of structural components of the skin. The majority of these diseases respond well to immunosuppressive and immunomodulatory treatment regimes. Therapeutic resistance converts the acute inflammatory response into a chronic and non-resolving inflammatory process that leads to tissue degeneration. In this article we will focus on the review of those autoinflammatory diseases that often display ulcerative cutaneous and aphthous lesions including pyoderma gangrenosum, Behçet disease, PAPA syndrome and hyperimmunoglobulinemia D with periodic fever syndrome (HIDS). Furthermore, the article will be complemented by an overview of those inflammatory diseases that are associated with non-ulcerative cutaneous manifestations.

Mitochondrial respiration controls neoangiogenesis during wound healing and tumour growth.

The vasculature represents a highly plastic compartment, capable of switching from a quiescent to an active proliferative state during angiogenesis. Metabolic reprogramming in endothelial cells (ECs) thereby is crucial to cover the increasing cellular energy demand under growth conditions. Here we assess the impact of mitochondrial bioenergetics on neovascularisation, by deleting cox10 gene encoding an assembly factor of cytochrome c oxidase (COX) specifically in mouse ECs, providing a model for vasculature-restricted respiratory deficiency. We show that EC-specific cox10 ablation results in deficient vascular development causing embryonic lethality. In adult mice induction of EC-specific cox10 gene deletion produces no overt phenotype. However, the angiogenic capacity of COX-deficient ECs is severely compromised under energetically demanding conditions, as revealed by significantly delayed wound-healing and impaired tumour growth. We provide genetic evidence for a requirement of mitochondrial respiration in vascular endothelial cells for neoangiogenesis during development, tissue repair and cancer.

Phase-specific functions of macrophages determine injury-mediated corneal hem- and lymphangiogenesis.

Macrophages are critical mediators of injury-associated corneal hemangiogenesis (HA) and lymphangiogenesis (LA). Yet, molecular regulators of the hem- and lymphangiogenic potential of corneal wound macrophages are poorly understood. Using two different mouse models of acute (perforating corneal incision injury) and chronic (corneal suture placement model) corneal injury, here we identified distinct functions of early- versus late-phase corneal wound macrophages in corneal HA and LA. Whereas early-phase wound macrophages are essential for initiation and progression of injury-mediated corneal HA and LA, late-phase wound macrophages control maintenance of established corneal lymphatic vessels, but not blood vessels. Furthermore, our findings reveal that the hem- and lymphangiogenic potential of corneal wound macrophages is controlled by the type of the corneal damage. Whereas perforating corneal incision injury induced primarily wound macrophages with lymphangiogenic potential, corneal suture placement provoked wound macrophages with both hem- and lymphangiogenic potential. Our findings highlight a previously unrecognized injury-context dependent role of early- versus late-phase corneal wound macrophages with potential clinical impact on therapy development for sight-threatening corneal neovascular diseases.

Skin fibrosis: Models and mechanisms.

Matrix synthesis, deposition and remodeling are complex biological processes that are critical in development, maintenance of tissue homeostasis and repair of injured tissues. Disturbances in the regulation of these processes can result in severe pathological conditions which are associated with tissue fibrosis as e.g. in Scleroderma, cutaneous Graft-versus-Host-Disease, excessive scarring after trauma or carcinogenesis. Therefore, finding efficient treatments to limit skin fibrosis is of major clinical importance. However the pathogenesis underlying the development of tissue fibrosis is still not entirely resolved. In recent years progress has been made unraveling the complex cellular and molecular mechanisms that determine fibrosis. Here we provide an overview of established and more recently developed mouse models that can be used to investigate the mechanisms of skin fibrosis and to test potential therapeutic approaches.

Genetically modified human keratinocytes overexpressing PDGF-A enhance the performance of a composite skin graft.

Skin loss due to burns and ulcers is a major medical problem. Bioengineered skin substitutes that use cultured keratinocytes as an epidermal layer with or without analogues of the dermis are one strategy for skin repair. However, none can achieve definitive wound closure, function, or cosmesis comparable to split-thickness autografts. Moreover, autograft donor sites, which require time to heal, may be limited or have attendant problems such as infection or functional/cosmetic deficiencies. To determine if the performance of composite skin grafts of keratinocytes on a dermal analogue could be enhanced, human keratinocytes were genetically modified to overexpress platelet-derived growth factor A chain (PDGF-A). Composite grafts of modified keratinocytes seeded onto acellular dermis, prepared from cryopreserved cadaver skin, secreted PDGF-AA protein in vitro [90 ng/graft (1.5 x 1.5 cm)/24 hr]. To test their performance in a wound healing model, composite grafts were transplanted to full-thickness excisional wounds on the back of athymic mice. PDGF-A grafts formed a stratified differentiated epidermis similar to control grafts. The acellular dermis was repopulated with host fibrovascular cells and by day 7, the PDGF-A grafts had significantly more cells in the dermis and increased staining for murine collagen types I and IV. At this early time point, wound contraction was also significantly inhibited in PDGF-A grafts versus control grafts. Thus, PDGF-A overexpression improves graft performance during the first critical week after transplantation.

Targeted expression of insulin-like growth factor to human keratinocytes: modification of the autocrine control of keratinocyte proliferation.

Somatomedin C/insulin-like growth factor-I (IGF-I) is required for the proliferation of keratinocytes in vitro. In skin, the cells known to synthesize IGF-I are melanocytes and fibroblasts of the dermis. To investigate the role of IGF-I as a mediator of keratinocyte proliferation, we have used retroviral-mediated gene transfer to introduce the gene encoding human IGF-I into diploid human keratinocytes, thus causing these cells to produce a growth factor they normally do not express. Modified cells synthesized and secreted significant levels of IGF-I (560 ng/10(7) cells/24 h) in vitro. Cells expressing IGF-I were no longer dependent on exogenously added IGF-I or insulin for their sustained growth in vitro under serum-free conditions. The growth of these cells did require added epidermal growth factor (EGF) and bovine pituitary extract. The addition of an antibody that neutralizes IGF-I inhibited cell growth, suggesting that IGF-I must be secreted by the cells to promote cell proliferation. To investigate the role of IGF-I in vivo, we grafted modified keratinocytes expressing IGF-I onto athymic mice. Grafts of epithelial sheets of modified cells formed a stratified epithelium comparable to control grafts of unmodified cells. When analyzed for keratin 16 expression and by quantitative staining for the nuclear proliferation antigen Ki-67, however, modified epithelia showed an increase in these markers of proliferation when compared with grafts of unmodified cells. This study demonstrates that genetic modification can be used to modify the autocrine control of keratinocyte proliferation. The de novo synthesis of IGF-I by keratinocytes could sustain keratinocytes growth in vitro and stimulate proliferation in vivo without significantly altering epidermal differentiation. These data further support the role of IGF-I as a paracrine mediator of epidermal proliferation and as a potential signal of mesenchymal-epithelial interactions.

Particle-mediated gene transfer of PDGF isoforms promotes wound repair.

Several techniques for cutaneous gene transfer have been investigated for either in vitro or in vivo applications. In the present study, we investigated whether the direct delivery of platelet-derived growth factor cDNA into skin results in improvement in tissue repair. Cutaneous transfections were carried out in rats using a particle-bombardment device (Accell). As revealed by reverse transcriptase-polymerase chain reaction, transgene expression in vivo was transient, with low level expression by day 5. When compared with wounds transfected with a control cytomegalovirus-luciferase plasmid, wounds transfected with platelet-derived growth factor A or B in the MFG vector showed a significant increase in wound tensile strength 7 and 14 d after transfection. At both time points platelet-derived growth factor A transfected wounds exhibited the highest increase in tensile strength over controls, resulting in a 3.5-fold increase at day 7 and a 1.5-fold increase at day 14. The degree of stimulation was not remarkably different between wounds transfected with platelet-derived growth factor B, which is predominantly cell associated, or a truncation mutant, platelet-derived growth factor B211, which is predominantly secreted. These findings demonstrate that in vivo gene transfer by particle bombardment can be used to improve the tissue repair response. This approach provides a robust tool to assess the biologic activity of various proteins and will aid in the development of therapeutic cutaneous gene delivery.

Evaluation of human skin reconstituted from composite grafts of cultured keratinocytes and human acellular dermis transplanted to athymic mice.

This study evaluates the use of composite grafts of cultured human keratinocytes and de-epidermalized, acellular human dermis to close full-thickness wounds in athymic mice. Grafts were transplanted onto athymic mice and studied up to 8 wk. Graft take was excellent, with no instances of infection or graft loss. By 1 wk, the human keratinocytes had formed a stratified epidermis that was fused with mouse epithelium, and by 8 wk the grafts resembled human skin and could be freely moved over the mouse dorsum. Immunostaining for keratins 10 and 16 and for involucrin revealed an initial pattern of epithelial immaturity, which by 8 wk had normalized to that of mature unwounded epithelium. Mouse fibroblasts began to infiltrate the acellular dermis as early as 1 wk. By 8 wk fibroblasts had completely repopulated the dermis, and blood vessels were evident in the most superficial papillary projections. Dermal elements, such as rete ridges and elastin fibers, which were present in the starting dermis, persisted for the duration of the experiment. Grafts using keratinocytes from dark-skinned donors as opposed to light-skin donors had foci of pigmentation as early as 1 wk that progressed to homogenous pigmentation of the graft by 6 wk. These results indicate that melanocytes that persist in vitro are able to resume normal function in vivo. Our study demonstrates that composite grafts of cultured keratinocytes combined with acellular dermis are a useful approach for the closure of full-thickness wounds.

Genetically modified human epidermis overexpressing PDGF-A directs the development of a cellular and vascular connective tissue stroma when transplanted to athymic mice--implications for the use of genetically modified keratinocytes to modulate dermal regeneration.

We investigated the hypothesis that keratinocyte-produced platelet-derived growth factor-AA (PDGF-AA) is involved in epidermal-dermal interactions and that PDGF-AA is an important mediator of the temporal and spatial events of tissue repair. Retroviral-mediated gene transfer was used to introduce the gene encoding human PDGF-A into cultures of human diploid keratinocytes. Genetic modification boosted the endogenous in vitro level of PDGF-AA secretion by over 300 fold. When PDGF-secreting cells were transplanted as epithelial sheets to athymic mice, modified keratinocytes underwent terminal differentiation and generated a stratified epithelium comparable to unmodified cells. Seven days after grafting the newly synthesized connective tissue layer subjacent to the PDGF-A-modified grafts was significantly thicker, was rich in mononuclear cells and fibroblasts, and had increased numbers of blood vessels when compared to control grafts of unmodified cells. These results suggest that PDGF-AA secreted by the epidermis is an important mediator of epithelial-mesenchymal interactions and helps to promote growth and vascularization of the underlying dermal tissue. Further, these data demonstrate the feasibility of using genetically modified cells to modulate tissue regeneration.

Expression and proteolysis of vascular endothelial growth factor is increased in chronic wounds.

Degradation of angiogenic mediators might be an underlying cause of chronic wounds. To test this hypothesis, we evaluated the expression and integrity of vascular endothelial growth factor, a potent angiogenic mediator, and its receptors, Flt-1 and KDR, in chronic venous leg ulcerations. Immunohisto- chemical, in situ hybridization, and semiquantitative reverse transcriptase polymerase chain reaction analyses all indicate that expression of vascular endothelial growth factor is elevated in ulcerative tissue, with vascular endothelial growth factor mRNA being especially pronounced in the hyperplastic epithelium of the wound margin. Flt-1 and KDR protein and mRNA were detected in the papillary vessels in close vicinity to the lesional epithelium of chronic wounds. Although increased expression of vascular endothelial growth factor protein was detected in the epidermis, the intensity of this staining was weak compared with the epidermal staining in psoriatic lesions and compared with the strong vascular endothelial growth factor mRNA signal in chronic wounds and psoriasis. To analyze whether this apparent decrease in immunoreactivity could be the result of degradation of vascular endothelial growth factor by proteolytic activities from the wound environment, we examined the stability of recombinant vascular endothelial growth factor in wound fluid from chronic leg ulcers. As demonstrated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, incubation of rVEGF165 with chronic, but not acute, wound fluid resulted in rapid proteolytic degradation of rVEGF165. Protease inhibitor studies indicate that serine proteases, such as plasmin, are involved in this degradation. Together, our data show that, although vascular endothelial growth factor expression is elevated in chronic wounds, increased proteolytic activity in this environment results in its degradation, which may contribute to an impaired wound healing response.