Recessive Dystrophic Epidermolysis Bullosa Patients Research Articles

Longitudinal study of wound healing status and bacterial colonisation of Staphylococcus aureus and Corynebacterium diphtheriae in epidermolysis bullosa patients.

Epidermolysis bullosa (EB) is an inherited disorder characterised by skin fragility and the appearance of blisters and wounds. Patient wounds are often colonised or infected with bacteria, leading to impaired healing, pain and high risk of death by sepsis. Little is known about the impact of bacterial composition and susceptibility in wound resolution, and there is a need for longitudinal studies to understand healing outcomes with different types of bacterial colonisation. A prospective longitudinal study of 70 wounds from 15 severe EB patients (Junctional and Recessive Dystrophic EB) from Chile. Wounds were selected independently of their infected status. Wound cultures, including bacterial species identification, composition and Staphylococcus aureus (SA) antibiotic susceptibility were registered. Wounds were separated into categories according to their healing capacity, recognising chronic, and healing wounds. Hundred-one of the 102 wound cultures were positive for bacterial growth. From these, 100 were SA-positive; 31 were resistant to Ciprofloxacin (31%) and only seven were methicillin-resistant SA (7%). Ciprofloxacin-resistant SA was found significantly predominant in chronic wounds (**P < .01). Interestingly, atoxigenic Corynebacterium diphtheriae (CD) was identified and found to be the second most abundant recovered bacteria (31/101), present almost always in combination with SA (30/31). CD was only found in Recessive Dystrophic EB patients and not related to wound chronicity. Other less frequent bacterial species found included Pseudomonas aeruginosa, Streptococus spp. and Proteus spp. Infection was negatively associated with the healing status of wounds.

102 Citrullinated histone H3, a marker for neutrophil extracellular traps, is associated with poorer prognosis in recessive dystrophic epidermolysis bullosa patients with cutaneous squamous cell carcinoma

Recessive dystrophic epidermolysis bullosa (RDEB) is a severe hereditary skin disease characterized by skin and mucosa fragility with blister formation due to type VII collagen deficiency. The major complication in RDEB patients is the development of squamous cell carcinoma (SCC) leading to premature mortality. Our study aims to correlate the immune profiles of SCCs with clinico-histopathological features in a cohort of RDEB patients. The immune cell profile was assessed by immunohistochemical analyses on 33 SCCs from 19 RDEB patients included between March 2016 and December 2021.

090 Inhibition of HMGB1 attenuates the inflammatory response in keratinocytes

Recessive dystrophic epidermolysis bullosa (RDEB) is a blistering skin disease caused by mutations in COL7A1, the gene encoding type VII collagen (C7). RDEB is characterized by chronic wounds and a high incidence of aggressive squamous cell carcinoma (SCC), the leading cause of death in this patient population. High mobility group box 1 (HMGB1) is a serum biomarker of disease severity in RDEB patients that may have a mechanistic role in promoting SCC carcinogenesis. HMGB1 is a chromatin-associated protein that has a dual role as a mediator of DNA repair in the nucleus and a damage-associated molecular pattern that stimulates the innate immune response when secreted extracellularly in response to inflammatory stimuli or cellular damage.

713 The RNA-based reprogramming of renal epithelial cells derived from recessive dystrophic epidermolysis bullosa patients into induced pluripotent stem cells

No effective therapies are available for recessive dystrophic epidermolysis bullosa (RDEB) at this time. Reprogramming adult cells into induced pluripotent stem cells (iPSCs) may allow for the development of new therapies for RDEB. iPSCs can be derived from many cell types, such as fibroblasts, keratinocytes, and blood cells. The isolation of fibroblasts and keratinocytes requires a skin biopsy, an invasive procedure commonly associated with scarring and inflammation, and venipuncture is potentially traumatic for RDEB patients due to the fragility of their skin.

498 Artesunate inhibits RDEB fibrosis by downregulating AKT signaling pathway

Patients with recessive dystrophic epidermolysis bullosa (RDEB) develop multiple skin wounds that heal with extensive scarring, contractures, and mitten deformities. RDEB patients display increased pro-fibrotic TGF-β signaling, a distinct pro-fibrotic gene expression profile, and elevated inflammation genes. No specific pharmacological treatment is currently available for RDEB fibrosis. Artemisinin and its derivatives are anti-malarial medications that also exhibit both anti-fibrotic and anti-inflammatory activity in animal and cell models.

490 Splice modulation as a therapeutic strategy applied to deep intronic mutations in COL7A1 causing recessive dystrophic epidermolysis bullosa

Recessive dystrophic epidermolysis bullosa (RDEB) is a rare autosomal inherited skin disorder caused by mutations in COL7A1 encoding type VII collagen (C7), the major component of anchoring fibrils. In this study, we report two cases of RDEB patients with new pathogenic deep-intronic point mutations in COL7A1. Patient 1 is a 45-year-old man with RDEB inversa from unrelated healthy parents, for whom sequencing of COL7A1 from genomic DNA and cDNA extracted from skin biopsies revealed two recessive loss-of-function mutations.

Therapeutic base editing and prime editing of COL7A1 mutations in recessive dystrophic epidermolysis bullosa

Recessive dystrophic epidermolysis bullosa (RDEB) is a severe skin fragility disorder caused by loss-of-function mutations in the COL7A1 gene, which encodes type VII collagen (C7), a protein that functions in skin adherence. From 36 Korean RDEB patients, we identified a total of 69 pathogenic mutations (40 variants without recurrence), including point mutations (72.5%) and insertion/deletion mutations (27.5%). For fibroblasts from two patients (Pat1 and Pat2), we applied adenine base editors (ABEs) to correct the pathogenic mutation of COL7A1 or to bypass a premature stop codon in Pat1-derived primary fibroblasts. To expand the targeting scope, we also utilized prime editors(PEs) to correct the COL7A1 mutations in Pat1- and Pat2-derived fibroblasts. Ultimately, we found that transfer of edited patient-derived skin equivalents (i.e., RDEB keratinocytes and PE-corrected RDEB fibroblasts from the RDEB patient) intothe skin of immunodeficient mice led to C7 deposition and anchoring fibril formation within the dermal-epidermal junction, suggesting that base editing and prime editing could be feasible strategies for exvivo gene editing to treat RDEB.

T-cell activation and bacterial infection in skin wounds of recessive dystrophic epidermolysis bullosa patients.

Recessive dystrophic epidermolysis bullosa (RDEB) patients develop poorly healing skin wounds that are frequently colonized with microbiota. Because T cells play an important role in clearing such pathogens, we aimed to define the status of adaptive T cell‐mediated immunity in RDEB wounds. Using a non‐invasive approach for sampling of wound‐associated constituents, we evaluated microbial contaminants in cellular fraction and exudates obtained from RDED wounds. Infectivity and intracellular trafficking of inactivated Staphylococcus aureus was accessed in RDEB keratinocytes. S. aureus and microbial antigen‐specific activation of RDEB wound‐derived T cells were investigated by fluorescence‐activated cell sorting‐based immune‐phenotyping and T‐cell functional assays. We found that RDEB wounds and epithelial cells are most frequently infected with Staphylococcus sp. and Pseudomonas sp. and that S. aureus essentially infects more RDEB keratinocytes and RDEB‐derived squamous cell carcinoma cells than keratinocytes from healthy donors. The RDEB wound‐associated T cells contain populations of CD4+ and CD8+ peripheral memory T cells that respond to soluble microbial antigens by proliferating and secreting interferon gamma (IFNγ). Moreover, CD8+ cytotoxic T lymphocytes recognize S. aureus‐infected RDEB keratinocytes and respond by producing interleukin‐2 (IL‐2) and IFNγ and degranulating and cytotoxically killing infected cells. Prolonged exposure of RDEB‐derived T cells to microbial antigens in vitro does not trigger PD‐1‐mediated T‐cell exhaustion but induces differentiation of the CD4high population into CD4highCD25+FoxP3+ regulatory T cells. Our data demonstrated that adaptive T cell‐mediated immunity could clear infected cells from wound sites, but these effects might be inhibited by PD‐1/Treg‐mediated immuno‐suppression in RDEB.

In vivo topical gene therapy for recessive dystrophic epidermolysis bullosa: a phase 1 and 2 trial

Recessive dystrophic epidermolysis bullosa (RDEB) is a lifelong genodermatosis associated with blistering, wounding, and scarring caused by mutations in COL7A1, the gene encoding the anchoring fibril component, collagen VII (C7). Here, we evaluated beremagene geperpavec (B-VEC), an engineered, non-replicating COL7A1 containing herpes simplex virus type 1 (HSV-1) vector, to treat RDEB skin. B-VEC restored C7 expression in RDEB keratinocytes, fibroblasts, RDEB mice and human RDEB xenografts. Subsequently, a randomized, placebo-controlled, phase 1 and 2 clinical trial (NCT03536143) evaluated matched wounds from nine RDEB patients receiving topical B-VEC or placebo repeatedly over 12 weeks. No grade 2 or above B-VEC-related adverse events or vector shedding or tissue-bound skin immunoreactants were noted. HSV-1 and C7 antibodies sometimes presented at baseline or increased after B-VEC treatment without an apparent impact on safety or efficacy. Primary and secondary objectives of C7 expression, anchoring fibril assembly, wound surface area reduction, duration of wound closure, and time to wound closure following B-VEC treatment were met. A patient-reported pain–severity secondary outcome was not assessed given the small proportion of wounds treated. A global assessment secondary endpoint was not pursued due to redundancy with regard to other endpoints. These studies show that B-VEC is an easily administered, safely tolerated, topical molecular corrective therapy promoting wound healing in patients with RDEB.

Immune Disregulation in Cutaneous Squamous Cell Carcinoma of Patients with Recessive Dystrophic Epidermolysis Bullosa: A Single Pilot Study.

Background: Cutaneous squamous cell carcinoma (cSCC) is one of the most devastating complications of recessive dystrophic epidermolysis bullosa (RDEB). We recently demonstrated a reduction in immune cell peritumoral infiltration in RDEB patients with cSCC, together with a reduction in CD3+, CD4+, CD68+ and CD20 lymphocytes as compared to primary and secondary cSCC in patients without RDEB. Recently, new molecules, such as high mobility group box 1 (HMGB1), T cell immunoglobulin, mucin domain 3 (TIM-3) and Heme oxygenase-1 (HO-1), have been shown to play a role in antitumoral immunity. Objective: Patients with RDEB are known to be at increased risk of developing skin cancers, including the dreaded squamous cell carcinoma of the. Tendentially, cSCCs that arise in the context of EBDR are more aggressive and lead to statistically significant bad outcomes compared to cSCCs developed on the skin of patients without EBDR. In an attempt to study the microenvironment of these lesions, we conducted an immunohistochemical analysis study of proteins that could be actively involved in the genesis of this type of malignant neoplasms. Methods: In this retrospective study, the OH1-HMGB1-TIM3 activation axis, as correlated to the T lymphocytes cell count, was assessed in biopsy samples from 31 consecutive cases consisting of 12 RDEB patients with cSCC, 12 patients with primary cSCC and 7 RDEB patients with pseudoepitheliomatous cutaneous hyperplasia. Parametric Student’s t-test was applied for normally distributed values, such as CD4+ and CD8+, and non-parametric Mann–Whitney test for non-normally distributed values, such as HMGB-1, TIM-3 and HO-1. Results: In RDEB patients with cSCC and with pseudoepitheliomatous hyperplasia, the expression of CD4 T helper lymphocytes was lower than in the peritumoral infiltrate found in primary cSCC. CD8 cytotoxic T lymphocytes were increased in primary cSCC compared to the other two groups. An increased HMGB1 expression was evident in both primary and RDEB cSCC. TIM3 expression was higher in RDEB patients with cSCC compared to the other two groups. A significantly reduced immunohistochemical expression of HO-1 was evident in the tumoral microenvironment of cSCC-RDEB as compared to primary cSCC. Conclusions: These data suggest that a reduced immune cell peritumoral infiltration in RDEB patients could be responsible, in the complexity of the mechanisms of carcinogenesis and host response, of the particular aggressiveness of the cSCC of RDEB patients, creating a substrate for greater local immunosuppression, which, potentially, can “open the doors” to development and eventual metastasis by this malignant neoplasm.

Oral microbiome in children with dystrophic recessive epidermolysis bullosa

Was to assess bacterial load in oral wounds in children with recessive dystrophic epidermolysis bullosa (RDEB). The study comprised 77 RDEB children aged 3-18 years (mean age 9.5±3.6 years) and 27 healthy children aged 4-18 years (mean age 9.8±4.1 years) who served as controls. Swabs for bacteriological study were taken from the oral wounds in RDEB patients and non-affected corresponding oral mucosa areas in controls. The microorganism growth was assessed after 24, 48 and 72 hours of incubation (37 °C and 30 °C) with subsequent identification in automatic bacteriological analyzer MicroScan Walk Away (Simens, USA). Results. The study revealed high prevalence and concentrations of Candida albicans (in 40.3% children), Staphilococcus aureus (23.4%), Enterobacter cloacae (9.1%), and Enterobacteria (10.4%) in RDEB children. From these species, only Candida albicans was present in controls (26%). The prevalence and concentration of commensal and pathogenic species correlated positively with age and significant difference was revealed between children at the age of 3-6 and 7-10 years (p=0.001). Thus, bacterial load in oral wounds correlates with the RDEB natural history and fibrosis progression. Delayed oral wound healing was associated with microbiome shift towards biofilm-producing bacteria Staphilococcus aureus and Enterobacter cloacae. Oral wounds microbiome may be an indicator of RDEB severity and tendency to oral fibrosis. The decrease of bacterial load in the oral wounds may remove one of the healing obstacles and serve as fibrosis prevention measure.

Development of Minicircle Vectors Encoding COL7A1 Gene with Human Promoters for Non-Viral Gene Therapy for Recessive Dystrophic Epidermolysis Bullosa.

Recessive dystrophic epidermolysis bullosa (RDEB) is a rare autosomal inherited skin disorder caused by mutations in the COL7A1 gene that encodes type VII collagen (C7). The development of an efficient gene replacement strategy for RDEB is mainly hindered by the lack of vectors able to encapsulate and transfect the large cDNA size of this gene. To address this problem, our group has opted to use polymeric-based non-viral delivery systems and minicircle DNA. With this approach, safety is improved by avoiding the usage of viruses, the absence of bacterial backbone, and the replacement of the control viral cytomegalovirus (CMV) promoter of the gene with human promoters. All the promoters showed impressive C7 expression in RDEB skin cells, with eukaryotic translation elongation factor 1 α (EF1α) promoter producing higher C7 expression levels than CMV following minicircle induction, and COL7A1 tissue-specific promoter (C7P) generating C7 levels similar to normal human epidermal keratinocytes. The improved system developed here has a high potential for use as a non-viral topical treatment to restore C7 in RDEB patients efficiently and safely, and to be adapted to other genetic conditions.

361 Identification of a new molecular modulator of severity in recessive dystrophic epidermolysis bullosa

Recessive dystrophic epidermolysis bullosa (RDEB) is a rare disorder characterized by cutaneous fragility of the skin and mucous, due to COL7A1 mutations. Patients with RDEB have a poor quality of life as result of chronic wounds that trigger fibrotic processes and facilitate the development of highly aggressive squamous cell carcinomas. To find new disease-severity modulators, we previously reported a transcriptomic study (RNA-Seq) in two RDEB siblings with marked phenotypic differences despite being genotypically identical for COL7A1 (c.6527insC/c.6527insC). Enrichment analysis of the differentially expressed genes suggested differences regarding the extracellular matrix (ECM), cell contraction, and response to TGF-β, among others. Here, we explored in detail the biological relevance of some of these differentially altered processes. In particular, fibroblasts from the less affected sibling (M-RDEB) showed reduced response to TGF-β stimulus and lower contractile capacity of collagen lattices, contrary to what was observed in other RDEB patients (including the sister with severe phenotype) and similar to the behavior of fibroblasts from healthy donors. Interestingly, M-RDEB fibroblasts upregulate a previously undisclosed ECM protein capable of inhibiting the TGF-β pathway, and presumably responsible -at least in part- for the mild phenotype observed in M-RDEB. Overexpression of the gene encoding this factor in fibroblasts from the sister with severe phenotype (S-RDEB) counteracted TGF-β response and cell contraction capacity in in vitro functional assays. A similar result was obtained when treating S-RDEB fibroblasts with the molecule as a recombinant protein. Thus, our study identified new modulators and druggable targets in RDEB pathophysiology, which would enable novel symptom relief therapies to tackle disease progression.

164 Molecular characterization of the response to haploidentical mesenchymal stem cells treatment in recessive dystrophic epidermolysis bullosa patients

Recessive dystrophic epidermolysis bullosa (RDEB) is an incurable rare skin fragility disorder characterized by deficient dermo-epidermal adhesion due to mutations in the gene encoding collagen VII (COL7A1). Patients with RDEB suffer chronic wounds and inflammation, exacerbated by pruritus, and ultimately trigger fibrosis and highly aggressive squamous cell carcinoma. Although classified as a skin disease, its symptoms affect multiple internal organs and tissues, which underscores the need for systemic treatment; allogenic mesenchymal stem cells (MSCs) derived from bone marrow have previously shown some benefit in patients with RDEB. The goal of this work is to understand the mechanisms by which MSCs exert their therapeutic effect in the unique context provided by a clinical trial (NCT04153630) in addition to characterization of the baseline profile of the recruited RDEB patients. Comparative transcriptomic analysis (RNA-Seq) of skin biopsies from 8 patients, before and after the MSC infusion, revealed differential expression of 41 genes, including some related to key functions such as inflammatory response (FOS, osteopontin) or pruritus (corneodesmosin). An enrichment analysis provided further insight highlighting some biological processes such as barrier function and oxidative stress. Finally, a further interactome network and mechanistic analysis showed that JAK-Stat and FoxO signaling pathways are modulated as a result of the MSC action. These preliminary data might be key for predicting the outcome of patient’s condition, understanding the global and individual response to MSCs and for optimization of treatment, as well as the development of new ones.

The clinical efficacy and safety of anti-IgE therapy inrecessivedystrophic epidermolysis bullosa.

The treatment of recessive dystrophic epidermolysis bullosa (RDEB) remains challenging. Elevated IgE levels have previously been reported in several RDEB patients. In this prospective, single-centre, open intervention study, elevated IgE levels were seen in 11 out of 12 patients with intense pruritus, and the patients with elevated IgE levels received anti-IgE therapy every 4 weeks for at least three cycles. Compared with the baseline, 10 patients with RDEB had good clinical outcomes with enhanced wound healing, a reduction in Birmingham (epidermolysis bullosa) EB severity score by 15%, a reduction in affected body surface area by 23.3%, amelioration of skin inflammation, and an increase in type VII collagen deposition by 13.1-fold. All the patients had a good tolerance to anti-IgE therapy. Furthermore, patients with higher IgE levels tended to have higher disease severity and more favorable clinical outcomes. Our report also suggested the potential role of IgE in the pathogenesis of inflammatory conditions associated with RDEB. (ChiCTR1900021437).

Role of transforming growth factor-β1 in recessive dystrophic epidermolysis bullosa squamous cell carcinoma.

Squamous cell carcinoma (SCC) develops in more than 80% of individuals with the skin blistering disorder recessive dystrophic epidermolysis bullosa (RDEB). In contrast with UV-induced SCC, RDEB-SCC results from skin damage and has a high proliferative and metastatic rate with 5-year survival near zero. Our objective is to determine the mechanisms underlying the increased metastatic tendencies of RDEB-SCC. RDEB-SCC cultured cell lines were treated with RDEB and non-RDEB fibroblast conditioned media and assayed for migration and invasion with and without small molecule inhibitors for TGFβ and other downstream signal transduction pathways. TGFβ1 secreted by RDEB dermal fibroblasts has been found to induce migration and invasion and to increase expression of epithelial-to-mesenchymal transition markers in an RDEB-SCC line. These effects were reversed upon inhibition of TGFβ signalling and its downstream pathways MEK/ERK, P38 kinase and SMAD3. A number of small molecule inhibitors for these pathways are in different phases of various clinical trials and may be applicable to RDEB-SCC patients. Studying the mechanisms of the extreme form RDEB-SCC may inform studies of other types of SCC, as well as lead to better therapies for RDEB patients.

Over-expression of stromal periostin correlates with poor prognosis of cutaneous squamous cell carcinomas.

Periostin, an extracellular matrix macromolecule implicated in tumorigenesis, serves as a prognostic marker for many cancer types. However, there are no data on periostin expression in cutaneous squamous cell carcinoma (cSCC). This study examined periostin expression in patients with cSCC and explored its clincopathological relationship and prognosis. Using immunohistochemistry and ImageJ analysis, we compared periostin expression in 95 cSCCs across a spectrum of cSCC aggressiveness: cSCC in situ (SCCIS) (n=25), low-risk cSCC (LR-cSCC) (n=26), high-risk cSCC (HR-cSCC) (n=38), and cSCC in recessive dystrophic epidermolysis bullosa patients (RDEB cSCC) (n=6). Immunohistochemistry demonstrated periostin expression within the intra-tumoral stroma but not within tumor cells. Periostin levels significantly (P<0.001) increased from SCCIS, LR-cSCC, HR-cSCC to RDEB SCC. The stroma of most of the cSCCs we evaluated contained cancer-associated fibroblasts with a myofibroblastic (α -SMA-positive) phenotype. Co-localization of periostin with α-SMA, evidence of fibroblast periostin expression, and absence of keratinocyte or tumor cell periostin expression suggest that, in cSCC, periostin is a product of the peritumoral microenvironment and not the tumor cells themselves. Our data indicate that fibroblast periostin expression is highly correlated with the aggressiveness of cSCC, and may thereby provide a molecular marker that will be useful for subtyping and diagnosing cSCCs according to their biological nature.

Inherited epidermolysis bullosa: epidemiology and patient care in Slovenia with a review of the updated classification

Inherited epidermolysis bullosa (EB) is a heterogeneous group of rare genetic skin disorders characterized by fragility of the skin and mucous membranes. The prevalence of all types of EB is estimated at approximately 11 per million, based on recent data from the American National Epidermolysis Bullosa Registry. A national registry of EB has not yet been established in Slovenia. Because all cases of EB are diagnosed and treated at our department, we have collected data on all known cases of EB in Slovenia. Based on our data, the prevalence of all EB types in Slovenia is about 20 per million. As of December 2020, our data consist of 29 EB simplex, three junctional EB, 10 dominant dystrophic EB, and four recessive dystrophic EB patients. The prevalence of all EB types in Slovenia is higher compared to the estimated prevalence in the United States. The multidisciplinary care of EB patients in Slovenia has been developed based on patients' needs, including a wide group of various specialists, and it has been adapted to the resources and treatment options available. This article also reviews the up-to-date classification and diagnostic protocol for EB, and international recommendations for interdisciplinary patient care.

Transplantation of autologous single hair units heals chronic wounds in autosomal recessive dystrophic epidermolysis bullosa: A proof-of-concept study

Autosomal recessive dystrophic epidermolysis bullosa (RDEB) is characterized by recurrent mucocutaneous blistering with non-healing ulcers which are often complicated by squamous cell carcinoma (SCC). Despite having as high as 80% death rate from SCC, RDEB still does not have an effective treatment. We report on the efficacy of single follicular unit extract (FUE) grafting to heal chronic ulcers of intermediate RDEB in a 54-year-old woman with extensive chronic wounds covering around 30% of the body surface area. On Day 17 post first graft session, the area of treated ulcers on her right upper back was reduced by 80%. Immunofluorescence study revealed positive type VII collagen expression along the epidermal and follicular basement membrane zone in the donor and recipient sites. A few grafted follicles continued to grow hair on the recipient sites. A total of 360 FUEs were grafted in nine sessions over five years, resulting in healing of most treated ulcers and reduced significantly her time for daily wound dressing. Importantly, FUE grafting using patient's own scalp follicles does not require any laboratory manipulation. It is safe and easy to perform. Autologous follicular grafting appears efficacious for healing of recalcitrant wounds and provides an innovative solution for RDEB patients with such wounds.

Aberrant recruitment of leukocytes defines poor wound healing in patients with recessive dystrophic epidermolysis bullosa

BackgroundPoorly healing wounds are one of the major complications in patients suffering from recessive dystrophic epidermolysis bullosa (RDEB). At present, there are no effective means to analyze changes in cellular and molecular networks occurring during RDEB wound progression to predict wound outcome and design betted wound management approaches. ObjectivesTo better define mechanisms influencing RDEB wound progression by evaluating changes in molecular and cellular networks. MethodsWe developed a non-invasive approach for sampling and analysis of wound-associated constituents using wound-covering bandages. Cellular and molecular components from seventy-six samples collected from early, established and chronic RDEB wounds were evaluated by FACS-based immuno-phenotyping and ELISA. ResultsOur cross-sectional analysis determined that progression of RDEB wounds to chronic state is associated with the accumulation (up to 90 %) of CD16+CD66b+ mature neutrophils, loss of CD11b+CD68+ macrophages, and a significant increase (up to 50 %) in a number of CD11c+CD80+CD86+ activated professional antigen presenting cells (APC). It was also marked by changes in activated T cells populations including a reduction of CD45RO+ peripheral memory T cells from 80 % to 30 % and an increase (up to 70 %) in CD45RA+ effector T cells. Significantly higher levels of MMP9, VEGF-A and cathepsin G were also associated with advancing of wounds to poorly healing state. ConclusionsOur data demonstrated that wound-covering bandages are useful for a non-invasive sampling and analysis of wound-associated constituents and that transition to poorly healing wounds in RDEB patients as associated with distinct changes in leukocytic infiltrates, matrix-remodeling enzymes and pro-angiogenic factors at wound sites.