Cryopreserved and lyophilized cultured epidermal allografts in the treatment of leg ulcers: a pilot study

BACKGROUND: In the conservative therapy of venous leg ulcers modern types of dressings are used most frequently. In the past 20 years 'active wound dressings' - cultured epidermal keratinocytes as autografts and allografts - were also introduced in the management of leg ulcers. METHODS: The aim of our study was to compare the effect of cryopreserved and lyophilized cultured epidermal allografts in the treatment of venous leg ulcers. Evaluation of the therapy was documented as photodocumentation, planimetry, healing time and evaluation of pain relief over a 3-month period after application. Fifty patients with venous leg ulcers were selected. Twenty-five patients (group I) were treated with cryopreserved keratinocytes and 25 (group II) with lyophilized keratinocytes. RESULTS: The final evaluation 3 months after the application of allografts showed 84% of healed ulcers in group I and 80% in group II. The number of healed ulcers and the healing rate both showed no statistically significant differences. The size of the ulcer was reduced by half during the first week in both groups. The size differences during the first week are statistically significant in both groups and they are comparable (P < 0.001). The intensity of the pain was statistically significantly reduced during the first week after application in both groups (P < 0.001). CONCLUSIONS: The cryopreserved and lyophilized cultured allografts are comparable in healing rate, course of healing and relief of pain, and also in planimetric changes during the healing of venous leg ulcers. Lyophilized allografts are more convenient for routine use than cryopreserved allografts as they can be stored at room temperature. These results could give rise to the more frequent use of lyophilized allografts in slow-healing venous leg ulcers.     

The longevity of a bilayered skin substitute after application to venous ulcers

BACKGROUND: A bilayered skin substitute composed of allogeneic keratinocytes and fibroblasts in a collagen gel has been approved by the US Food and Drug Administration for the treatment of venous and diabetic ulcers. Its mechanism of action has not been fully determined. OBJECTIVE: To determine the longevity of allogeneic fibroblasts and keratinocytes in a bilayered skin substitute in patients with venous leg ulcers. METHODS: Ten patients with venous leg ulcers were treated with a bilayered skin substitute on day 0, days 3 to 5, and weeks 1 through 3. Biopsy specimens of the grafted wound were taken. We used polymerase chain reaction analysis to determine whether allogeneic DNA was present in the biopsy specimens. RESULTS: We detected allogeneic DNA in 2 of 8 specimens at 1 month after initial grafting. Neither of the 2 patients showed persistence of allogeneic DNA at 2 months after initial grafting. CONCLUSIONS: Allogeneic cells from a bilayered skin substitute do not appear to survive permanently after grafting for treatment of venous leg ulcers. Other mechanisms of action might include cytokine release, structural support, or provision of a moist wound environment.     

Unterschiedliche Abheilungsdauer und Häufigkeit der Hospitalisation bei Ulcus cruris verschiedener Ursachen

Background

Leg ulcers are a symptom of a heterogeneous group of diseases. Their treatment causes substantial costs due to the long healing times and extensive wound care measures. There is a paucity of information about healing times and the necessity of hospital treatment for leg ulcers of different etiologies.

Materials and methods

In this retrospective study, healing times and the frequency of in-hospital treatment of 355 patients with leg ulcers attending a wound care clinic of a university hospital were examined.

Results

The proportion of healed ulcers was 32.0?% after 3 months and 54.3?% after 6 months with an average treatment duration of 6.1 months for all ulcers. This proportion of healed ulcers was higher for venous ulcers with 45.5?% after 3 months and 63.0?% after 6 months, whereas only 30.0?% of mixed arterial-venous ulcers and 35.0?% of hypertensive ischemic leg ulcers (HYTILU) were healed after 6 months. Of the latter group, 71?% of patients were hospitalized at least once during the observation period as compared to 47?% of patients with a venous ulcer. The duration of the hospital stay was longer for mixed ulcers and HYTILU with an average of 30 days vs. 23 days for venous ulcers.

Conclusions

These data indicate that the healing times of ulcers of different etiologies differ substantially and that especially ulcers with arteriosclerosis as a causative factor have longer healing times. The fact that they require in-hospital treatment more frequently and for longer periods has significant socio-economic consequences.     

Oral doxycycline for the treatment of chronic leg ulceration

This pilot study investigated oral doxycycline as an adjunct to compression therapy for non-healing venous leg ulcers. Ten patients received doxycycline 20 mg twice daily (low-dose doxycycline) and ten patients received doxycycline 100 mg twice daily (high-dose doxycycline). Utilising a pre-test post-test study design, ulcer area was measured and wound fluid was collected before and after 4 weeks of treatment. In the high-dose doxycycline group, the reduction in median ulcer area was 48% (p = 0.1) and there was a significant reduction in wound fluid total matrix metalloprotease-1 (p = 0.02). These effects were not observed with low-dose doxycycline. There were no significant changes in wound fluid tumour necrosis factor-α or quantitative bacteriology following treatment with low-dose or high-dose doxycycline. There was no significant relationship between change in ulcer area and matrix metalloprotease-1, -8 or -9 activities in wound fluid at the end of treatment. Median wound fluid doxycycline concentrations after 4 weeks of treatment were 0.2 mg/L(0.45 lM) and 2.3 mg/L (5.18 lM) [DOSAGE ERROR CORRECTED] in the low-dose and high-dose groups, respectively, which are lower than that previously shown to inhibit matrix metalloproteases and tumour necrosis factor-α. Our study suggests that doxycycline 100 mg twice daily may improve the healing rate of recalcitrant leg ulcers, however the mechanism remains unclear.     

Allogeneic cultured keratinocytes in the treatment of leg ulcers. A pilot study.

Forty-two patients (10 males and 32 females) with 52 chronic leg ulcers were treated with sheets of cultured allogeneic keratinocytes. Sixty-five % of the ulcers healed completely and the healing rate differed between various diagnostic groups. The best results were obtained in patients with venous ulcers and wounds with mixed etiology, whereas less improvement was observed with ischaemic ulcers. Rheumatic ulcers also responded well in combination with oral corticosteroids. The overall impression was that the grafting procedure markedly enhanced wound healing.     

Epithelial tissue-type plasminogen activator expression, unlike that of urokinase, its receptor, and plasminogen activator inhibitor-1, is increased in chronic venous ulcers

BACKGROUND: The plasminogen activation system represents a potent mechanism of extracellular proteolysis and is an essential component of normal wound healing. It has also been implicated in the pathogenesis of chronic, nonhealing ulcers. Traditionally, urokinase-type plasminogen activator (uPA) has been associated with pericellular proteolytic activity involved in tissue remodelling processes, and tissue-type plasminogen activator (tPA) mainly with intravascular fibrinolysis. OBJECTIVES: The present study was conducted to characterize the spatial distribution of the various plasminogen activation system components in chronic ulcers and acute, well-granulating wounds. METHODS: The expression of uPA, tPA, urokinase receptor (uPAR), plasminogen activator inhibitor-1 (PAI-1), and vitronectin was investigated by immunohistochemical staining, in addition to uPA, tPA and PAI-1 expression by in-situ hybridization, in samples from eight chronic venous ulcers, five decubitus ulcers, five well-granulating acute wounds and five normal skin samples. RESULTS: In chronic venous leg ulcers tPA mRNA was detected in basal and suprabasal keratinocytes at the leading wound edge, while in well-granulating wounds and in decubitus ulcers tPA mRNA was expressed only in a few keratinocytes. However, tPA was widely expressed in fibroblast- and macrophage-like cells in the stroma of well-granulating wounds, while less tPA was detected in the granulation tissue of chronic ulcers. tPA mRNA and protein were localized in the superficial granular layers in normal skin. Although no qualitative differences in expression of uPA, PAI-1 or uPAR in the wound edge keratinocytes in chronic ulcers vs. normally granulating wounds were found, their expressions were more pronounced in the granulation tissue of well-granulating wounds. CONCLUSIONS: These results suggest that in poorly healing venous leg ulcers, the pattern of tPA expression is altered in keratinocytes at the leading edge of the wound, and the patterns of tPA, uPA and PAI-1 expression are altered in the granulation tissue.     

Wound teleconsultation in patients with chronic leg ulcers

BACKGROUND: The treatment of chronic leg ulcers requires frequent assessments of local wound status and adjustment of therapy. The availability of reasonably priced photographic equipment and quick electronic transfer of high-quality digital images should make it possible that the assessment of wound status can be made by remote experts. OBJECTIVE: This study examines the feasibility of using teledermatology for wound assessment and therapeutic suggestions for patients with chronic leg ulcers. METHODS: One hundred ten chronic leg ulcers of different origins were examined in face-to-face consultations. The examining doctor assessed the wound, made therapeutic recommendations and took 1-4 photographs of the wound using a digital camera. The digital images and relevant clinical information were then transmitted via a web application to an expert in wound care, who provided an independent teledermatological assessment of wound status and therapeutic recommendations. RESULTS: In our study, a high accordance between direct consultations and electronic consultations was found in the assessment of chronic leg ulcers, especially for important features like slough (concordance: 84.6%), necrosis (concordance: 98.2%) and granulation tissue formation (concordance: 76.4%). Furthermore, the teledermatologist generally felt confident in recommending further treatment strategies and in planning further wound assessments via the internet. CONCLUSIONS: Our results suggest that teledermatology offers great potential for the future in chronic wound care. By reducing the need to travel long distances to the hospital or to consult a physician with expertise in wound care, wound teleconsultation might lower health care costs and improve the quality of life for patients with chronic wounds, while still maintaining a high quality of wound care.     

Allogeneic cultured epidermal grafts heal chronic ulcers although they do not remain as proved by DNA analysis

To investigate whether allogeneic cultured keratinocytes are rejected or not, and to find out how beneficial their effect on wound healing could be, patients with chronic ulcers were grafted with allogeneic cultured human keratinocytes. In order to examine the epidermal origin of the healed wound, DNA analysis was performed and compared to donor and recipient blood-cell DNA. Healing was observed in 84% of the grafted ulcers by granulation tissue stimulation and would edge effect. In little time 60% of the grafted chronic ulcers healed completely. Although no rejection was observed, DNA analysis revealed that the grafted allogeneic keratinocytes were finally replaced by the patient's own epidermis. This study confirmed that cultured allogeneic keratinocytes that have been grafted on ulcers, play an important role in the wound healing process.     

Healing effect of ketanserin on chronic leg ulcers in patients with diabetes

BACKGROUND: The treatment of chronic leg ulcers remains a stubborn problem in many patients. Topical 2% ketanserin ointment, a 5HT2-serotoninergic blocking agent, has been reported to improve healing of decubitus, venous, diabetic and ischaemic ulcers. METHOD: The present double-blind intra-individual comparative study was performed in 12 women with diabetes presenting with at least two similar leg ulcers. In each subject, the two lesions were randomly assigned to be treated for 8 weeks by 2% ketanserin ointment or its unmedicated vehicle. OBJECTIVE: assessments of the dynamics of wound healing were performed using computerized morphometry. Evaluations were performed at 2-week intervals for 8 weeks. RESULTS: A significant decrease in relative wound area was observed on the ketanserin-treated ulcers compared with the placebo group. CONCLUSION: Topical ketanserin is a valuable therapy for difficult-to-treat leg ulcers.     

Amnia for intractable skin ulcers with recessive dystrophic epidermolysis bullosa: report of three cases

Recessive dystrophic epidermolysis bullosa (RDEB) is a disease characterized by recurrent blistering and chronic ulceration of the skin. In these patients, recurrent blisters frequently result in intractable skin ulcers due to impaired wound healing caused by mutations in the type VII collagen gene and malnutrition as well as by increased collagenase activity. To evaluate the efficacy of amnia for intractable ulcers in RDEB, we treated RDEB patients with amnia. The amniotic membrane was simply placed on the cleansed wound surface. The procedure was repeated once a week for up to 10 weeks. As a result, wound conditions improved remarkably after treatment with amnia for 2-10 weeks in all the patients, resulting in total re-epithelization of the ulcers. Amnia could be an effective therapy for intractable skin ulcers in RDEB patients, and should be considered as a re-emerging therapeutic option for the disease.     

Behandlung von Wundheilungsstörungen mit autologen Epidermisäquivalenten

A 63 year old man developed non-healing ulcerations after en-bloc resection of multiple cylindromas of the scalp. After excision and granulation-stimulating local therapy, the wound was covered with mesh grafts from the thighs. He developed widespread tension blisters with superficial ulcerations in the occipital region which did not heal despite adequate topical therapy. We then treated him successfully with EpiDex, a tissue engineered fully differentiated epidermal equivalent derived from keratinocytes of the outer root sheath of plucked anagen hair follicles. We introduce this treatment modality as a non-invasive effective option in treating non-healing ulcers.     

Phase IIa randomized,placebo‐controlled study of antimicrobial photodynamic therapy in bacterially colonized,chronic leg ulcers and diabetic foot ulcers: a new approach to antimicrobial therapy

Background With increasing problems of antibiotic resistance, photodynamic therapy (PDT) is being developed as a novel antimicrobial treatment. Following light activation, cationic photosensitizer PPA904 [3,7‐bis(N,N‐dibutylamino) phenothiazin‐5‐ium bromide] kills a broad spectrum of bacteria in vitro and this has a variety of potential clinical applications. Objectives To determine if PDT in bacterially colonized chronic leg ulcers and chronic diabetic foot ulcers can reduce bacterial load, and potentially lead to accelerated wound healing. Methods Sixteen patients with chronic leg ulcers and 16 patients with diabetic foot ulcers (each eight active treatment/eight placebo) were recruited into a blinded, randomized, placebo‐controlled, single‐treatment, Phase IIa trial. All patients had ulcer duration > 3 months, bacterially colonized with > 10⁴ colony‐forming units cm^?2. After quantitatively assessing pretreatment bacterial load via swabbing, PPA904 or placebo was applied topically to wounds for 15 min, followed immediately by 50 J cm^?2 of red light and the wound again sampled for quantitative microbiology. The wound area was measured for up to 3 months following treatment. Results Treatment was well tolerated with no reports of pain or other safety issues. In contrast to placebo, patients on active treatment showed a reduction in bacterial load immediately post‐treatment (P < 0·001). After 3 months, 50% (four of eight) of patients with actively treated chronic leg ulcer showed complete healing, compared with 12% (one of eight) of patients on placebo. Conclusions This first controlled study of PDT in chronic wounds demonstrated significant reduction in bacterial load. An apparent trend towards wound healing was observed; further study of this aspect with larger patient numbers is indicated.     

The microbiology of infected and noninfected leg ulcers

BACKGROUND: A clinical study was undertaken to investigate and compare specifically the aerobic and anaerobic microbiology of infected and noninfected leg ulcers. METHODS: Leg ulcers, defined as being infected on the basis of clinical signs, were swab sampled and investigated for aerobic and anaerobic microorganisms using stringent isolation and identification techniques. RESULTS: Two hundred and twenty isolates were cultured from 44 infected leg ulcers, in comparison with 110 isolates from 30 noninfected leg ulcers. Statistical analysis indicated a significantly greater mean number of anaerobic bacteria per infected ulcer (particularly Peptostreptococcus spp. and Prevotella spp.) in comparison with the noninfected ulcer group (2.5 vs. 1.3, respectively) (P < 0.05). Also, anaerobes represented 49% of the total microbial composition in infected leg ulcers compared with 36% in noninfected leg ulcers. The mean numbers of aerobes per wound in the two ulcer groups were not statistically different (P > 0.05). The study failed to demonstrate a clear correlation between commonly implicated facultative pathogens and wound infection. The isolation rate of Pseudomonas aeruginosa was generally low and, although Staphylococcus aureus was a frequent isolate in both wound types, it was more prevalent in noninfected leg ulcers. CONCLUSIONS: This study has demonstrated the complex aerobic-anaerobic microflora which exists in leg ulcers, the prevalence of anaerobes in infected wounds, and a poor correlation between the presence of specific aerobic pathogens and wound infection. In view of these findings, the role of microbial synergistic interactions in the pathogenesis of chronic wound infection may be of greater clinical importance than the isolated involvement of any specific potential pathogen.     

Genese des Ulcus cruris

The etiology of chronic leg ulcers is very heterogeneous. The primary aim of our investigation was to analyze the etiology of leg ulcers in patients treated in our wound care center over a period of 5 years. In this open retrospective study, data of 354 patients was analyzed. We found a venous leg ulcer in 203 (57%), and in 52 (15%) patients a combined arterio-venous induced leg ulcer. The third most common cause was leg ulcers due to different types of vasculitis in 47 (13%) patients. An exclusively arterial induced leg ulcer was diagnosed in 13 (3.7%) patients. Overall 16 different etiologies were identified. In contrast to the few comparable other investigations, we found a higher incidence of immunologic und infectious causes for leg ulcers in our patients. Even if our results do not allow a representative overview about the prevalence of leg ulcer in Germany, they demonstrate the wide spectrum of leg ulcer causes in a dermatologic wound care centre.     

Autologous suction blister grafting for chronic leg ulcers

Background Non‐healing leg ulcers represent a treatment problem. Objective Investigate grafting of autologous suction blister roofs as treatment. Methods Twenty‐nine chronic, non‐healing leg ulcers of various aetiologies in 18 inpatients were treated by autologous epidermal grafting using the roofs of suction blisters. Results 55% of ulcers completely healed 2 to 6 weeks after grafting. A 50–90% reduction in size was documented in 34% and no change was observed in 11% of ulcers. Twelve weeks after grafting, 89% of ulcers were healed completely. In most ulcers, we observed a stimulation of reepithelialization from the wound edge (‘edge effect’) and an accelerated formation of healthy granulation tissue. During a follow‐up period of 12 months, 90% of the ulcers remained healed. Conclusion Grafting of autologous suction blister roofs is an effective treatment option for non‐healing leg ulcers. The advantages of the method are its lack of pain, low costs and immediate availability.     

Superabsorbent wound dressings versus foams dressings for the management of moderate-to-highly exuding venous leg ulcers in French settings: An early stage model-based economic evaluation

Venous leg ulcers (VLUs) are the most common hard-to-heal ulcers associated with the high humanistic and economic burden. Especially clinically challenging are moderate-to-highly exuding venous leg ulcers, and current management guidelines recommend several different wound dressings for this indication.The study aimed to determine the cost-effectiveness of managing moderate-to-highly exuding venous leg ulcers with superabsorbent wound dressings versus foams dressings in French settings from Haute Autorité de Santé perspective.For evidence synthesis decision-analytic model was developed in line with international and French good modelling practices recommendations. All model inputs were informed by data identified through systematic literature reviews that, in our best knowledge, represent the best available evidence at this moment.According to the prediction from this early stage health economic evaluation, using superabsorbent wound dressings versus foam dressings mix in patients with moderate-to-highly exuding leg ulcers in French settings will result in an improved healing rate of 2.58%, incremental HRQoL of 0.143 QALWs and total direct cost savings of € 435 (USD 473) per patient over six months period.The clinical outcomes results from this evaluation are in line with management guidelines that have recommended superabsorbent wound dressings as a first-line choice in the therapy of moderate-to-highly exuding venous leg ulcers. In addition, this evaluation demonstrates that using superabsorbent wound dressings will also lead to economic savings from the Haute Autorité de Santé perspective.     

Treatment of leg ulcers with cryopreserved allogeneic cultured epithelium. A multicenter study.

BACKGROUND--In the past few years, several authors have described the usefulness of cultured allogeneic epidermal sheets in promoting wound healing of burns, leg ulcers, and donor sites. This study reports clinical results obtained by different departments in the treatment of chronic leg ulcers by cryopreserved cultured allogeneic epithelium. The freezing procedure and the assessment of viability of the cryopreserved epithelium are also described. A total of 30 ulcers were treated using 138 cryopreserved allografts. OBSERVATIONS--Twenty ulcers (66.6%) healed completely within 12 weeks. Four ulcers showed a 30% to 84.4% reduction in size by 3 weeks but did not heal completely; the remaining six ulcers did not show any improvement. A strong stimulation of granulation tissue formation and of reepithelialization from the wound edge were observed. RESULTS--The results indicate that frozen cultured epidermis, stored in a skin bank, is a valid and generally applicable alternative therapy for the treatment of chronic ulcers.     

Repeated cultured epidermal allografts in the treatment of chronic leg ulcers of various origins.

Twelve patients with 16 leg ulcers, existing for at least 3 months and not responsive to conventional inpatient therapy of at least 3 weeks, were treated with repeated applications of cultured allogenic keratinocyte sheets. A marked decrease in size was seen in all ulcers but 2. Complete closure of the ulcer was seen in 62% of the ulcers within 8 weeks. Healing was due to enhanced granulation and increased epithelialization, starting from the periphery of the wound. This edge effect suggests that the epidermal allografts act by stimulation of migration and/or multiplication of the acceptor's keratinocytes, rather than by take of the allograft.     

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.