Treatment of full skin thickness burn injury using cultured epithelial grafts

This report presents the experience gained from 26 patients treated with autogenic cultured epithelial grafts (auto-CEG). All auto-CEG were applied to wounds clinically defined as full skin thickness injury. In total 89 separate sites were grafted. The overall estimate of ‘take’ ranged from 0 to 98 per cent with a mean value of 15 per cent. The highest level of ‘take’ (43 per cent) was observed when auto-CEG were applied to wounds which had been previously covered with allogenic split-thickness skin grafts. An increased incidence of wound colonization with pathogenic species of bacteria corresponded with a decreased graft ‘take’. Ps. aeruginosa and Staph. aureus were found to be present on 32.6 per cent and 60.5 per cent of wound swabs respectively, where 10 per cent or less ‘take’ of auto-CEG was seen, indicating that bacterial infection is in part responsible for graft failure. However, in 20.9 per cent of such instances, no growth of bacteria was detected, perhaps suggesting that certain wound beds may not present the correct physical environment necessary to support proliferating epithelial cells isolated from their underlying dermal component.     

A policy of early excision and grafting in elderly burn patients shortens the hospital stay and improves survival

Early excision and grafting of the burn wound appears to shorten the hospital stay and decrease mortality in children and adults. However, whether an early surgical approach is safe in elderly burn patients has not been resolved. To answer this question we carried out a prospective study of early surgery in 114 consecutive patients over the age of 50 years. Patients were generally operated on between post-burn days 2 and 5. The mean age of the patients was 68 years, with a burn size of 22 per cent, of which 13 per cent was full thickness skin loss. The mean hospital stay of the surviving patients was reduced by 40 per cent compared to national averages (P less than 0.001). The mortality rate for the entire group of patients was 17 per cent, with 2 deaths in the 65 patients with burns less than 20 per cent total body surface area (TBSA). Although the mortality rate for patients with burns greater than 20 per cent TBSA was 35 per cent, this was less than predicted (P less than 0.05). The improvement in survival appeared to be due to a decrease in the incidence of lethal burn wound infections.     

EFFECTS OF PROLONGED NITROUS OXIDE ON WOUND HEALING

The effect of long-term nitrous oxide treatment on wound healingwas assessed in rats using wound tensiometry, to assess woundstrength in surgical incised wounds, and estimation of the rateof healing of a small area of full thickness skin loss. No effecton wound strength could be detected in animals treated for threedays with 25 or 60 per cent nitrous oxide. Similarly, no significantdifference was present in the rate of healing of the area offull thickness skin loss after two days treatment with nitrousoxide.     

Successful homografting in an elderly patient with extensive burns using his identical twin brother as skin donor

A 60-year-old patient with 50 per cent body surface area (BSA) flame burns (of which 46 per cent were full skin thickness loss) was treated successfully with skin grafts taken from his identical twin brother to cover 20 per cent of the wounds. On theoretical grounds, it can be assumed that the period in hospital was reduced by 28 days. Blood for tissue and blood group typing should be taken at an early stage, before the patient is given a blood transfusion.     

Use of cultured epidermal autografts in the treatment of large burns

Mortality in patients with large areas of full skin thickness burns is, in part, due to complications developing during the period of prolonged delay required to obtain enough wound healing to permit skin grafting from limited donor sites. Cultured epithelial autograft (CEA) has become available as an alternative measure to the use of expanded skin autografts and regrafting. Small biopsies are taken and transported to the laboratories of BioSurface Technology where keratinocytes are grown to cover large areas during a 3-week period. The cultured keratinocytes are then available on petroleum jelly gauze which is applied to the patient. The gauze is used as a temporary dressing. To date, 37 patients have been biopsied. Grafts have been applied in 15. Graft ‘take’ averaged 71.5 per cent at our institution. Two of the patients grafted with CEA died of sepsis. One patient had a 100 per cent loss of the CEA grafts. In 12 patients, the use of CEA probably contributed significantly to wound coverage and survival. Such grafts are more susceptible to mechanical loss than routine autograft, although long-term coverage after several years is considered to be satisfactory. The cost of the process is high.     

Preliminary clinical studies of a biological skin equivalent in burned patients

The possibility of covering large areas of full thickness skin loss with 'living skin equivalent' produced by a modification of Bell's method was studied. Living skin equivalents, composed of a dermal equivalent (fibroblasts plus collagen) covered by epithelial cells were grafted, meshed or non-meshed, onto granulation tissue and, in one patient, onto fascia. Eight patients with full skin thickness burn wounds covering over 15 per cent of the body surface area were thus partially covered. The graft 'take' was evaluated every 48 h. In every patient grafted, an extensive lysis (60-90 per cent) of the skin equivalent graft was observed at the first dressing (48 h). In one patient only, a significant percentage of 'take' (40 per cent) was observed 14 days after grafting. These disappointing results were probably related to the presence of collagenases or proteases produced on the wound bed either by bacteria or by surrounding human cells. It appears that at the present time the biochemical nature of the dermal equivalent used is not yet completely appropriate to serve routinely as a substitute for human skin.     

Micrografting in the treatment of severely burned patients

The micrografting technique, employed to treat patients with extensive TBSA burns and reduced areas of healthy donor skin, was assessed simultaneously with traditional mesh grafts (STSG) or Tiersch grafts in the treatment five severely burned subjects (average TBSA burned 35.6 per cent; average per cent of full thickness wounds 32.6 per cent).

At the first clinical control on day 6 post-surgery 93 per cent of the micrografted area was in situ and healthy: epithelialization of the wound sites was complete at day 21. Meshed STSG yielded 90 per cent attachment at day 6 post-surgery, with epithelialization complete at 12 ± 2 days post-surgery.

Functional results were equivalent no matter what technique was used, while aesthetic results were better in the areas where micrografting was carried out.

Although the micrograft technique is labour-intensive, if the expansion needed is at least 1:6, the aesthetic and functional results obtained are comparable to, or better than, those with meshed grafts. Also, large segments of micrograft are not compromised if a small area of mesh becomes detached, and epithelialization is faster and more uniform, enabling a reduction in both infection and length of hospital stay.     

Ultrastructural and surface-area changes of acute and traditionally expanded full thickness skin grafts.

The purpose of this study was to determine the effect of expansion on the survival of full thickness skin grafts. In eight pigs 300 ml rectangular tissue expanders were placed into subcutaneous pockets. In four pigs (group I), the expanders were inflated rapidly with a mean 200 ml saline. In the other four pigs (group II), a mean total of 300 ml saline was injected in weekly increments over eight weeks. At the completion of expansion, full thickness skin grafts were taken from the expanded area and sutured back to the donor defects. In both groups surviving graft areas were similar to controls (p greater than 0.06). This study showed that acute and traditional expansion does not lead to increased loss of full thickness skin grafts of the expanded skin.     

自体皮与异体皮混合移植中的层粘蛋白表达

目的　通过对大张异体皮打洞嵌植小块自体皮混合移植皮片中层粘蛋白 (L N )的表达研究 ,探讨自体皮与异体皮混合移植促进 度烫伤创面愈合的机制。方法　清洁级 SD大鼠背部脱毛后 ,造成占体表总面积10 %～ 15 %的 度烫伤。伤后 3天切痂 ,移植大张打洞的异体皮。异体皮移植后第 3天 ,嵌植小块自体皮 ,完成皮肤混合移植。在异体皮移植后 3、5、7、14和 2 1天 ,采用免疫组织化学方法测定移植皮片中 L N的表达。结果　异体皮移植后第 7天出现排异反应 ,表现为异体表皮逐渐被爬行而来的自体表皮所铲除替代 ,异体真皮表面仍可见 L N阳性表达。异体皮移植后第 14天 ,从自体皮岛迁移来的表皮细胞下有零星 L N表达 ,异体皮中的 L N仍基本保持完整。异体皮移植后第 2 1天 ,自体表皮已基本覆盖异体皮区 ,L N表达已基本完整。结论　大张异体皮打洞嵌植小块自体皮混合移植中的异体皮为创面愈合提供了基底膜成分 ,有助于改善创面愈合的外观和功能     

Fish skin grafts compared to human amnion/chorion membrane allografts: A double‐blind,prospective, randomized clinical trial of acute wound healing

Chronic, nonhealing wounds consume a great deal of healthcare resources and are a major public health problem, associated with high morbidity and significant economic costs. Skin grafts are commonly used to facilitate wound closure. The grafts can come from the patient's own skin (autograft), a human donor (allograft), or from a different species (xenograft). A fish skin xenograft from cold‐water fish (Atlantic cod, Gadus morhua) is a relatively recent option that shows promising preclinical and clinical results in wound healing. Chronic wounds vary greatly in etiology and nature, requiring large cohorts for effective comparison between therapeutic alternatives. In this study, we attempted to imitate the status of a freshly debrided chronic wound by creating acute full‐thickness wounds, 4 mm in diameter, on healthy volunteers to compare two materials frequently used to treat chronic wounds: fish skin and dHACM. The purpose is to give an indication of the efficacy of the two therapeutic alternatives in the treatment of chronic wounds in a simple, standardized, randomized, controlled, double‐blind study. All volunteers were given two identical punch biopsy wounds, one of which was treated with a fish skin graft and the other with dehydrated human amnion/chorion membrane allograft (dHACM). In the study, 170 wounds were treated (85 wounds per group). The primary endpoint was defined as time to heal (full epithelialization) by blinded assessment at days 14, 18, 21, 25, and 28. The superiority hypothesis was that the fish skin grafts would heal the wounds faster than the dHACM. To evaluate the superiority hypothesis, a mixed Cox proportional hazard model was used. Wounds treated with fish skin healed significantly faster (hazard ratio 2.37; 95% confidence interval: (1.75–3.22; p = 0.0014) compared with wounds treated with dHACM. The results show that acute biopsy wounds treated with fish skin grafts heal faster than wounds treated with dHACM.     

In vivo expansion and regeneration of full‐thickness functional skin with an autologous homologous skin construct: Clinical proof of concept for chronic wound healing

A new cell‐tissue technology uses a patient's skin to create an in vivo expanding and self‐organising full‐thickness skin autograft derived from potent cutaneous appendages. This autologous homologous skin construct (AHSC) is manufactured from a small full‐thickness skin harvest obtained from an uninjured area of the patient. All the harvested tissue is incorporated into the AHSC including the endogenous regenerative cellular populations responsible for skin maintenance and repair, which are activated during the manufacturing process. Without any exogenous supplementation or culturing, the AHSC is swiftly returned to the patient's wound bed, where it expands and closes the defect from the inside out with full‐thickness fully functional skin. AHSC was applied to a greater than two‐year old large (200 cm²) chronic wound refractory to multiple failed split‐thickness skin grafts. Complete epithelial coverage was achieved in 8 weeks, and complete wound coverage with full‐thickness functional skin occurred in 12 weeks. At 6‐month follow‐up, the wound remained covered with full‐thickness skin, grossly equivalent to surrounding native skin qualitatively and quantitatively equivalent across multiple functions and characteristics, including sensation, hair follicle morphology, bio‐impedance and composition, pigment regeneration, and gland production.     

依创面定形厚断层皮片在全手背瘢痕整复中的应用

目的:研究皮肤移植重建手背的理想整形方法。方法:选择38例41只手背瘢痕。在局部肿胀麻醉下全层切除手背瘢痕并作关节畸形复位。用布片制作与创面相同形状和大小的取皮模板,以此制作定形厚断层皮片,植至手背创面。结果:41个定形皮片中36个皮片完全成活,术后6￣28个月随访28例31只手,手外形及功能良好,皮片色泽接近正常皮肤,无环形瘢痕形成。5个皮片有点状皮片坏死,后期随访结果也较好。所有病例未发生明显的环形瘢痕。结论:定形厚断层皮片移植是手背瘢痕整形的理想方法。     

032 Treatment of Chronic Ulcer with Allogeneic Cultured Dermal Substitute (CDS) composed of Collagen Sponge with Fibroblasts

We have been cultivating human epidermal cells for therapeutic purpose according to the original methods developed by Rheinwald and Green. Cultured epithelium (CE) was applied to patients with severe skin defects, burn wounds, chronic skin ulcers and cutaneous disorders like hypomelanosis. Autologous CE allows to restore massive skin surface in a short period compared with other conventional treatments. For grafts take, it is important to manage wound beds properly prior to CE grafting. The CDS was applied to prepare wound bed acceptable for CE grafting. The CDS was designed to secrete various types of cytokines, i.e., VEGF and KGF to stimulate wound healing. The successful management of deep wounds like chronic skin ulcer or burn ulcer requires granulation tissue formation and epithelialization to wound closure. This study aimed to evaluate the application of CDS in conjunction with CE for patients with chronic skin ulcer and burn ulcer. In some cases the wounds were cured by using CDS, and followed by CE grafting. All clinical trials achieved excellent or good results, showing no contracture and no hypertrophic scar after wound closure. The CDS was found to be useful to prepare wound beds and to facilitate wound management.     

Release of burn scar contractures of the neck in paediatric patients

Burn scar contractures of the neck represent a continuing problem for plastic surgeons. A review of 143 neck-release procedures performed at the Cincinnati Shriners Burns Institute documented a high rate of contracture recurrence. This was especially true in those patients who had previously suffered burns to the entire anterior neck. Treatment of this condition with Z-plasties, or releasing incisions with split thickness skin grafts, had a recurrence rate of 81 per cent and 62 per cent, respectively. The use of a neck hyper-extension brace for over 1 year following skin grafting decreased the recurrence rate to 17 per cent. In patients who were not compliant in wearing a brace, the best result was obtained by using a full thickness skin graft in the release site. Contractures resulting from smaller burns were successfully handled by a variety of techniques.     

Use of cultured human epidermal keratinocytes for allografting burns and conditions for temporary banking of the cultured allografts

Five children who suffered burns clinically regarded as full skin thickness loss were grafted with cultured allogeneic skin from newborn prepuce. The wounds had remained open and infected without healing for about 20 days before the patients were received in the burn unit. To avoid losing surviving deep epidermal cells the wounds were débrided but not deeply excised and, a few days before allografting, they were washed with isodine solution and sterile water, and treated with silvadene cream application. All children received 76 cultured allografts of about 60 cm2 each. After allografting, the wounds were epithelized in 7-10 days and the allogeneic grafted skin began desquamation suggesting that the allograft did not 'take' permanently but was replaced by the newly formed skin. On the other hand, since allografting is an adequate therapy to provide early temporary coverage in extensively burned patients, we developed conditions for banking cultured skin to make it available for immediate use. The conditions described allow banking of the cultured grafts for 15-20 days with retention of clonal growth ability similar to that of unstored epithelia. The results show that cultured epidermal cells obtained from human newborn foreskin, when used as allografts for coverage of full skin or deep partial skin thickness burns, allow rapid epithelization of the burn wounds.     

In vitro skin expansion: Wound healing assessment

For treatments requiring split‐thickness skin grafts, it is preferable to mesh the grafts. This reduces the amount of excised skin and covers more wound area. The mesh technique, however, destroys surface continuity, which results in scarring. Strain‐based bioreactors, on the other hand, have successfully expanded split‐thickness skin grafts in vitro within a 7‐day period, increasing graft coverage. After in vitro expansion, the expanded skin grafts were tested in a porcine full‐thickness excisional wound model. Expanded graft take rate was 100%. Volumetric, histologic, and mechanical assessments indicated that expanded grafts were comparable to unexpanded grafts (positive control). While there was considerable variation in expansion (31% to ?3.1%), this technique has the potential to enhance the coverage area of skin grafts while reducing or eliminating scarring.     

嵌合体大鼠诱导异种皮肤移植免疫耐受的初步研究

目的 探讨嵌合体大鼠诱导异种皮肤移植免疫耐受的可行性。方法 采集兔骨髓干细胞，经行宫内胎鼠移植以及对新生子鼠行肝内注射，制作兔骨髓干细胞嵌合体大鼠模型。6周后将15只嵌合体大鼠分为A组（8只）、B组（7只）。将A组大鼠皮肤移植给供髓兔，将7只非嵌合体大鼠皮肤移植给非供髓兔作A组对照；将B组大鼠、7只非嵌合体大鼠（B组对照）皮肤同时移植给供髓兔。记录移植后皮片成活时间、创面愈合时间。结果A组对照移植皮片成活（9．3±1．8）d，创面于（20．9±2．1）d愈合；A组移植皮片成活（15．1±2．6）d，创面于（18．5±1．3）d愈合。B组及其对照移植皮片的成活时间、创面愈合时间与A组相似。与各自对照皮肤移植相比，A、B组皮片成活时间延长（P〈0．01），创面愈合时间缩短（P〈0．05或P〈0．01）。结论 嵌合体大鼠行异种皮肤移植后能诱导移植皮片免疫耐受．使其成活时间明显延长．创面愈合时间缩短。     

A comparison of calcium alginate and scarlet red dressings in the healing of split thickness skin graft donor sites

Forty-six patients had split thickness skin grafts harvested from the upper inner thigh. Calcium alginate (Kaltostat) and scarlet red dressings were applied to each half of the wound. Dressings were changed after 10 days and healing of the donor site was assessed. Seventy-two per cent of wounds dressed with calcium alginate and 84% of wounds dressed with scarlet red were healed at 10 days. Scarlet red was shown to be significantly better than Kaltostat in the healing of split thickness skin graft donor sites when assessed at 10 days (p less than 0.04).     

The role of sampling in the detection of microbial contamination on cadaveric allograft skin used as a biological wound dressing

The availability of cryopreservation and low temperature storage techniques for cadaveric allograft skin allows it to be preserved while microbial assessments are made before its use as a temporary biological dressing on burn wounds. In a 300-donor, 5-year prospective study, we tested ten skin samples from defined areas on each donor for microbiological contamination. Although the skin from 52.3 per cent of the donors possessed some detectable residual microbial contamination after surgical body preparation and skin removal, such contamination was limited to an average of 1.4 areas per body, leaving 86 per cent of all skin obtained free from detectable contamination and suitable for use as biological wound dressings. The number of skin samples tested per donor body determined the accuracy of detection of the presence of contamination. Testing one skin sample per donor body yielded a correct skin assessment 92 per cent of the time, while testing five skin samples increased the accuracy to 96 per cent, and testing ten skin samples yielded a 99.9 per cent accuracy in detection of skin contamination. Thus, it is within the ability of a skin bank to set the limits of microbiological risk to patients receiving processed cadaveric allograft skin.     

Rejection, after a slightly prolonged survival time, of Langerhans cell-free allogeneic cultured epidermis used for wound coverage in humans

Autologous cultured epidermis (CE) grown from small skin biopsies in vitro has been successfully applied for wound grafting in humans. Since it has been reported recently that allogeneic CE might be tolerated as permanent wound cover, we investigated the properties of CE and its use as autologous and allogeneic grafts. Except for some differences, such as the absence of Langerhans cells and the lack of a stratum corneum, CE resembled its natural analogue. Autologous CE applied for grafting of leg ulcers and various surgical skin defects adhered firmly and permanently to the wound bed within 2 weeks, became regularly stratified, and formed a stratum corneum. Langerhans cells gradually entered the grafts; the dermis contained no inflammatory infiltrate. Allogeneic CE unmatched for MHC and blood group antigens used to partially cover tangentially excised third-degree burns, donor sites of split-thickness skin, and a defect after tumor excision initially survived well like the autografts. However, they were completely rejected after 10-22 (mean, 14.5) days, which is 4-5 days later than reported for split-thickness skin allografts. Clinically, rejection presented as "melting" of the graft. (Immuno)histologically, we found a dense mononuclear dermal infiltrate consisting predominantly of activated T cells, vacuolization, and single-cell necrosis of keratinocytes, as well as HLA-DR expression on keratinocytes, and finally separation and lysis of the epidermis. Limiting dilution analysis in 2 out of 4 allograft recipients revealed a considerable increase of circulating donor-specific cytotoxic T cell precursors during graft rejection. We conclude that grafting of allogeneic CE does not lead to permanent but to slightly prolonged graft survival.