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PubMedDate not supplied

Fluorescence imaging of reepithelialization from skin explant cultures on acellular dermis.

Reconstituted skin models are valuable tools in studies of cutaneous biology although they are not generally devised to visualize and quantify the time course of reepithelialization. We describe a skin explant culture technique coupled with vital microscopy allowing sequential imaging of epithelial outgrowth while maintaining the tissue in culture. Radial expansion of neo-epidermis was initiated from a 2-mm skin punch biopsy explanted onto acellular human dermis and maintained at the air-liquid interface in serum-containing keratinocyte medium. Microscopic viewings and surface area measurements of newly formed epidermis were performed repeatedly using fluorescein-based cell staining and subsequent image analysis. The labeling and visualization procedures did not interfere with neo-epidermal growth or tissue architecture. In order to appraise the versatility of the model, we studied the effect of epidermal growth factor supplementation on the course of skin resurfacing as related to tissue morphology and proliferative activity over a 10-day cultivation period. Exogenous epidermal growth factor at 10 ng/ml increased the radial outgrowth rate (mean, + 13.3 percent), papillomatosis index (+ 19.2 percent), epithelial thickness (+ 49.8 percent), and fraction of Ki-67 positive basal cells (+ 18.4 percent) in neo-epidermis produced from a biopsy of normal human skin. The contribution of this miniaturized and convenient format for concurrent studies of dynamic and qualitative features of neo-epidermis should be a useful complement to existing assays of epidermalization.

Cell Differentiation↗
PubMed2003

Early results of transcutaneous injection laryngoplasty with micronized acellular dermis versus type-I thyroplasty for glottic incompetence dysphonia due to unilateral vocal fold paralysis.

Medialization thyroplasty (type I) has become the gold standard to improve glottic closure due to unilateral vocal fold paralysis. A newer injection method utilizing homologous collagen from cadaveric human tissue has been described as an attractive alternative as no donor site is required, there is a very low risk of hypersensitivity, and the intact, acellular collagen fibers may suffer a reduced long-term reabsorption rate. Preliminary results on eight patients comparing presurgical and postsurgical parameters (perceptual, stroboscopic, acoustic, and aerodynamic) revealed comparable results when compared with a control group of individuals, age- and sex-matched, that had undergone standard medialization thyroplasty (type I). Further study is needed to assess the long-term results with this minimally invasive method of vocal fold medialization.

Adult↗
PubMedDate not supplied

Evaluation of acellular human dermis as a dermal analog in a composite skin graft.

This study evaluated the structure and function of a dermal substitute composed of de-epidermalized, acellular human dermis, and tested its performance as part of a composite skin graft with cultured human keratinocytes. The acellular dermis retained much of the complex structure of native dermis, as demonstrated by immunostaining for collagen Types IV and VII. Collagen Types IV and VII were found on the papillary surface, and collagen Type IV was found at sites throughout the acellular dermis. To further demonstrate the functionality of the acellular dermis material, we seeded the papillary surface with cultured keratinocytes. In response to the architecture of the dermal papillae, the keratinocytes formed a three-dimensional epidermal structure that was several cell layers thick, and recreated the original skin rete ridges at the interface with the dermis. When these composite grafts were transplanted to athymic mice, host fibrovascular cells repopulated the acellular dermis. Some vessels grew into the acellular dermis along the original pathways of the human blood vessels, as demonstrated by co-localization of human and mouse collagen Type IV. The skin that developed from these grafts repigmented completely via passenger melanocytes from the keratinocyte cultures, was durable, and remained stable for more than 5 months.

Animals↗
PubMedDate not supplied

A new system for the cultivation of keratinocytes on acellular human dermis with the use of fibrin glue and 3T3 feeder cells.

The growth of human keratinocytes on human acellular dermis in 4 different culture systems was compared. Epidermis was completely separated and removed from dermis after skin samples had been soaked in 0.1% trypsin at 4 degrees C for 1 week. Forty pieces of saline-washed dermis, 1 cm2 each, were randomized into 4 groups: in group A, human keratinocytes that had undergone 2 to 3 cell passages were seeded (30 x 10(4) cell/cm2) onto the dermis and sprayed with a thin layer of fibrin glue and proliferative 3T3 feeder cells that had been growing separately on the culture dish; in group B, the dermis was only sprayed with fibrin glue; in group C, the dermis was treated with 3T3 cells only; and in group D, the dermis was not sprayed with anything. The dermis samples in all groups were raised on a grid to provide an air-liquid culture system. Histology results of the composite grafts at 2 weeks were assessed as having either scanty colonies of keratinocytes (SCK), continuous stratified epithelium (CSE), or no observable keratinocyte growth. Eight out of the ten dermis samples (80%) in group A demonstrated CSE, and 30% of the samples in group B showed SCK. There were 10% SCK and 20% CSE in group C, and in group D, 30% SCK and 10% CSE were found. The good results in group A indicated that the fibrin glue facilitated the seeding efficiency of the keratinocytes on the dermis and that the vital factors released from the 3T3 feeder cells enhanced the growth and differentiation of the keratinocytes. This model provides an optimal system for the cultivation of keratinocytes on acellular dermis.

3T3 Cells↗
PubMed2003

Culture of rat keratinocytes with acellular pig dermis.

This study evaluated the growth rate and the cell activity of cultured keratinocytes on acellular pig dermis in order to develop a composite skin in vitro for burn injuries or other skin defects. Full thickness skin was cultivated from neonatal SD rats, and separated into epidermal layer and dermal layer with enzyme digestion. The keratinocytes were then seeded on the prepared acellular pig dermis soaked in the culture medium. The cultures were incubated and the growth status of keratinocytes on acellular pig dermis evaluated by phase contrast microscope, histological examination with hematoxylin-eosin staining and acridine orange staining, immunohistochemistry, observation of growth curve plotted by MTT colorimetry and analysis of changes in keratinocytes proliferation cycle with flow cytometer. Almost all keratinocytes anchored in 48-72 h, and most inosculated at days 6 and 7. The growth curve showed that the keratinocytes grew in logarithmic phase at days 3-6 after seeding. More than four layers of keratinocyte structure and the basement membrane between keratinocytes and porcine dermis were observed. Pancytokeratin was strongly positive in the cultured keratinocytes. Laminin and collagen IV were positive in the basement membrane. It is concluded that the cultured keratinocytes on acellular pig dermis grow well and the structure of composite skin which has been established is satisfactory.

Animals↗
PubMed2003

[Clinical application of acellular allogenic dermis with autogenous epithelium in wound repair].

OBJECTIVE: To observe the clinical effect of acellular allogenic dermis with split-thickness autogenous skin graft for coverage of wound. METHODS: Acellular allogenic dermis with split-thickness autogenous skin graft was used to repair 34 wounds of head, neck, trunk and extremities. The area of wounds was from 5 cm x 10 cm to 12 cm x 19 cm. Out of 34 wounds, there were 2 due to old granulation, 4 due to excision of giant pigmented nevus, 6 due to excision of capillary hemangioma of skin and 22 due to excision of scar. RESULTS: All grafts survived and had the smooth surface without obvious pigmentation and with slight wound contraction. CONCLUSION: Acellular allogenic dermis with autologous epithelium for coverage of various wounds is an ideal procedure.

Adolescent↗
PubMed2001

Experimental assessment of the revascularization of acellular human dermis for soft-tissue augmentation.

The purpose of this experimental study was to determine whether acellular human dermis was capable of complete revascularization in a subcutaneous implantation site with various placement geometries. In young adult rabbit ears, four different sheet and rolled configurations were placed and harvested after 3, 7, 14, and 28 days with silicone rubber microangiographic injections followed by histologic analysis. Revascularization of single-layer acellular human dermis occurred rapidly and was essentially complete by 14 days after surgery. No differences were observed in the ingrowth of vessels regardless of how the basement membrane was oriented. In rolled configurations, vascular ingrowth throughout the implant was slower and had not completely penetrated the grafts by 28 days after surgery at study completion. Vessel ingrowth occurred through the implant surfaces contacting the surrounding soft tissue and along the open seam of the roll. No differences were seen whether the basement membrane was oriented on either the inside or the outside of the roll. Acellular human dermis is capable of significant revascularization of its compact collagen composition in the early postoperative period. In thicker geometries, the rate and completeness of vessel ingrowth are predictably slower. Whether complete revascularization of multilayered or rolled grafts is achieved cannot be determined from this study.

Animals↗
PubMed2000

Both dermal matrix and epidermis contribute to an inhibition of wound contraction.

Contracture is a major detriment to functional recovery from large wounds. To determine the relative value of dermal replacement and epidermal coverage in inhibiting wound contraction, five full-thickness wounds (all 5 x 5 cm2) were placed on the back of 8 swine and treated in the following manner: (1) open wound, (2) porcine acellular dermis (analogous to AlloDerm for human use), (3) porcine acellular dermis with epidermal autograft placed 7 days postwounding, (4) porcine acellular dermis with immediate epidermal autograft, and (5) conventional-thickness autograft. Scar dimensions and punch biopsies were taken at days 14 and 30 postwounding. The planimetry results demonstrated that wound contraction was significantly greater with the open wounds (group 1) than all other wounds with a dermal substitute. Furthermore, wounds with initial epidermal coverage had significantly less contraction than unepithelialized wounds (14.8 +/- 1.1 cm2 at day 14 in wound group 2 vs. 20.4 +/- 0.6 cm2 in wound group 4; p < 0.05). Biopsy results revealed that wounds with initial epithelial coverage had the least amount of inflammation. These findings suggest that both dermal matrix and epidermal coverage contribute to an inhibition of wound contraction and that prompt epithelial coverage appears to impede contraction by reducing inflammation.

Animals↗
PubMedDate not supplied

Acellular allogenic dermis does not hinder initial engraftment in burn wound resurfacing and reconstruction.

Donor site morbidity, including pain and potential hypertrophic scarring, increases directly as donor site thickness increases. Autograft hypertrophy increases inversely to the thickness of autograft harvest. Both of these liabilities might be lessened by the ready availability of a functioning, "off-the-shelf" dermal substitute. The most immunogenic components of transplanted allograft skin are the cellular elements of the epidermis and dermis. If these components are removed, the remaining noncellular dermal tissue is relatively immunologically inert. We grafted 10 sites with matched control areas in six patients, with limited (< 25%) areas of the body surface appropriate for donor harvest, who required either acute resurfacing or a reconstructive procedure requiring split-thickness autografting. The six children (three girls and three boys) had an average age of 5.2 +/- 0.9 years (range, 2.8 to 10), and an average burn size of 68.7% +/- 6.7% (range, 47% to 85%). The 10 study and control sites were all parts of 10 separate procedures in these massively burned children, nine sites being reconstructive releases, and one excised acute burn. Successful epithelialization was noted at 7 days over 83% +/- 3.4% (range, 60% to 95%) at the cryopreserved acellular human dermis sites and 83.3% +/- 4.3% (range, 60% to 98%) at the control sites (NS, p = 0.96). Duration of follow-up has been 11.9 +/- 3.5 weeks (range, 2 to 29), with no difference in results by Vancouver Scar Scores. We conclude that cryopreserved acellular human dermis will engraft successfully and support engraftment of overlying thin autograft. Initial results relating to the effectiveness of cryopreserved acellular human dermis in optimizing appearance and function are encouraging, but longer follow-up is required before definitive conclusions can be made.

Burns↗
PubMed2004

[Clinical application of composite skin graft consisting of acellular allogenic dermis and autologous skin].

OBJECTIVE: To study the procedures of applying composite skin graft consisting of acellular allogenic dermis and autologous skin in the treatment of full-thickness burns and scar formation. METHODS: Fifteen burn wounds in 8 patients were treated with the composite skin graft using one-stage grafting. RESULTS: Complete wound healing was achieved in 13 wounds. The external appearance, texture and flexibility of the grafted skin were satisfactory in most cases and no obvious scar formation was observed. CONCLUSION: One-stage grafting of the composite skin graft yields a high success rate for most burn wounds with good basal layer condition.

Adolescent↗
PubMed2006

Reconstruction of parasacral hernia with acellular human dermis graft.

Parasacral hernias are defects through the pelvic floor which occur as the result of sacral resection. These defects are often large, and are frequently the result of treatment for sacral malignancies. This report documents the case of a 71-year-old woman who underwent radical coccygectomy and partial sacrectomy for a chordoma and subsequently presented 1 year later with a large parasacral hernia. The defect was repaired using an acellular human dermis graft with a gluteus maximus muscle-advancement flap overlay. This article summarizes the current literature of this challenging clinical problem, and examines the use of acellular human dermis in the repair of complex hernias.

Aged↗
PubMed2001

Tympanic membrane repair with a dermal allograft.

OBJECTIVES: To assess the use of an acellular dermal allograft in the repair of chronic tympanic membrane perforations. Chronic tympanic membrane perforations are a common problem in otolaryngology, and although surgical tympanoplasty using either temporalis fascia or rice paper has proven to be highly successful, these materials are not without their own limitations. The search has continued for a simpler, yet equally effective, means of repairing persistent tympanic membrane perforations in an office setting. In this study we experimentally evaluated the use of an acellular dermis (AlloDerm, (LifeCell Corporation, The Woodlands, TX) as an alternative to traditional tympanoplasty materials. STUDY DESIGN: Prospective study using 28 adult chinchillas. METHODS: Subtotal tympanic membrane perforations were created bilaterally in 28 adult chinchillas. Animals with noninfected, stable perforations that showed no signs of epithelial regeneration after 5 to 8 weeks were used to compare the use of rice paper patch with AlloDerm in patch tympanoplasties. RESULTS: Eighteen of 23 tympanoplasties (78%) that were performed using AlloDerm showed no signs of perforation after 5 to 6 weeks. In those performed using rice paper control, 14 of 21 (66%) showed no signs of perforation after 5 to 6 weeks. In addition, histological evaluation of the healed tympanic membranes demonstrated that the acellular dermis had been incorporated within the middle fibrous layer of the tympanic membrane. CONCLUSIONS: The results and histological studies suggest that acellular dermis may be a suitable alternative to traditional materials currently used for patch tympanoplasty. Future studies to evaluate the efficacy of acellular dermis in humans are warranted.

Animals↗
PubMedDate not supplied

Efficacy of "thick" acellular human dermis (AlloDerm) for lower eyelid reconstruction: comparison with hard palate and thin AlloDerm grafts.

OBJECTIVES: To evaluate the efficacy of thick acellular human dermis (thick AlloDerm [LifeCell Corporation, The Woodlands, Tex]) grafts for posterior and middle lamellae reconstruction to correct lower eyelid retraction and to compare the long-term efficacy of thick AlloDerm with thin AlloDerm and hard palate grafts. METHODS: Retrospective analysis of patients undergoing lower eyelid reconstruction, which encompassed subperiosteal midface lifting, middle lamellae scar lysis, and placement of lower eyelid thick AlloDerm graft. Analysis included 21 surgical procedures in 11 patients. All patients had undergone at least 1 previous lower eyelid surgery with resultant lower eyelid retraction and scleral show. Preoperative and postoperative photographs were used for analysis. Measurements of the corneal diameter and distance from pupil center to lower eyelid margin were obtained, standardized, and compared. RESULTS: Of 21 procedures, 16 (8 of 11 patients) demonstrated improvement of lower eyelid position. The mean improvement of the median marginal reflex distance was 1.6 mm (range, 0.4-2.2 mm). The average follow-up after surgery was 215 days (range, 3-12 months). Of 21 procedures (3 patients), 5 failed to demonstrate improvement of lower eyelid position, with the mean final eyelid position lower postoperatively by 0.8 mm (range, 0.4-1.4 mm). CONCLUSIONS: We demonstrated long-lasting improvement of lower eyelid position with placement of thick AlloDerm grafts during lower eyelid reconstruction. The patients in our study had undergone previous lower eyelid blepharoplasty with resultant middle lamellae tethering. Surgical correction included subperiosteal midface-lift and middle lamellae scar lysis, in addition to thick AlloDerm graft placement to the lower eyelid. The results are comparable to hard palate grafts but perhaps superior to thin AlloDerm grafts.

Blepharoplasty↗
PubMed1991

In vitro reconstitution of skin: fibroblasts facilitate keratinocyte growth and differentiation on acellular reticular dermis.

Extensive full-thickness burns require replacement of both epidermis and dermis. We have described a method in which allogeneic dermis from engrafted cryopreserved cadaver skin was combined with cultured autologous keratinocytes. In the present study we combined human keratinocytes and fibroblasts, and acellular human dermis in vitro and transplanted this "reconstituted skin" into athymic mice. Both human papillary dermis in which the basement membrane zone has been retained and human reticular dermis that has been repopulated with human dermal fibroblasts are good substrates for keratinocyte attachment, stratification, growth, and differentiation. Both of these dermal preparations can be lyophilized and stored at room temperature without losing their ability to support keratinocyte growth. In contrast, human papillary dermis that has been treated with trypsin lacks laminin and collagen type IV in the BMZ and supports keratinocyte attachment and differentiation less well.

Animals↗
PubMed2006

Fibroblasts accelerate culturing of mucosal substitutes.

Reconstruction of large mucosal defects of the floor of the mouth is typically performed with keratinizing skin. Drawbacks include donor site defects and hair bearing of the flaps. Cultured mucosal substitutes (CMSs) have been developed for clinical use to replace keratinizing skin. Acellular dermis is often used as a dermal carrier for autologous cells, because it reduces wound contraction and is easier for the surgeon to handle than, for example, collagen gels. A major problem of CMSs using acellular dermis is variation in epidermal quality. To improve the quality of the CMSs, human fibroblasts were incorporated into the acellular dermis and seeded with human keratinocytes. To study the role of the fibroblasts in epidermal morphology and basement membrane formation, CMSs were stained for differentiation markers beta1 integrin, cytokeratin 10, and involucrin after 1 and 2 weeks in culture. Basement membrane formation was analyzed using laminin 5 and collagen IV and VII staining; proliferation was analyzed using Ki-67 staining. The epidermises of fibroblast-containing CMSs matured faster into a well-organized epithelium than did those that did not contain CMSs. A 52.7% increase in basal cells, a 53.5% increase in mitosis index, and a 78.0% increase in keratinocyte cell layers were observed. Addition of fibroblasts reduced culturing time and enhanced proliferation, maturation, and quality of the epidermis.

Cells, Cultured↗
PubMed2005

Utility of acellular allograft dermis in the care of elderly burn patients.

BACKGROUND: Because skin thins with advancing age, traditional thickness skin grafts cannot always be obtained in very elderly burn patients without creating a new full-thickness wound at the skin graft donor site. MATERIALS AND METHODS: In an attempt to circumvent this problem, acellular allograft dermis (Alloderm, Life Cell Corp., The Woodlands, TX) and thin autograft (depth 0.005 inches) was used in skin grafting 10 elderly burn patients (age 78 year +/- 2, TBSA burn 17% +/- 2; mean +/- SEM) over a 1-year period. The outcome of patients receiving Alloderm was compared retrospectively to a similar group of 18 elderly patients admitted over the prior year, eight of whom underwent operative wound excision and autografting (depth 0.014 inches) without Alloderm. RESULTS: Length of hospital stay was significantly reduced in patients treated with Alloderm compared to the total group of elderly in whom selective use of operative debridement and skin grafting was used. Functional outcome was improved in those patients who underwent skin grafting regardless of operative technique. Donor site healing time was significantly reduced with Alloderm (12 days +/- 1 versus 18 days +/- 2), while graft take was similar to conventional autografting. Unfortunately, 3-month mortality remained poor regardless of operative skin grafting or technique used. CONCLUSIONS: This initial experience suggests that use of Alloderm may allow more elderly burn patients to undergo operative wound closure, thus improving functional outcome and reducing hospitalization. Unfortunately, long-term survival for very elderly burn patients remains poor.

Aged↗