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• W2103075603 abstract "We describe a 17-year-old Taiwanese boy in whom vitiligo of the right leg developed when he was 8 years old. The vitiligo stopped spreading 6 years later. He had received medical treatments without significant improvement. In September 1997, he received an initial transplantation of autologous cultured melanocytes to a 50-cm2 depigmented area as a trial. After successful pigmentation was observed in that area, he received 4 similar treatments to other vitiliginous skin areas of various sizes (each area was 70 to 90 cm2, with a total area of 360 cm2). After 5 transplantations, approximately 95% of treated vitiliginous skin was repigmented. Two suction blisters (each 1.80 cm2 in area) were raised on normal skin of the abdominal area. The roofs of the blisters were excised for cell culture. The specimens were incubated in 0.25% trypsin solution for 15 minutes at 37°C, followed by treatment with 0.02% EDTA solution for 10 minutes. The epidermal sheets were gently manipulated with forceps to dissociate the epidermal cells and to yield a cell suspension. The cells were seeded into a flask with a modified melanocyte medium consisting of Ham's F12 nutrient mixture supplemented with 50 μg/mL of gentamicin, 20 ng/mL of recombinant human basic fibroblast growth factor (bFGF), 20 μg/mL of isobutylmethylxanthine, 10 ng/mL of cholera toxin, and 20% fetal calf serum. The flask was incubated at 37°C in an atmosphere of 95% humidified air and 5% carbon dioxide (CO2). Geneticin was added (100 μg/mL) for 3 days to eliminate keratinocytes and fibroblasts. After reaching confluence (mean interval between seeding and confluence was 2 weeks), the melanocytes were detached by means of trypsin-EDTA solution, centrifuged, diluted, and seeded for subculture. After 17 to 22 days of 1 to 2 subcultures, the number of melanocytes had increased 50- to 100-fold, with a doubling time of 2.60 to 3.68 days. At the time of transplantation, the melanocytes were detached by means of trypsin-EDTA solution, centrifuged, and resuspended with F12 medium. The epidermis at the recipient site was removed by superficial ablation using a SilkTouch Flashscanner attached to a Sharplan 1030 CO2 laser at the setting of 6 W with a 0.2-second pulse duration. The melanocyte suspension was applied to the denuded areas with a pipette at a density of 7 × 104 to 1 × 105 cells per square centimeter. The number of transplanted melanocytes was 4.5 × 106 in the first transplantation, and ranged from 7 × 106 to 8 × 106 in each of the following 4 transplantations. The transplant was secured with Trex nonadherent silicone-treated gauze covered with gauze moistened with medium, followed by Tegaderm dressing. After 8 to 10 days, all dressings were removed. The patient was then encouraged to expose the treated areas to natural sunlight. Clinical repigmentation gradually took place within 4 weeks after each treatment. A slight hyperpigmentation presented initially, but then faded gradually. After 6 months, the treated areas had acquired a color similar to the surrounding skin, and this appearance remained unchanged at 10 months' follow-up. The repigmented portion amounted to 95% of the total treated areas. Treatment of vitiligo with epidermal grafting or the transplantation of cultured melanocytes has been previously reported. Both methods obtained good results; the transplantation of cultured melanocytes produced superior, often cosmetically perfect results.1Falabella R Repigmentation of leukoderma by autologous epidermal grafting.J Dermatol Surg Oncol. 1984; 10: 136-144Crossref PubMed Scopus (55) Google Scholar, 2Olsson MJ Juhlin L Transplantation of melanocytes in vitiligo.Br J Dermatol. 1995; 132: 587-591Crossref PubMed Scopus (97) Google Scholar Repigmentation has remained stable for several years in most cases. We use the CO2 laser to rapidly create an uncomplicated superficial epidermal ablation, which appears to be a fertile growth surface for transplanted melanocytes. CO2 laser abrasion can create a more homogeneous dermabrasion than traditional dermabrasion procedures even in an uneven skin surface. The culture medium used widely for cultivation of epidermal melanocytes is TIC medium, which consists of 12-o-tetradecanoyl-phorbol-13-acetate (TPA), isobutylmethylxanthine, and cholera toxin.3Halaban R Ghosh S Duray P Kirkwood JM Lerner AB Human melanocytes cultured from nevi and melanomas.J Invest Dermatol. 1986; 87: 95-101Abstract Full Text PDF PubMed Scopus (131) Google Scholar TPA is a tumor promoter and should be avoided. We have studied the growth regulation of melanocytes in vitro and found that basic fibroblast growth factor can be a substitute for TPA. It is a safe natural substance and causes a significantly greater increase in cell growth than TPA.4Hu DN McCormick SA Ritch R Studies of human uveal melanocytes in vitro: growth regulation of cultured human uveal melanocytes.Invest Ophthalmol Vis Sci. 1993; 34: 2220-2227PubMed Google Scholar Many melanocytes can be obtained with the use of our modified culture medium, enabling transplantation to larger areas than are possible with epidermal grafting methods. The advantages of autologous cultured melanocyte suspensions include ease of preparation, ability to perform controlled application, coverage of large depigmented lesions, and production of a homogeneous skin color. Although the mechanical force used to produce a suction blister may cause damage to epidermal cells, the time of enzymatic exposure can be shortened, thereby decreasing the trauma produced by prolonged exposure to enzymes in shaved skin methods. Other advantages of suction blister sampling include the avoidance of local anesthesia and less scarring or pigmentation change at the donor area. Although the culture of melanocytes is time-consuming, the encouraging results obtained indicate that the procedure can be a valuable treatment for large segmental vitiligo." @default.
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• W2103075603 date "2001-03-01" @default.
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• W2103075603 title "Transplantation of autologous cultured melanocytes for treatment of large segmental vitiligo" @default.
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