Vitiligo is an acquired skin disease that significantly impacts the quality of life of patients. Medical treatment of vitiligo includes the use of melanocyte transplant, but the results are variable. We have treated 4 patients with either focal or generalized stable vitiligo using a graft of autologous melanocytes' culture on a denuded amniotic membrane (AM). A culture biopsy was obtained in every patient and grown in melanocytes' media for 10-14 days after which cells were transferred to a denuded AM and transplanted into the achromic lesions. Patients were followed for up to 6 months using clinical assessment of achromic lesions. Treated areas ranged between 4 cm 2 and 210.6 cm 2 . Response to treatment was excellent in all patients with 90-95% repigmentation success rate. Our results demonstrate that transplantation of autologous melanocytes cultured on AM is a new, simple, and effective treatment for stable vitiligo.

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PURPOSE. The purpose of this study was to develop a culture method for expansion of corneal endothelial cells (CEC) based on the combined activation of PI3K/Akt and Smad2.METHODS. Morphology, proliferation, and migration of cultured rabbit and nonhuman primate CEC were examined in the presence of the PI3K/Akt activators IGF-1 and heregulin beta in combination with the Smad2 activator activin A. Phenotypic characterization of CEC was performed at the RNA and protein levels. Cell pump function and transepithelial electric resistance were used for in vitro functional assessment of CEC. Finally, ex vivo-expanded rabbit CEC were transplanted into a model of endothelial damage in rabbit corneas.RESULTS. Treatment of rabbit and nonhuman primate CEC in vitro with IGF-1, heregulin beta, and activin A induced an upregulation of PI3K/Akt and Smad2 signaling pathways and an increase in proliferation and migration of CEC expressing ZO-1, connexin-43, and Na þ /K þ -ATPase. Cell pump function evaluation revealed the complete functionality of cultured CEC. Injection of rabbit CEC successfully produced recovery of normal corneal thickness in a rabbit model of endothelial dysfunction.CONCLUSIONS. We demonstrated that the combined activation of PI3K/Akt and Smad2 results in in vitro expansion of phenotypic and functional CEC. Expanded cells were able to contribute to restoration of corneal endothelium in a rabbit model. These findings may represent a new therapeutic approach for treating corneal endothelial diseases.

Vitiligo is a disfiguring disease with no definitive treatment options that significantly affects patients' quality of life. We aimed to compare, for what we believe to be the first time, the repigmentation efficacy of cultured epidermal cell suspension (CES) and amniotic membrane (AM)-cultured epidermal cell grafting (CEG) in the treatment of stable vitiligo. (December 15, 2010, to June 5, 2013, randomized, double-blind, intraindividually placebo-controlled clinical trial with a 6-month posttreatment follow-up period (last follow-up, November 26, 2012) was carried out in the dermatology department of the University Clinic of Navarra, Spain. The study was approved by the local institutional review board (Comité Ético de Investigación Clínica de Navarra, Health Department, Government of Navarra, Spain). Written informed consent was received from all patients. The participants did not receive financial compensation.Of 30 eligible patients with stable leukoderma, 24 individuals (15 women; age range, 18-57 years) were included in the final analyses. Dermatologic examination was performed on each patient to select one large vitiligo lesion (≥90 cm 2 ) or several smaller vitiligo lesions (up to 5 lesions, ≥90 cm 2 in total) per patient.Amniotic membranes were obtained during elective cesarean delivery as described. 1 Melanocyte growth medium M2 (M2; PromoCell) was used for the culture. A superficial shave biopsy (0.5 cm 2 ) was taken from pigmented buttock skin under local anesthesia. Epidermal cells were obtained (Dispase II neutral protease, grade II; Roche; and TrypLE Select enzyme; Gibco-Life Technologies) as described. 1 Cells were subcultured in two 75-cm 2 culture flasks. When 70% to 80% confluence was reached,