[Culture of human keratinocytes in serum free medium]

Keratinocytes are useful for cellular transplantation studies in order to improve functional outcome in burn patients and chronic wounds. Recently they have been used for generation of iPS cells with high efficiency. In this study, we described the details on separation, culture and proliferation of human keratinocytes from foreskin samples. In this experimental and qualitative study we obtained neonatal foreskin samples following newborn circumcision under sterile conditions. We used dispase enzyme to separate the epidermis from the dermis. Trypsin enzyme was used for isolation of keratinocyte cells from the epidermis layer. Isolated cells were cultured in type I human collagen-coated dishes and serum-free Epilife medium. We assessed morphological and immunocytochemical aspects of the isolated cells. Morphological and Immunocytochemical analyses revealed that isolated cells have typical keratinocyte morphology and they could express CK14 which is a specific marker of the keratinocytes. The cells were successfully sub-cultured at a split ratio of 1:2 every 4 to 5 days. After passage 10, a significant decrease in the proliferation of the human keratinocyte cells was observed. Morphologically the cells were flat and thin. In this study we improved the method of isolation and cultivation of human keratinocyte from foreskin. Using this method, we isolated human keratinocytes for more than 20 times. Thus, it can be concluded that this method is reproducible in other laboratories

Report of an unusual cast of perineal burn leading to anal sphincter dysfunction with fecal incontinence due to formation of an annular scar around anus

Isolated deep bum of perineal area is rare and usually occurs in cases with extensive bum. The etiology of this kind of bum may be of thermal, chemical or electrical origin. Treatment in most cases is conservative and surgical intervention rarely required. Isolated deep bum of perineal area has been reported in some special circumstances such as falling into furnace or pots containing hot fluids and sitting on exhaust pipe of vehicles; but no dysfunction of anal sphincter alter formation of perineal annular scar had been detected. Our objective is to focus on particular cases of perineal burn and its complications. A 12 years old Afghan boy with history of previous burn of perincal area at the age of 6 months, due to falling into boiled water, referred to Tohid Hospital with chief complaint of fecal incontinence. At that time the patient had been managed conservatively for his burn. As far as the patient's parents remember, he had been suffering from fecal incontinence since his childhood. In physical exam an annular scar around anus together with lack of sphincter tone were detected. By releasing the scar and use of skin graft, sphincter tone improved gradually. In special cases of isolated perineal deep burn, conservative management may lead to scarring and dysfunction of anal sphincter with fecal incontinence, imposing physical, social and psychological impacts specially on children. Proper and timely treatment is recommended to prevent this complication