Gene Transfer into Cultured Human Epidermis and its Transplantation onto Immunodeficient Mice: An Experimental Model for Somatic Gene Therapy

Birgit Sehested Hansen, Jens Fogh, Uffe Birk Jensen, Thomas Gryesten Jensen, Peter K A Jensen, Jørgen Rygaard, Steen Kølvraa, Lars Bolund
1994 Journal of Investigative Dermatology  
To try epidermis as a target for somatic gene therapy we studied transfected primary human keratinocytes grovvn in culture and grafted onto athymic mice. We have developed a novel technique for grafting cultured epidermal sheets onto mice. First, the graft is placed on the dorsal muscle fascia underneath the mouse skin using the latter as a bandage. Secondly, the mouse skin above the graft is removed, which exposes the grafted skin to open air and thus stimulates terminal difFerentiation. A
more » ... l method for the discrimination between murine and human epidermal cells is also presented, employing in situ hybridization with hUJDan Alu repeated DNA sequences. During monolayer culture the keratinocytes were lipof"ected with the gene for human growth hormone in an Epstein-Barr virus-based expression vector. The cells were allowed to develop a multilayered tissue for T he transplantation of cultured keratinocytes onto athymic animals has numerous applications. It may be used to optimize the grafting techniques of severe burns [1], or to study events occurring during the terminal differentiation of normal or pathologic epidermis [2, 3] . It may also be used in the context of gene therapy either to produce an organoid deliveri~g or :emoving specific products systemically [4] [5] [6] [7] or as a geneticall y Improved graft that locally can deliver products like growth factors that could stimulate wound healing and the formation of a more developed dermis. Fenjves et at [4] showed that human apolipoprotein E (Apo E) was secreted into the blood of nude mice after the transplantation of cultured human primary keratinocytes. Using a carcinoma cell line transfected with the gene encoding human growth hormone (hGH), Teumer et at [5] showed it possible to detect this exogenous protein after the transplantation of the c~rcinoma cell sheet upside down. under the mou~e skin . The coagulation factor IX has also been translently detected m the blood of nude mice after the grafting of retrovirally transduced h uman primary keratinocytes [6] . In the present study, we have transfected primary keratinocytes Manuscript
doi:10.1111/1523-1747.ep12395402 pmid:8077706 fatcat:vnw2rsptijaxhhxf55i4femkli