Maintenance of human skin in organ culture: role for insulin-like growth factor-1 receptor and epidermal growth factor receptor

Recent studies have shown that adult skin incubated in low-Ca2+ (0.15 mM) m edium rapidly degenerates but that normal architecture is maintained when t he tissue is incubated in high-Ca2+ medium (1.4 mM Ca2+). To investigate wh ether the skin cell-produced growth factors insulin-like growth factor-1 (I GF-1) and epidermal growth factor (EGF) play a role in these events, 2-mm s kin punch biopsies were obtained and maintained for 8 to 10 days in a basal medium containing 0.15 mM Ca2+ with and without growth factors, or contain ing 1.4 mM Ca2+ with and without antibodies to the same growth factors. In parallel experiments, cultured human keratinocytes were incubated for 2 day s in the same basal medium in the presence or absence of the same growth fa ctors and antibodies. Consistent with previous reports, organ cultures incu bated in the low-Ca2+ (0.15 mM) medium rapidly degenerated. Neither IGF-1 n or EGF prevented the complete degeneration of epidermis and dermis in these organ cultures. Interestingly, the addition of an anti-IGF-1 receptor (IGP -1R) antibody to the organ cultures maintained in high-Ca2+ medium induced changes reminiscent of those seen when the organ cultures were maintained i n low-Ca2+ medium, i.e. tissue degeneration. In contrast, antibodies to EGF receptor, used for comparison, only produced focal areas of epidermal necr osis. In vitro, IGF-1 is a known mitogen for keratinocytes. In cultured hum an keratinocytes, anti-IGF-1R antibody partially inhibited the IGF-1-mediat ed stimulation of human keratinocyte proliferation without affecting normal spontaneous growth. Additionally, IGF-1R immunolocalized to basal keratino cytes in vivo, exhibited specific binding to IGF-1 in vitro. This indicated a critical role for IGF-1R in both organ cultures ex vivo and cultured cel ls in vitro. Messenger RNA encoding both IGF-1 and IGF-1R were readily dete cted by RT-PCR in organ cultures incubated in both low- and high-Ca2+ mediu m. There were no detectable differences in IGF-1 mRNA in organ cultures gro wing in the low- or high-Ca2+ medium, but lower levels of IGF-1R mRNA were observed in the organ cultures maintained in low-Ca2+ medium than in those in high Ca(2+)medium. These findings are consistent with homeostatic change s in the tissue grown under different calcium concentrations. IGF-1 mRNA wa s detected in several skin cell populations in vitro, even though it was un detectable in cultured keratinocytes. Taken together these findings indicat e that (1) the IGF-1/IGF-1R loop is critically involved in maintenance of h uman skin organ cultures ex vivo, and (2) IGF-1, locally produced by skin c ells other than keratinocytes, interacts with its receptor, predominantly e xpressed in basal keratinocytes, to maintain tissue homeostasis.