Psoriasis is characterized by a marked T cell dependent hyperproliferation of the keratinocyte population within lesions of afflicted individuals. We have recently identified a clonogenic population of progenitor keratinocytes which, in normal skin, rarely undergo transition from the G0 stage of the cell cycle into the G1 cell cycle stage, however, in psoriatic lesions these progenitor cells are all actively cycling in G1 /S/G2/M. Relative to normals, clonogenic G0 keratinocytes hyper-respond to lymphokines from lesional T cell clones, in an ex vivo model of synchronized cell cycle induction from G0, particularly if stimulated with fibronectin (Fn). Changes in the clonogenic keratinocyte interactions with dermal/epidermal junction (DEJ) extracellular matrix may be a key link that confers psoriatic proliferation potential and T cell activation dependence in this disease. Both involved and uninvolved areas of psoriasis reveal increased Fn deposition at the DEJ. We show preliminary data that demonstrates that the Fn deposited is of an embryonic splice variant that splices in the EDA segment (EDA Fn). EDA which confers enhanced display of the RGD site, but is rarely stably expressed in adult human tissue. Further, keratinocytes from psoriatic individuals show increased alpha5beta1 integrin expression, a fibronectin receptor, enhanced integrin dependent spreading and focal adhesion kinase (FAK) activation, specifically on Fn. It is our hypothesis that increased Fn, Fn receptor, and Fn dependent functional activation in psoriasis are in response to the DEJ EDA Fn, and that an EDA-Fn rich versus poor ECM will regulate keratinocyte cell cycle progression and responsiveness to in combination with immune mediators. We propose to identify the cell type(s) in psoriasis responsible for production of EDA Fn, using assays for protein and mRNA with in situ localization. We will test whether or not EDA Fn regulates the FAK activation and proliferation of keratinocytes in a skin equivalent model in which keratinocytes engineered to overexpress EDA Fn production are used to form the epidermis. We will also test whether EDA Fn protein alone induces FAK phosphorylation, and its downstream kinase linked to cell cycle, MAPK, as well as cell cycle markers, in fresh ex-vivo normal and psoriatic keratinocytes. Blocking fragments of EDA Fn to the Fn matrix and specific integrin pair inhibitors will identify therapeutic intervention targets. This project will provide important and novel information that will translate rapidly to new treatments for psoriasis as well as to understand morphogenesis, with broad relevance to development, aging and cancer.
