The homeostasis of the corneal epithelium, a rapid self-renewing tissue, is ultimately achieved by orderly proliferation and differentiation of limbal stern cells (SC). The main objective of this research proposal is to explore how limbal SC functions are regulated by their vascularized stromal microenvironment. The healthy state of SC requires a fine balance of SC functions between SC renewal and SC differentiation. The former restores SC population, but the latter depletes it by differentiation into transient amplifying cells (TAC) and then into post-mitotic differentiated cells. Although both are proliferative progenitor cells, limbal SC differ from corneal TAX in location, life span, cell-cycle length and differentiation. The Pi now knows that these mitotic kinetic differences result from intrinsic differences in the autocrines and paracrines generated by epithelial-fibroblast interactionsand by access to serum factors that modulate epithelial cells directly or indirectly via fibroblasts. In this proposal, he focuses on the role of limbal fibroblasts in regulating SC renewal by testing the folowing hypotheses: (H1) SC survival is controlled by an anti-apoptotic survival factor produced by limbal (also 3T3) fibroblasts under hydrocortisone stimulation. (H2) SC renewal is dependent on the above anti-apoptotic activity and preferentially stimulated by keratinocyte growth factor (KGF). The PI has shown that KGF is preferentially produced by limbal fibroblasts and hepatocyte growth factor (HGF) is preferentially produced by corneal fibroblasts. Furthermore, limbal clonal growth is enhanced by KGF HGF IGF-I, but corneal clonal growth is enhanced by EGF KGF TGF-aHGF in the presence of the fibroblast-derived anti-apoptotic activity. Thus, he also propose (h3) kgf acts by stimulating the IGF-IR/IGF-I but not EGF/TGF-a autocrine pathway during SC renewal. (H4) Selective upregulation of KGF expression by IL-1B is a mechanism to explain how SC can be activated during wounding. (H5) TGF-B's differential modulation of KGF and HGF produced by limbal and corneal fibroblasts is a mechanism to explain how SC are paradoxically spared or activated while TAC are inhibited, and explains how SC can be indirectly modulated by non-proteinous factor(s) in the serum. This research will use biochemical cell and molecular biological techniques to unravel the pathogenic mechanisms underlying various corneal diseases that involve limbal SC dysfunctions, and it may, thereby, result in new therapeutic strategies for treating these difficult diseases.
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