The long-term objective of this research is to elucidate the cellular mechanisms underlying the squamous metaplasia that develops in the conjunctival epithelium of patients with aqueous tear deficiency. The hypothesis tested during this project period is that squamous metaplasia of the conjunctival epithelium is a state of chronic activation of the conjunctival epithelium that is perpetuated by increased levels of inflammatory cytokines produced by """"""""stressed"""""""" conjunctival epithelium as well as reduced concentrations of lacrimal gland-secreted factors in the tear fluid that normally promote homeostasis.
Specific Aim 1 test the prediction that the conjunctival epithelium of patients with aqueous tear deficiency and squamous metaplasia has increased proliferation and abnormal differentiation in comparison to control tissues from patients with tear film disorders who do not have squamous metaplasia and from normal controls.
Specific Aim 2 tests the prediction that the severity of squamous metaplasia is inversely correlated with the concentration of epidermal growth factor in the tear fluid, and directly correlates with concentrations of inflammatory cytokines in the conjunctival epithelium.
Specific Aim 3 test in culture systems of human conjunctiva epithelium whether proliferative rate and production of differentiation markers can be directly regulated by lacrimal gland-secreted tear factors (vitamins A, EGF, and TGF-B1) and inflammatory cytokines (TNF-a, ILI-a, IL-6, IL-8). Methods include immunofluorescent staining, confocal microscopy, Western Blot, RT-PCR, ELISA, and organotypic culture systems. These studies will provide information about the underlying cellular pathology of squamous metaplasia that may be used to target specific therapies to prevent or treat this morbid condition in patients with dry eye, as well as information about the mechanisms that regulate the growth and differentiation of the conjunctival epithelium in healthy eyes.
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