Our broad, long-term objective is to understand the mechanisms that regulate movement of fluid and solutes across the woman's reproductive tract epithelium. Our main objective in the present study is to understand how aging in women leads to decreased production of cervical mucus and vaginal fluid. We will utilize a new method that the PI has developed to culture human and cervico/vaginal epithelial cells on filters, and we propose to test four hypotheses: A. That aging is associated with decreased pericellular permeability of the cervical and vaginal epithelia to transport of fluids and solutes, which leads to decreased fluid secretion. B. That aging shifts the steady-state equilibrium of actin filaments in cervical and vaginal epithelial cells towards F- actin; this produces a rigid cytoskeleton which does not allow the cells to contract in response to stimuli and to increase the volume of the intercellular space, thus decreasing the pericellular permeability. Estrogen, in contrast to aging shifts the equilibrium towards G-actin, thus producing a more dynamic cytoskeleton. C. That human cervical and vaginal cells produce nitric oxide (NO), and that NO can inhibit actin polymerization and increase pericellular permeability. D. That in human cervical and vaginal cells, estrogen increases production of NO by up=regulation of calcium-dependent NO- synthase (ecNOS). Methods will include flux and electrophysiological studies done in a modified diffusion/ Using chamber, fluorescence microscopy of attached cells for determinations of changes in cell size, biochemical and molecular biology assays. The present study may provide novel information on age- and estrogen-related effects on cervical- and vaginal-cell function. Health relatedness of the project: the knowledge gained in present study may be important for understanding mechanisms of disease in the cervix e.g. infertility (""""""""cervical factor""""""""), and conditions of excessive or diminished secretion of cervical mucus and vaginal fluid. Understanding mechanisms and regulation of cervical mucus production is important for developing compounds to alter cervical mucus characteristics. These data may improve our understanding of estrogen replacement to postmenopausal women. They may also provide efficient and simple modalities for birth control by targeting known mechanisms of transcervical transport.
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