ELECTROLYTE TRANSPORT IN CULTURED CERVICAL CELLS

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The applicant, George Gorodeski, is an Obstetrician-Gynecologist and an Assistant Professor of Reproductive Biology at Case Western Reserve University with a long-term commitment to research career in Physiology of Reproduction. His past work on steroid function in the human uterus in-vivo generated several publications. At present, his research is directed at the cellular level and his broad, long term objective is to study mechanisms and regulation of fluid and solute transport in the human female reproductive tract and their role in human reproduction and women's health. The specific aims of the present study are: 1. to optimize the methodology of culturing human ectocervical epithelial (hECE) cells, which are a good in-vitro model of the woman's lower genital tract epithelium, as a polarized epithelium on a porous filter and to characterize the cultured epithelium using morphological, biochemical and functional criteria. 2. to characterize electrolyte transport by the cultured hECE cells with particular emphasis on active NaCl secretion and absorption, and the regulatory mechanisms involved. The hypothesis is that this epithelium controls the luminal fluid environment by actively secreting fluid and solutes from the blood into the lumen, that this process is under the tonic control of steroid hormones and retinoids and that it can be acutely modulated by neurohumoral secretagogues or by mediators of local inflammatory response. The health relatedness of the project is: 1. the data will provide new knowledge of cellular mechanisms and regulation of fluid secretion in the woman's reproductive tract. 2. The information gained will make it possible to design clinical studies of the local use (i.e. luminal) of agents which regulate ion transport in order to control fluid secretion in the woman's genital tract. The experimental design is to study transepithelial NaCl transport using cellular electrophysiology properties. The methods to be used are: 1. to quantitate active Na+ and CI- transport as short circuit current (Isc) generated by cultured hECE cells mounted in Ussing chamber. 2. The nature of the actively transported ions under specific conditions will be determined from Isc changes following treatment with transport inhibitors or following ion replacement in the bathing solution. 3. To quantitate Cl- transport in non-confluent hECE cultures using fluorescence microscopy. The study will be done under the sponsorship of Dr. s' Scarpa and Hopfer at the Departments of Physiology and Biophysics and Reproductive Biology at Case Western Reserve University.