Tumor Necrosis Factor-alfa (TNF) is a cytokine that inhibits cell proliferation and prolactin release from anterior pituitary cells. TNF release as well as TNF effects are higher in anterior pituitary cells from proestrus rats and are estrogen-dependent. Also, TNF induces death by apoptosis in anterior pituitary cells, specially in lactotropes and somatotropes. The major goal of the present project is to determine the mechanisms involved in anterior pituitary cell renewal, in particular the role of TNF/TNF-R1 and the Fas/FasL systems in cell death by apoptosis in the anterior pituitary during the estrous cycle. First, we will evaluate whether apoptosis of anterior pituitary cells is induced by TNF-R1 and Fas activation. Then, we will explore the effect of TNF/TNF-R1 and Fas/FasL system son anterior pituitary cell apoptosis during the estrous cycle. In order to investigate whether TNF and FasL effects are directly exerted on lactotropes and somatotropes or mediated by paracrine actions on other cell types, we will localize the presence of TNF-R1, Fas and FasL in different anterior pituitary cell subpopulations by inmmunofluorescence. The expression of TNF-R1, Fas and FasL during the estrous cycle as well as the modulation of TNF-R1, Fas and FasL gene expression by gonadal steroids will also be investigated. Further, we will study the influence of gonadal steroids on the induction of apoptosis of anterior pituitary cells by theTNF/TNF-R1 and Fas/FasL systems. We will as well examine the influence of gonadal steroids on TNF- and Fas induced apoptosis by infecting anterior pituitary cells with adenoviral vectors expressing TNF or FasL. This research will be done primarily in Argentina at Buenos Aires University in collaboration with Adriana Seilicovich as a complement of an NIH grant. The experiments involving the generation, purification and characterization of recombinant adenovirus vectors will be performed at the Gene Therapeutics Institute, Cedars-Sinai Medical Center, LA. The same applyies for the viral infections of anterior pituitary cells in primary cultutre, their implementation, analysis of the data and interpretation of the results.