Although cell-hormone interactions during anterior pituitary gland (APG) development have been studied extensively, these studies have not significantly clarified which factors or conditions are necessary to establish the major cell types in the APG. Unlike cell-hormone interactions, cell-matrix interactions in the APG have not been investigated thoroughly, but are known to be essential for normal differentiation in a number of tissues. These interactions could be vital to APG differentiation. Preliminary data shows that fetal APG cells differentiate to secrete significantly more hormones on reconstituted basement membrane gels than on single ECM components, plastic or glass. Basement membrane gels have the major components of the natural basement membrane in contact with embryonic APG cells and therefore can serve as an in vitro model to examine cell-basement membrane interactions that may occur in vivo. The specific hypotheses to be tested are: 1) that laminin, collagen IV, and heparan sulfate proteoglycan are present in the form of a basement membrane among developing APG cells and appear, along with integrin receptors, prior to detectable differentiation of gonadotrophs and lactotrophs; 2) that developing gonadotrophs and lactotrophs respond to reconstituted basement membranes or mixtures of basement membrane components by elevating their secretion of LM, FSH, and PRL; by increasing the corresponding mRNA of these hormones, and by increasing their mitotic rate; 3) that gonadotrophs and lactotrophs are more responsive to releasing and/or inhibiting factors on reconstituted basement membranes or mixtures of basement membrane components than they are on glass or individual basement membrane components; 4) that the observed effects of basement membranes on gonadotrophs and lactotrophs are mediated, in part, by a family of cell-surface extracellular matrix receptors (integrins); and 5) that through integrin receptors, reconstituted basement membranes and/or gel-like mixtures of basement membrane components effect changes in cytoskeletal proteins (such as actin and tubulin) that play a role in regulating the secretion of LH, FSH and PRL and their corresponding mRNA levels. These hypotheses will be tested using a variety of techniques including: light, electron and confocal microscopy; SDS-PAGE, immunoprecipitation and affinity chromatography; cell culture, RIA, in situ and blot hybridizations; Western blots, and perturbation studies. These studies will begin to characterize cell- matrix interactions that appear to significantly influence APG differentiation and function.
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