TESTICULAR INSULIN-LIKE GROWTH FACTOR BINDING PROTEINS

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Understanding of spermatogenesis is limited by lack of knowledge about local control mechanisms within the seminiferous tubule. The objective of this proposal is to address this gap in understanding by demonstrating that insulin-like growth factor-I(IGF-I) and IGF binding protein-3 (IGFBP-3) are locally-produced autocrine/paracrine modulators of spermatogenesis. IGFBP-3 is the predominant IGF carrier protein produced by rat and mouse Sertoli cells in culture. FSH markedly lowers (IGFBP-3 mRNA and protein concentrations in the Sertoli cell and IGFBP-3 is a potent inhibitor of IGF-I actions on the cultured Sertoli cell. There is preliminary evidence from other investigators that IGF-I stimulates spermatogonial cell proliferation. Based on these data, the principal hypothesis is that IGF-I stimulates spermatogenesis and IGFBP-3 inhibits these actions of IGF-I. The strategy to test this hypothesis is to employ two model systems; organ culture of isolated seminiferous tubules and transgenic mice in which the expression IGF-I and/or IGFBP-3 is under the direction of the regulatory elements of the Sertoli cell-specific androgen binding protein gene. Specific aim 1 proposes to examine the effects of IGF-I on spermatogenesis. Transgenic mice with targeted overexpression of IGF-I and isolated seminiferous tubules from normal mice will be used. Proliferation will be assessed by tritiated thymidine incorporation, bromodeoxyuridine labelling and counting of the testicular homogenates; differentiation will be assessed by analysis of specific markers of the different stages of spermatogenesis, histological staging and measurement of lactate and aromatase activity. The second specific aim is to determine whether IGFBP- 3 inhibits IGF-I actions. The same methods will be used as in Specific Aim 1 except that the transgenic mice will have targeted overexpression of IGFBP-3. Finally, the mice overexpressing IGF-I and IGFBP-3 will be crossed to determine if their effects can be neutralized. These studies should establish whether IGFBP-3 is a physiological autocrine/paracrine modulator of IGF-I activity in the testis,. These results will contribute to an understanding of unexplained male infertility and will have general implications pertaining to how the IGFs and their BPs function with in different tissues of the body.