Use of Gene Therapy: A Tool to Study Reproductive Function

这个项目的总体目标是探索通过使用 独特的遗传模型系统，用GnRH替代GnRH的性腺功能低下的HPG小鼠 靶向基因治疗。这是对病毒载体系统成功地定向细胞特异性、 生理调节的GnRH表达和释放是HPG生殖功能的完全恢复 老鼠。我们的病毒转基因的独特特征之一是对生殖的再创造至关重要 完整性是利用具有自身同源启动子的GnRH基因。内源性促性腺激素释放激素基因的应用 启动子可能允许调节的、脉动的GnRH的产生和释放，从而刺激脑垂体促黄体生成素和促性腺激素释放。 FSH的产生和导致生殖轴的激活和生育的诱导。我们的初步研究 在注射HSV的HPG女性中，GnRH.GFP扩增子显示出对垂体酮和黄体生成素的协同刺激 FSH的分泌与卵巢和子宫的生长。在HPG雄性中，我们已经证明了在 HSV.GnRH.GFP扩增载体的传递。我们假设GnRH基因的传递和表达 对成年雌性和雄性HPG小鼠下丘脑神经元的影响将导致生殖的完全重演 功能。在第一个目标中，我们建议更充分地描述实现生殖恢复的程度 通过GnRH基因治疗，包括卵泡发生和排卵、精子发生、性生殖 行为和生育能力。在这个模型中生殖功能的成功激活也表明 在HPG小鼠中，GnRH神经元的输入保持不变，因此适当地调节GnRH的输出 可能会发生。在第二个目标中，GnRH救援对调控机制的影响 将探索生殖，包括外围调节器，如正反馈和负反馈效应 性类固醇和中枢调节剂，如Kispeptin。拟议的研究提供了一种遗传模型 选择性和受调控的基因替代疗法，因此可能不仅对 生殖障碍，如不孕不育，但也用于神经科学中的基因治疗应用， 在正常基因调控至关重要的情况下，治疗神经和其他疾病。

The overall goal of this project is to explore the regulatory mechanisms that control reproduction using a unique genetic model system, the hypogonadal, GnRH-deficient hpg mouse with replacement of GnRH by targeted gene therapy. The ultimate test for the ability of viral vector systems to successfully direct cell-specific, physiologically regulated GnRH expression and release is the full recovery of reproductive function in the hpg mice. One of the unique features of our viral transgene that is essential for the recreation of reproductive integrity is the use of the GnRH gene with its own cognate promoter. The use of the endogenous GnRH gene promoter may allow regulated, pulsatile GnRH production and release, and hence stimulate pituitary LH and FSH production and lead to activation of the reproductive axis and induction of fertility. Our preliminary studies in hpg females injected with HSV.GnRH.GFP amplicon demonstrate coordinated stimulation of pituitary LH and FSH secretion and ovarian and uterine growth. In the hpg male, we have demonstrated spermatogenesis after delivery of HSV.GnRH.GFP amplicon vector. We hypothesize that delivery and expression of the GnRH gene to hypothalamic neurons in adult female and male hpg mice will result in the full recapitulation of reproductive function. In the first aim, we propose to more fully characterize the extent of reproductive recovery achieved by GnRH gene therapy, including ovarian folliculogenesis and ovulation, spermatogenesis, sexual reproductive behavior and fertility. Successful activation of reproductive function in this model would also suggest that neuronal inputs to GnRH neurons remain intact in hpg mice, so that appropriate modulation of GnRH output can occur. In the second aim, the effects of GnRH rescue on the regulatory mechanisms that control reproduction will be explored, including peripheral modulators such as positive and negative feedback effects of sex steroids, and central modulators such as kisspeptin. The proposed studies provide a genetic model for selective and regulated gene replacement therapy, and thus may have important implications not only for reproductive disorders such as infertility, but also for applications of gene therapy in the neurosciences, for treatment of neurological and other disorders where normal gene regulation is critical.