CHARACTERIZATION OF GONADOTROPIN SECRETING CELL LINES

生殖内分泌学领域在促性腺激素β亚单位基因调控的许多方面都有困难，这在很大程度上是由于缺乏分化良好的β亚单位表达细胞系。LbetaT2细胞系被证明表达和分泌Lh(1)，现在已经被证明也表达和分泌FSH(2)，第二个细胞系FSHtsT5也分泌FSH。这两个细胞系是第一个表达所有三个促性腺激素基因的细胞，初步研究表明，它们对这些基因表现出不同的调节。然而，这两种细胞系对促性腺激素基因研究的适用性取决于它们对正常促性腺激素生理学的概括程度，而这一点尚未得到证实。我们假设LbetaT2和FSHtsT5系的细胞保留了分化的促性腺激素功能的成分，并建议将它们作为成熟的、分化的促性腺激素细胞的模型进行评估。我们计划测量他们的促性腺激素特异基因的表达，这些基因对促性腺激素亚单位的表达非常重要，包括激活素、抑制素、卵泡素及其受体。这不仅证实了它们在研究促黄体生成素和卵泡刺激素方面的适用性，而且也证实了激活素在细胞内的信号传递以及这些性腺肽之间复杂的相互作用的研究。我们还建议评估这些细胞系对已知的促性腺激素激活剂包括GnRH、激活素、抑制素和性腺类固醇的反应。这是对影响LHbeta、FSHbeta和Alpha亚单位差异调节和分泌的因素进行更复杂研究的重要基础。此外，该项目结合K08-DK 02477支持首席调查员的职业发展计划，并为持续的科学生产力和进一步的专门研究技术培训提供技术支持。

The field of reproductive endocrinology has struggled with many aspects of the regulation of the gonadotropin beta subunit genes, in large part due to the lack of a well-differentiated beta-subunit expressing cell line. The development of the LbetaT2 cell line which was demonstrated to express and secrete LH (1) has now been shown to also express and secrete FSH(2) and a second cell line, the FSHtsT5 line, secretes FSH as well. These two cell lines are the first to express all three gonadotropin genes and preliminary studies indicate that they exhibit differential regulation of those genes. However, the applicability of these two cells lines to study of the gonadotropin genes is dependent upon how well they recapitulate normal gonadotroph physiology and that has not yet been established. We hypothesize that cells of the LbetaT2 and FSHtsT5 lines retain elements of differentiated gonadotroph function and propose to evaluate them as models of mature, differentiated gonadotrophs. We plan on measuring their expression of gonadotroph-specific genes important for the expression of the gonadotropin subunits including activin, inhibin, follistatin and their receptors. This will not only confirm their appropriateness for study of LH and FSH, but also the study of the intracellular signalling of activin and the complex interactions between these gonadal peptides. We also propose to evaluate the response of these cell lines to known activators of gonadotroph function including GnRH, activin, inhibin, and the gonadal steroids. This is essential groundwork for more complex studies of factors which contribute to the differential regulation and secretion of LHbeta, FSHbeta and alpha-subunit. In addition, this project supports the Principal Investigator's Career Development Plan in conjuction with K08-DK 02477 with technical support for continued scientific productivity and further specialized research technique training.