CHARACTERIZATION OF GONADOTROPIN SECRETING CELL LINES

促性腺激素分泌细胞系的表征

• 批准号: 6033239
• 负责人: KATHRYN G SCHUFF
• 金额: $ 7.55万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6033239
• 关键词: RNase protection assay; Sf9 cell line; affinity chromatography; androgens; estrogens; follicle stimulating hormone; follistatin; gene expression; genetic regulation; green fluorescent proteins; hormone biosynthesis; hormone receptor; inhibin; luteinizing hormone; polymerase chain reaction; radionuclides; secretion; tissue /cell culture; transfection

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.

生殖内分泌学领域一直在努力解决促性腺激素β亚基基因调控的许多方面，这在很大程度上是由于缺乏分化良好的β亚基表达细胞系。 已证明表达和分泌LH的LbetaT 2细胞系的开发（1）现已证明也表达和分泌FSH（2），第二个细胞系FSHTsT 5系也分泌FSH。这两个细胞系是第一个表达所有三种促性腺激素基因的细胞系，初步研究表明它们表现出对这些基因的差异调节。 然而，这两个细胞系的适用性，研究促性腺激素基因是取决于如何以及他们重演正常的促性腺激素生理学，尚未建立。我们假设LbetaT 2和FSHtsT 5细胞系保留了分化的促性腺激素功能的元素，并建议将其作为成熟、分化的促性腺激素细胞的模型进行评估。我们计划测量它们的促性腺激素特异性基因的表达，这些基因对促性腺激素亚基的表达很重要，包括激活素、促性腺激素结合素、卵泡抑素及其受体。这不仅证实了它们适用于LH和FSH的研究，而且也适用于激活素的细胞内信号传导和这些性腺肽之间的复杂相互作用的研究。我们还建议评估这些细胞系对促性腺激素功能的已知激活剂（包括GnRH、激活素、促性腺激素和性腺类固醇）的反应。 这是对有助于LH β、FSH β和α亚基的差异调节和分泌的因子进行更复杂研究的必要基础。此外，该项目支持主要研究者的职业发展计划，结合K 08-DK 02477，为持续的科学生产力和进一步的专业研究技术培训提供技术支持。