Targeted expression of human growth hormone gene to the fat or to the cartilage in spontaneous dwarf rats.

人类生长激素基因在自发性侏儒大鼠的脂肪或软骨中的靶向表达。

• 批准号: 15590982
• 负责人: KATAKAMI Hideki
• 金额: $ 1.98万
• 依托单位: University of Miyazaki (2004-2005)宮崎医科大学 (2003)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590982/
• 关键词: fat cell-specific hGH expression; chondrocyte-specific hGH expression GH of fat cells; development and differentiation by; hGH-transgenic rats; leptin gene; alphal-collagene gene; IGF-1-independent GH action; α1-collaen遺伝子; 脂肪細胞特異的ヒトGH遣伝子発現

A strain of spontaneous dwarf rats (SDR, dr) is discovered and fully characterized in Japan. They are small and short caused by a point mutation at the splice junction between the third intron and the forth exon of the rat GH gene, which was identified by us and others. They provide us an opportunity to study the IGF-1-independent GH actions to various tissues, such as fat and cartilage. In order to elucidate the IGF-1-independent direct actions of growth hormone to the fat tissue or cartilage, we first generated kimeric genes of the human GH gene (2.1kbp) coupled with leptin gene promoter (-3.5kbp, Lep-hGH : 5.7kbp) or human alpha 1-collagne gene promoter (-5.2kbp, α 1C-hGH : 7.3kbp). By injecting the Lep-hGH gene and frozen sperm into oocytes of dr, we successfully generated a line of human GH-transgenic dr with targeted expression to the fat tissue (Lep-hGH-dr). Lep-hGH-dr expressed hGH not only in fat tissues, but also in testes or ovaries. Small amounts of the hGH gene expressed in the stomach, adrenal and kidney. Lep-hGH-dr showed increased both in weight (1.3-1.5 times, vs. control dr) and body fat (140-150%, vs. control dr). Serum hGH levels in Lep-hGH-dr were detectable by an ultrasensitive EIA, 8.1-949.1pg/ml.Routine bacteriological examination detected contamination with M. pulmonis in the transgenic rat colony. All of the transgenic rats, including Lep-hGH-dr, were sacrificed. Subsequent physiological and histological analyses of Lep-hGH-dr, and generation of α 1C-hGH-dr were both terminated.Based on these results, we conclude that the segment of -2.4kbp upstream promoter of human leptin gene is sufficient to target the hGH gene expression to the fat tissue. The questions have remained unsolved, i.e., whether insufficient amount of expressed hGH gene in the fat or low levels of circulating hGH resulted in unexpected obesity and modest acceleration of growth in Lep-hGH-dr.

在日本发现了一种自发性矮小大鼠(SDR，DR)，并对其进行了充分的特征分析。它们是由我们和其他人鉴定的大鼠GH基因第三内含子和第四外显子之间拼接连接处的点突变造成的。它们为我们提供了一个研究IGF-1非依赖性生长激素对各种组织的作用的机会，如脂肪和软骨。为了阐明生长激素对脂肪组织或软骨的非胰岛素样生长因子-1的直接作用，我们首先构建了人GH基因与瘦素基因启动子(-3.5kBP，α-hGH：5.7kBP)或人α1-胶原基因启动子(-5.2kbp，LEP-1C-hGH：7.3kBP)偶联的同源基因。通过将Lep-hGH基因和冷冻精子注射到DR的卵母细胞中，我们成功地获得了一株脂肪组织靶向表达的人GH转基因DR(Lep-hGH-DR)。LEP-hGH-DR不仅在脂肪组织中表达，而且在睾丸和卵巢中也有表达。少量hGH基因在胃、肾上腺和肾脏中表达。LEP-hGH-DR显示体重(1.3-1.5倍，与对照DR相比)和体脂(140-150%，与对照DR相比)增加。用超灵敏的EIA法检测到LEP-hGH-DR的血清hGH水平为8.1-949.1pg/ml。常规细菌学检查发现转基因大鼠群体中存在肺支原体污染。所有转基因大鼠，包括LEP-hGH-DR，被处死。结果表明，人瘦素基因上游启动子-2.4kbp的片段足以将α基因的表达靶向脂肪组织。这些问题尚未得到解决，即脂肪中hGH基因表达不足或循环中hGH水平低是否导致LEP-hGH-DR意外肥胖和生长轻度加速。