基因修饰鼠所致下丘脑-垂体-性腺轴的功能改变与性别关联的心肌肥大

The clinical observations indicate that there is a significant gender-related difference in incidence and mortality of cardiovascular diseases. Gender-related phenotypes in cardiovascular system have also been observed in various genetically modified mice. Cardiac hypertrophy is a common complication of many cardiovascular disease such as hypertension and atherosclerosis of etc., and its occurrence is related with the increases of intracellular free calcium in cardiomyocytes. We observed that disruption of FKBP12.6 gene in mice resulted in cardiac hypertrophy in adult male mice, but not in females despite the fact that both male and female knockout mice display similar dysregulation of calcium release. Interestingly, the female knockout mice treated with tamoxifen, an estrogen receptor antagonist, develop cardiac hypertrophy. It has also been reported that the adult male CD38 knockout mice develop cardiac hypertrophy and we observed that the CD38 male knockout mice have a marked enhance in cardiovascular functions along with a significant increase of blood androgen. These results suggest that the changes of sex-hormones in various genetically modified mice play an important role in the cardiac hypertrophy induced by disruption of a gene. This proposal is to investigate the molecular mechanisms of the function changes of hypothalamic-pituitary-gonad axis regulated by the FKBP12.6-mediated CaN-NFAT signaling pathway or CD38-related Sirt1. Elucidation of the mechanisms will provide important experimental evidences for prevention and treatment of the gender-related cardiovascular diseases.

临床观察表明，心血管疾病的发生率及死亡率存有明显的性别差异。动物实验显示，许多基因敲除鼠所致的心血管系统功能和病理改变也与性别密切相关。心肌肥大是心血管疾病如高血压、动脉粥样硬化等的一种常见并发症，其发生发展与细胞内游离钙浓度升高密切相关。我们曾观察到敲除FKBP12.6基因可引起小鼠心肌细胞的钙释放异常，但仅成年雄性基因敲除鼠发生心肌肥大，给予雌性基因敲除鼠它莫西芬则可诱导其心肌肥大。有研究表明，敲除CD38基因可导致成年雄性小鼠心肌肥大，我们的研究也显示雄性CD38基因敲除鼠的心功能明显增强，且与其血浆雄激素水平升高有关。提示性腺激素水平改变在基因缺失引起的心肌肥大中起重要作用。本项目旨在探讨FKBP12.6介导的CaN-NFAT信号传导通路和CD38介导的SIRT1对下丘脑-腺垂体-性腺轴功能的调节作用及其分子机制，从而为基因缺失或突变所致性别相关的心血管疾病的防治提供实验依据。

临床观察及动物研究表明，心血管疾病的发生率及死亡率存有明显的性别差异。我们曾观察到敲除FKBP12.6基因可引起成年雄性鼠心肌肥大，给予雌性基因敲除鼠它莫西芬则可诱导其心肌肥大，提示性腺激素水平改变在基因缺失引起的心肌肥大中起重要作用。据此，我们提出假说：基因修饰（如基因缺失或突变）所致的下丘脑-腺垂体-性腺轴功能异常是导致性别相关的心血管系统改变如心肌肥大发生的主要机制。本课题利用已经证实存有性别差异表型的多种基因敲除鼠模试图验证我们的假说，从而为基因缺失或突变所致性别相关的心血管疾病防治提供实验依据。我们的结果证明，基因修饰如敲除FKBP12.6或CD38基因，或下丘脑特异性高表达GnRH均可显著激活小鼠的下丘脑-腺垂体-性腺轴，进而导致性别相关的心血管表型，并对其可能的分子机制进行了深入的系统探讨。我们的主要发现有：1）敲除FKBP12.6基因所致成年雄性小鼠心肌肥大的心肌细胞主要钙离子通路蛋白如CaMKⅡ、CaN和性激素受体蛋白表达与对照组比较无显著变化，提示FKBP12.6缺失所致的心肌肥大不完全依赖于FKBP12.6缺失引起心肌细胞内钙离子异常释放，激活钙离子相关的心肌肥大通路，而可能与性激素水平的升高有关。2）敲除FKBP12.6基因可致成年小鼠的性腺轴功能增强，性激素水平显著升高，而摘除FKBP12.6-/-雄性小鼠睾丸则可预防其心肌肥大的发生，表明雄激素升高是FKBP12.6基因敲除所致心肌肥大的主要原因。3）证明FKBP12.6缺失通过激活脑垂体的CaN/NFAT等信号通路而增强下丘脑-腺垂体-性腺轴（HPGs）的功能，其机制与FKBP12.6缺失促进细胞内钙信号相关基因表达，进而显著提升细胞内Ca2+浓度以及促进性激素相关基因如LHβ，FSHβ表达有关。4）CD38缺失可显著促进小鼠下丘脑组织中GnRH1表达，进而激活下丘脑-脑垂体-性腺轴，导致性激素水平升高，其机制与CD38缺失导致细胞内NAD+升高，进而激活Sirt1信号通路有关。5）我们制备的可诱导的下丘脑特异性GnRH1转基因鼠可成功模拟下丘脑-脑垂体-性腺轴激活所呈现的性别差异性心血管表型。

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