Conditional steroidogenic cell-targeted deletion of TSPO unveils a crucial role in viability and hormone-dependent steroid formation
Conditional steroidogenic cell-targeted deletion of TSPO unveils a crucial role in viability and hormone-dependent steroid formation
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DOI：
10.1073/pnas.1502670112
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发表时间：
2015-06-09
影响因子：
11.1
通讯作者：
Papadopoulos, Vassilios
中科院分区：
文献类型：
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作者：
Fan, Jinjiang;Campioli, Enrico;Papadopoulos, Vassilios
转运蛋白(TransLocator Protein，TSPO)是类固醇合成组织中线粒体胆固醇转运复合体的关键成员。为了评估TSPO的功能，我们建立了两个Cre介导的TSPO条件性基因敲除(CKO)小鼠。首先，将性腺细胞靶向的Amhr2-Cre小鼠与TSPO-Flosed小鼠杂交，获得F1 TSPO Amhr2 CKO小鼠(TSPO(f1/fl)；Amhr2-Cre(/+))。性交后12.5天(DPC)胚胎的基因分型证实了4.4%的CKO小鼠意外的孟德尔比例。由于Amhr2-Cre在12.5dpc的性腺中表达，这些发现提示胚胎的植入前选择。对表达数据库的分析显示，在2-细胞和8-细胞受精卵中，Amhr2水平升高，这表明TSPO在桑椹胚期之前是异位沉默的，表明TSPO提高了胚胎致死率，并参与了胚胎发育。为了绕过这个问题，以类固醇生成细胞为靶标的Nr5a1-Cre小鼠与TSPO-Flobled小鼠杂交。由此产生的TSPO(f1/fl)；Nr5a1-Cre(/+)小鼠以正常的孟德尔比例出生。Nr5a1驱动的TSPO CKO小鼠的肾上腺皮质和性腺中的TSPO水平显著降低。用人绒毛膜促性腺激素(HCG)治疗小鼠后，尽管大量脂滴耗尽，但循环中的睾酮水平仍升高。相比之下，Nr5a1驱动的TSPO CKO小鼠在肾上腺皮质激素(ACTH)的作用下失去了形成皮质酮的能力。对于ACTH依赖的类固醇合成很重要，Mc2r、Stard1和Cypa11a1的水平没有受到影响，而Scar1的水平增加，并观察到脂滴的积累，这表明胆固醇用于类固醇的利用被阻断。TSPO在肾上腺髓质中的表达和肾上腺素的增加也被观察到。综上所述，TSPO对植入前胚胎发育和ACTH刺激的类固醇生物合成是必需的。
Translocator protein (TSPO) is a key member of the mitochondrial cholesterol transport complex in steroidogenic tissues. To assess the function of TSPO, we generated two lines of Cre-mediated Tspo conditional knockout (cKO) mice. First, gonadal somatic cell-targeting Amhr2-Cre mice were crossed with Tspo-floxed mice to obtain F1 Tspo Amhr2 cKO mice (Tspo(fl/fl); Amhr2-Cre(/+)). The unexpected Mendelian ratio of 4.4% cKO mice was confirmed by genotyping of 12.5-day-postcoitum (dpc) embryos. As Amhr2-Cre is expressed in gonads at 12.5 dpc, these findings suggest preimplantation selection of embryos. Analysis of expression databases revealed elevated levels of Amhr2 in two-and eight-cell zygotes, suggesting ectopic Tspo silencing before the morula stage and demonstrating elevated embryonic lethality and involvement of TSPO in embryonic development. To circumvent this issue, steroidogenic cell-targeting Nr5a1-Cre mice were crossed with Tspo-floxed mice. The resulting Tspo(fl/fl); Nr5a1-Cre(/+) mice were born at a normal Mendelian ratio. Nr5a1-driven Tspo cKO mice exhibited highly reduced Tspo levels in adrenal cortex and gonads. Treatment of mice with human chorionic gonadotropin (hCG) resulted in increased circulating testosterone levels despite extensive lipid droplet depletion. In contrast, Nr5a1-driven Tspo cKO mice lost their ability to form corticosterone in response to adrenocorticotropic hormone (ACTH). Important for ACTH-dependent steroidogenesis, Mc2r, Stard1, and Cypa11a1 levels were unaffected, whereas Scarb1 levels were increased and accumulation of lipid droplets was observed, indicative of a blockade of cholesterol utilization for steroidogenesis. TSPO expression in the adrenal medulla and increased epinephrine production were also observed. In conclusion, TSPO was found necessary for preimplantation embryo development and ACTH-stimulated steroid biosynthesis.