性别决定关键基因RSPO1调控能量平衡与糖脂代谢的机制研究

RSPO1 is one of the few key genes that determine gonadal development. Female patients with RSPO1 homozygous mutations develop male phenotypes, e.g. presence of testis or ovotestis in females. Given reproduction and metabolism are inseparable, genes related to reproductive development may also play an important role in energy metabolism. However, the role of RSPO1 in energy metabolism balance has not been reported. In this proposal, we set up an adipose tissue-specific overexpression RSPO1 mouse model and global RSPO1 gene knockout mice, and blocked serum RSPO1 level with neutralization antibody, and we found that RSPO1 could inhibit browning, reduce basal metabolic rate and therefore promote fat accumulation, while RSPO1 deletion or its function blocking could reduce fat accumulation and resist to high-fat diet induced obesity. Further, when the potential receptor LGR4 was knocked out or their downstram classical Wnt signal pathway blocker was added, the inhibition of RSPO1 on browning was almost absence, suggesting that RSPO1 regulates metabolism dependent on the LGR4-β-catenin signal pathway. We intend to explore and clarify the role and mechanism of RSPO1 in glucose and lipid metabolism in this project, and to identify a potential new hub molecule for core life activities of reproduction and metabolism.

RSPO1是决定雌性性腺发育的少数关键基因之一，RSPO1纯合突变的女患者将出现睾丸/卵睾等男性性腺。生殖与代谢密不可分，生殖发育相关基因往往在能量代谢过程中发挥重要作用。然而，RSPO1在能量代谢平衡中的作用尚未见报道。本项目通过构建脂肪组织特异性过表达RSPO1小鼠、RSPO1基因敲除小鼠、中和抗体阻断等手段，发现RSPO1抑制棕色化、降低基础代谢率而促进脂肪累积，RSPO1缺失或功能阻断则减少脂肪累积，抵抗肥胖。机制探讨发现，RSPO1可抑制脂肪前体细胞棕色化，当敲除其潜在配体LGR4后、或加入经典Wnt信号通路阻断剂后，其抑制棕色化作用几近消失，提示RSPO1依赖于LGR4-β-Catenin信号通路调控代谢。本项目拟继续探索RSPO1在糖脂代谢中的作用与机制，以期阐明RSPO1在调控性别发育的同时也发挥调控能量和糖脂平衡的作用，是“生殖与代谢”核心生命活动的新枢纽分子。

最近的遗传证据将WNT下游突变与脂肪分布联系起来。然而，WNT在人类肥胖中的作用仍不清楚。在这里，我们使用全外显子组测序(WES)筛选1,994名肥胖病例和2,161名对照的所有Wnt相关旁分泌因子，并确定12名肥胖患者在RSPO1(p.R219W/Q) 中具有相同位点突变，增加人类肥胖症的易感性。RSPO1主要在内脏脂肪中表达，特别是在成纤维细胞簇中富集，并且随着肥胖而增加。在脂肪组织中过表达人RSPO1小鼠在高脂饮食(HFD)下会出现肥胖，原因子阿姨棕色/米色脂肪产热减少。相反，RSPO1敲除通过增加产热作用来抵抗HFD诱导的肥胖。从机制上讲，RSPO1过表达或给药通过LGR4–Wnt/β-catenin信号通路显著抑制脂肪细胞线粒体呼吸和产热。重要的是，携带热点突变 (p.R219W) 的人源化敲入小鼠表现出抑制产热的表型，并模拟了肥胖携带者的肥胖特征。该突变破坏了RSPO1与细胞外基质的静电相互作用，导致RSPO1过度释放，从而激活 LGR4-β-catenin信号并减弱成熟米色脂肪细胞中的产热能力。因此，我们的研究结果表明，作为旁分泌Wnt信号增强分子RSPO1的功能获得性突变和过表达会抑制脂肪产热，并导致人类肥胖。