DataSheet1_Chaigui granule exerts anti-depressant effects by regulating the synthesis of Estradiol and the downstream of CYP19A1-E2-ERKs signaling pathway in CUMS-induced depressed rats.doc

Background: There is a significant gender difference in the prevalence of depression. Recent studies have shown that estrogen plays a crucial role in depression. Therefore, studying the specific mechanism of estrogen’s role in depression can provide new ideas to address the treatment of depression. Chaigui granule has been shown to have exact antidepressant efficacy, and the contents of saikosaponin (a, b1, b2, d) and paeoniflorin in Chaigui granule are about 0.737% and 0.641%, respectively. Some studies have found that they can improve depression-induced decrease in testosterone (T) levels (∼36.99% decrease compared to control). However, whether Chaigui granule can exert antidepressant efficacy by regulating estrogen is still unclear. This study aimed to elucidate the regulation of estrogen levels by Chaigui granule and the underlying mechanism of its anti-depressant effect.
Methods: Eighty-four male Sprague-Dawley (SD) rats were modeled using a chronic unpredictable mild stress (CUMS) procedure. The administration method was traditional oral gavage administration, and behavioral indicators were used to evaluate the anti-depressant effect of Chaigui granule. Enzyme-linked immunosorbent assay (ELISA) was adopted to assess the modulating impact of Chaigui granule on sex hormones. Then, reverse transcription-quantitative PCR (RT-qPCR), and Western blot (WB) techniques were employed to detect extracellular regulated protein kinases (ERK) signaling-related molecules downstream of estradiol in the hippocampus tissue.
Results: The administration of Chaigui granule significantly alleviated the desperate behavior of CUMS-induced depressed rats. According to the results, we found that Chaigui granule could upregulate the level of estradiol (E2) in the serum (∼46.56% increase compared to model) and hippocampus (∼26.03% increase compared to model) of CUMS rats and increase the levels of CYP19A1 gene and protein, which was the key enzyme regulating the synthesis of T into E2 in the hippocampus. Chaigui granule was also found to have a significant back-regulatory effect on the gene and protein levels of ERβ, ERK1, and ERK2.
Conclusion: Chaigui granule can increase the synthesis of E2 in the hippocampus of CUMS-induced depressed rats and further exert antidepressant effects by activating the CYP19A1-E2-ERKs signaling pathway.

背景：抑郁症患病率存在显著的性别差异。近期研究表明，雌激素(estrogen)在抑郁症发病机制中发挥关键作用。阐明雌激素在抑郁症中的具体作用机制，可为抑郁症治疗提供全新思路。已有研究证实柴桂颗粒具有确切的抗抑郁功效，其所含柴胡皂苷(a、b1、b2、d)与芍药苷(paeoniflorin)的含量分别约为0.737%与0.641%。部分研究发现，上述成分可改善抑郁症模型中睾酮(testosterone, T)水平的降低（较对照组降低约36.99%）。但柴桂颗粒是否可通过调控雌激素发挥抗抑郁功效，目前尚不明确。本研究旨在阐明柴桂颗粒对雌激素水平的调控作用，及其抗抑郁效应的潜在分子机制。
方法：采用慢性不可预见性温和应激(chronic unpredictable mild stress, CUMS)造模法，对84只雄性斯普拉格-道利(Sprague-Dawley, SD)大鼠构建抑郁症模型。给药方式采用传统灌胃给药，通过行为学指标评估柴桂颗粒的抗抑郁效应。采用酶联免疫吸附测定(enzyme-linked immunosorbent assay, ELISA)法，检测柴桂颗粒对性激素水平的调控作用。随后，通过逆转录实时定量聚合酶链反应(reverse transcription-quantitative PCR, RT-qPCR)与蛋白质印迹(Western blot, WB)技术，检测海马组织中雌二醇下游的细胞外调节蛋白激酶(extracellular regulated protein kinases, ERK)信号通路相关分子。
结果：柴桂颗粒给药可显著缓解CUMS诱导的抑郁模型大鼠的绝望行为。本研究结果显示，柴桂颗粒可上调CUMS模型大鼠血清与海马组织中的雌二醇(estradiol, E2)水平（较模型组分别升高约46.56%与26.03%），同时可提升海马组织中调控睾酮向雌二醇合成的关键酶CYP19A1的基因与蛋白表达水平。此外，柴桂颗粒可显著反向调控ERβ、ERK1与ERK2的基因与蛋白表达水平。
结论：综上，柴桂颗粒可提升CUMS诱导的抑郁模型大鼠海马组织中雌二醇的合成水平，并通过激活CYP19A1-E2-ERKs信号通路发挥抗抑郁效应。