亚精胺诱导自噬抗鹅卵巢氧化应激的作用机制

Spermidine, a natural autophagy-inducer, functions as an antioxidant to prevent oxidative stress. Autophagy can enhance the capacity of ovarian follicles against oxidative stress and preserve the follicle pool reserve in female animals. Our studies suggested that spermidine feeding enhanced ovarian autophagy and protected granulosa cells against 3-nitropropionic acid-induced oxidative stress in goose ovaries. Therefore, it is speculated that supplementation of spermidine could protect ovaries against oxidative stress in geese by inducing autophagy. In this project, the geese and primary granulosa cells were treated with spermidine and 3-nitropropionic acid. Based on both in vivo and in vitro, the actions of autophagy-induced by spermidine and protective effects of spermidine against oxidative stress will be clarified in this project. The key signal transduction pathways related to autophagy-induced by spermidine will be screened and identified using transcriptome sequencing technology. The mechanisms of action of autophagy-induced by spermidine in granulosa cells will be determined using mTOR and AMPK inhibitor and the siRNAs for EP300 and MAP1S. Combined treatment with 3-nitropropinonic acid, spermidine, autophagy inducer and autophagy inhibitor, the action mechanism of spermidine against oxidative stress by inducing granulosa cell autophagy will be revealed. This study will be significant support and supplement for studies on action mechanisms of spermidine inducing autophagy and protecting against oxidative stress. And the results in this project will be also provide a novel approach and entry-point to improve the goose egg-laying performance and exploitation of safe and functional feedstuff.

亚精胺是自噬的天然诱导剂并具有抗氧化应激的功能，而自噬也有助于增强卵巢卵泡抗氧化应激能力，并提高卵泡池储备。我们研究表明，亚精胺不仅可诱导鹅卵巢自噬，还可抗3-硝基丙酸诱导的鹅卵巢氧化应激。因此，推测外源性亚精胺可通过诱导自噬抗鹅卵巢氧化应激。项目采用亚精胺和3-硝基丙酸处理鹅和原代卵泡颗粒细胞，从在体和离体水平研究并阐明亚精胺诱导鹅卵巢和颗粒细胞自噬及抗氧化应激的作用；转录组测序筛选并富集亚精胺诱导颗粒细胞自噬的关键信号途径；用mTOR和AMPK抑制剂，EP300和MAP1S的siRNA处理颗粒细胞，研究并揭示亚精胺诱导颗粒细胞自噬的作用机制；用3-硝基丙酸、亚精胺、自噬诱导剂和抑制剂处理颗粒细胞，研究并揭示亚精胺通过诱导颗粒细胞自噬抗氧化应激的作用机制，以期为阐明亚精胺诱导自噬和抗卵巢氧化应激的机制研究提供支撑和补充，将有助于改善鹅产蛋性能，并为开发安全的功能性饲料提供新途径和切入点。

亚精胺作为一种天然的自噬诱导剂，可预防细胞免受自由基介导的氧化损伤，被认为是延长寿命、抗氧化应激和抗衰老的非常有前景的物质。本项目通过3-硝基丙酸建立四川白鹅卵巢及卵泡颗粒细胞氧化应激模型，联合亚精胺灌胃处理，探究亚精胺对鹅卵巢自噬水平及抗氧化能力的影响。在此基础上，应用转录组测序技术筛选亚精胺诱导颗粒细胞自噬的关键基因及信号途径；同时应用自噬诱导剂和抑制剂，干扰自噬关键基因及抑制关键通路，阐明亚精胺诱导卵泡颗粒细胞自噬发挥抗氧化的作用机制。.项目主要获得以下结果：（1）亚精胺能够通过调控自噬关键蛋白表达诱导自噬，提高抗氧化酶活性清除细胞中活性氧和丙二醛进而发挥抗氧化应激作用；（2）亚精胺能够通过多种途径诱导自噬，干扰单一自噬关键信号分子对亚精胺诱导自噬无显著影响；而抑制mTOR/AMPK通路亚精胺能够进一步诱导颗粒细胞自噬；（3）亚精胺能够通过降低颗粒细胞内活性氧水平和调节抗凋亡相关蛋白表达发挥抗凋亡作用；（4）多胺能够通过调控卵泡类固醇激素合成相关基因表达参与卵泡的选择和发育过程；（5）亚精胺能提高肠道组织抗氧化酶活性，减少丙二醛累积；亚精胺还能调节肠道菌群结构，促进有益代谢物的产生进而提高肠道健康水平。综上所述，本研究明确了亚精胺诱导自噬抗氧化应激的作用，探究了亚精胺诱导自噬的作用机制，发掘了亚精胺作为新型饲料原料的潜力，为研究亚精胺调控雌性动物的繁殖功能提供数据支撑，同时为提高畜禽繁殖力提供新途经。