Zhao Xiaoxuan, Jiang Yuepeng, Ren Jiajie, Wang Yunrui, Zhao Yan, Feng Xiaoling
Heilongjiang University of Chinese Medicine, Harbin, China.
School of Basic Medical Sciences, Zhejiang Chinese Medical University, Hangzhou, China.
Front Pharmacol. 2022 Jan 24;13:794938. doi: 10.3389/fphar.2022.794938. eCollection 2022.
Impaired decidualization was recognized as one of the crucial pathomechanisms accounting for unexplained recurrent spontaneous abortion (URSA). Currently, the exact molecular mechanism and targeted clinical decision are still in exploration. Bushen Huoxue decoction (BSHXD) has previously been proved effective in treating URSA, but its mechanism remains to be elucidated. This study aimed to explore the regulation mechanism of BSHXD in decidualization from its intervention in autophagy so as to rationalize its potential as a novel therapeutic regime for URSA. Decidua tissues were collected from patients with URSA and healthy pregnant women who underwent legal terminations for non-medical reasons at the first trimester. Besides, cell line T-hESCs was utilized to establish induced decidualization model, and were randomly divided into ESC group, DSC group, 3-MA group, AMPK siRNA group, scrambled siRNA group and AMPK siRNA + BSHXD group. Transmission electron microscopy, Monodansylcadaverine (MDC) assay, qRT-PCR, immunohistochemistry, immunofluorescence and Western blotting were used to evaluate the level of decidualization, autophagy and activation of AMPK signaling pathway in decidua tissues and cell experiments. Experiments on decidua tissues showed that decidualization was impaired in URSA with inhibited autophagy. Besides, pAMPK T172 and pULK1 S556 were decreased, and pmTOR S2448 and pULK1 S757 were increased. Cell experiments showed that the level of autophagy increased during induced decidualization, but when autophagy was inhibited, decidualization was impaired. In addition, AMPK/mTOR/ULK1 affected decidualization by mediating autophagy, and BSHXD improved decidualization through this mechanism. In conclusion, this study clarified that the inhibition of autophagy mediated by AMPK/mTOR/ULK1 was associated with impaired decidualization, and the intervention of BSHXD on this pathological process may be a vital mechanism for its treatment of URSA. This study laid the foundation for further research and application of BSHXD.
蜕膜化受损被认为是不明原因复发性自然流产（URSA）的关键病理机制之一。目前，确切的分子机制和针对性的临床决策仍在探索中。补肾活血汤（BSHXD）先前已被证明对治疗URSA有效，但其机制仍有待阐明。本研究旨在从其对自噬的干预作用来探讨BSHXD在蜕膜化中的调控机制，从而阐明其作为URSA新型治疗方案的潜力。收集URSA患者以及孕早期因非医学原因依法终止妊娠的健康孕妇的蜕膜组织。此外，利用细胞系T-hESCs建立诱导蜕膜化模型，并将其随机分为胚胎干细胞（ESC）组、蜕膜细胞（DSC）组、3-甲基腺嘌呤（3-MA）组、腺苷酸活化蛋白激酶（AMPK）小干扰RNA（siRNA）组、乱序siRNA组和AMPK siRNA + BSHXD组。采用透射电子显微镜、单丹磺酰尸胺（MDC）检测、实时定量聚合酶链反应（qRT-PCR）、免疫组织化学、免疫荧光和蛋白质免疫印迹法评估蜕膜组织和细胞实验中蜕膜化水平、自噬水平以及AMPK信号通路的激活情况。蜕膜组织实验表明，URSA患者的蜕膜化受损且自噬受到抑制。此外，磷酸化AMPK（pAMPK）T172和磷酸化UNC-51样激酶1（pULK1）S556水平降低，磷酸化哺乳动物雷帕霉素靶蛋白（pmTOR）S2448和磷酸化ULK1 S757水平升高。细胞实验表明，诱导蜕膜化过程中自噬水平升高，但当自噬受到抑制时，蜕膜化受损。此外，AMPK/雷帕霉素靶蛋白（mTOR）/ULK1通过介导自噬影响蜕膜化，而BSHXD通过该机制改善蜕膜化。总之，本研究阐明了AMPK/mTOR/ULK1介导的自噬抑制与蜕膜化受损有关，BSHXD对这一病理过程的干预可能是其治疗URSA的重要机制。本研究为BSHXD的进一步研究和应用奠定了基础。
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