Effects of Erbuzhuyu Decoction Combined with Acupuncture on Endometrial Receptivity Are Associated with the Expression of miR‐494‐3p

Abstract

Background/Aim. Erbuzhuyu decoction (EBZYD) is a traditional Chinese medicine (TCM) formula and has been used in infertility treatment. Meanwhile, acupuncture is also used to treat female infertility. However, it is unclear whether EBZYD combined with acupuncture has better therapeutic effect. The aim of this study was to explore the effect of EBZYD combined with acupuncture and investigate its mechanism in superovulation mice. Methods. The mice received the treatment of EBZYD, acupuncture, EBZYD combined with acupuncture, or miR-494-3p agomir combined with EBZYD and acupuncture. The blastocysts’ number, endometrial microstructure, and endometrial thickness were observed, followed by the detection of endometrial receptivity-related factors, PI3K/Akt/mTOR pathway-related proteins, and miR-494-3p expression using quantitative real-time polymerase chain reaction (qRT-PCR) or western blot. Luciferase reporter assay was performed to confirm the targeting relationship between HOXA10 and miR-494-3p. Results. EBZYD combined with acupuncture treatment could increase the number of blastocysts, pinopodes, endometrial thickness, and the expression of endometrial receptivity-related factors, and the treatment effect of EBZYD combined with acupuncture was better than EBZYD or acupuncture alone. In addition, EBZYD combined with acupuncture treatment activated PI3K/Akt/mTOR pathway and inhibited the expression of miR-494-3p. HOXA10 is one of the target genes of miR-494-3p. Overexpression of miR-494-3p reversed the therapeutic effect of EBZYD combined with acupuncture and suppressed the expression of HOXA10 and the activity of PI3K/Akt/mTOR pathway. Conclusion. This study suggests that EBZYD combined with acupuncture could improve endometrial receptivity in superovulation mice via miR-494-3p/HOXA10 axis.

Regulation mechanism of miR-494-3p on endometrial receptivity in mice via PI3K/AKT/mTOR pathway

“…Another study found that miR-494-3p was related to endometrial receptivity in the microRNA expression profiling of endometriaassociated infertility patients (Xu et al 2017). Furthermore, overexpression of miR-494-3p has been shown in previous studies to block the activity of PI3K/AKT/mTOR pathway (Yuan et al 2020). However, up to now, there has been no study on the relevant mechanism of how miR-494-3p affects endometrial receptivity via the PI3K/AKT/mTOR pathway, so its mechanism of action cannot be clearly defined.…”

“…Twenty mice with normal estrous cycle were selected for the test, and were randomly divided into four groups: NC (negative control) group, GnRHa group (injecting GnRHa), GnRHa+NC-antagomir group (injecting GnRHa and NC-antagomir), and GnRHa+miR-494-3p antagomir group (injecting GnRHa and miR-494-3p antagomir). Specifically, except for the NC group, mice in the other groups were injected intraperitoneally with 40 μg/100 g of GnRHa to induce implantation disorder (Gong et al 2013;Yuan et al 2020) at 9:00 a.m. every day for 9 consecutive days. At the 9th day, the mice were injected with 0.04 IU/ kg HMG for ovulation induction, and 0.1 IU/kg HCG was injected 48 h later.…”

The Treatment of Complementary and Alternative Medicine on Female Infertility Caused by Endometrial Factors

“…Additionally, some researchers also conducted high-throughput RNA sequencing and bioinformatics analysis on the samples of patients who had received acupuncture treatment or not, and concluded that acupuncture treatment could play a role in changing endometrial receptivity by regulating the differential expression of circular RNAs in infertility patients [ 127 ]. Yuan et al [ 128 ] performed Erbuzhuyu decoction (EBZYD; two-step Evodia decoction) combined with acupuncture on mice, and the results showed that it could promote the expression of endometrial tolerance-related factors and increase blastocyst number and endometrial thickness through activating PI3K/Akt/mTOR signaling pathway, and its treatment effect was superior to using EBZYD or acupuncture alone (see Table 4 ).…”

Autophagy as a Therapeutic Target of Natural Products Enhancing Embryo Implantation

“…Several reports have suggested that traditional herbal formulas and medicinal herbs successfully improve endometrial receptivity and might be an alternative option for improving the outcome of embryo implantation [192][193][194]. Many herbal formulas have been studied for enhancing the embryo implantation rate in animal studies and clinical study [195][196][197][198][199][200][201][202][203][204][205][206][207][208][209][210][211][212][213]. Among these formulas, Erbu Zhuyu decoction and Tokishakuyakusan (TJ-23, Danggui Shaoyao san in Chinese) have been reported to induce autophagy via the expression of Beclin1 and LC3 [208,214,215].…”

“…BaelanChagsangBang -- [236] Bangdeyun and its component DS147 -- [196,197] Buganshen recipe -- [198] BuShenAnTai recipe -- [201] Bushen Tiaoxue Granules and Kunling Wan -- [202] Dingkun Pill -- [206,207,212] Erbu Zhuyu decoction -Increases the Beclin1 and LC3B [204,208] Gushen'antai pills -- [195] Liuwei Dihuang Granule -- [199] Shoutaiwai recipe -- [200] Tokishakuyakusan(Danggui Shaoyao san) -Induces autophagy and mitophagy via increasing PINK1 and LC3 but reducing p62 [203,214,215] Wenshen Yangxue decoction -- [205,211] Xianziyizhen Recipe -- [237] Yeosin-san Paeonia lactiflora and Cyperus rotundus - [213,238] Yiqixue buganshen recipe -- [198] Zhuyun recipe -- [210] Angelica gigas Decusirol Block autophagic flux by suppressing cathepsin C expression [216,221] Cnidium officinale -- [239] Cyperus rotundus -Increases LC3B II/LC3B and Beclin1 [223,229] Paeonia lactiflora Paeoniflorin Induces autophagy via inhibition of AKT/mTOR [114,225,230] Panax quiquefolius (American Ginseng) Ginsenoside Rb1 and Rg1…”