Effect and underlying mechanism of Bu-Shen-An-Tai recipe on ovarian apoptosis in mice with controlled ovarian hyperstimulation implantation dysfunction

Abstract

The effect and underlying mechanism of Bu-Shen-An-Tai recipe on ovarian apoptosis in mice with controlled ovarian hyperstimulation (COH) implantation dysfunction were studied. The COH implantation dysfunction model in mice was established by intraperitoneal injection of 7.5 IU pregnant mare's serum gonadotrophin (PMSG), followed by 7.5 IU human chorionic gonadotrophin (HCG) 48 h later. Then the female mice were mated with male at a ratio of 2:1 in the same cage at 6:00 p.m. The female mice from normal group were injected intraperitoneally with normal saline and mated at the corresponding time. Day 1 of pregnancy was recorded by examining its vaginal smears at 8:00 a.m. of the next day. Fifty successfully pregnant mice were equally randomly divided into 5 groups: normal control pregnant group (NC), COH implantation dysfunction model group (COH), low dosage of Bu-Shen-An-Tai recipe group (LOW), middle dosage of Bu-Shen-An-Tai recipe group (MID) and high dosage of Bu-Shen-An-Tai recipe group (HIGH). Then from day 1, the mice in different groups were respectively intragastrically given corresponding treatments at 9:00 a.m. for 5 consecutive days. The concentrations of 17β-estradiol (E) and progesterone (P) were determined by radioimmunoassay (RIA). The ultrastructural changes of ovarian tissues were observed by transmission electron microscope (TEM). The histopathological changes of ovarian tissues were observed by HE staining. The number of atretic follicles and pregnant corpus luteum were also recorded. TUNEL was applied to measure apoptotic cells of ovarian tissues. Western blotting was used to detect the protein expression of apoptosis- related factors like Bax, Bcl-2 and cleaved-caspase-3 in ovarian tissue of mice. The results showed that ovarian weight, the concentrations of E2 and P4, the number of atretic follicles and pregnant corpus luteum, as well as the apoptosis of granulosa cells were significantly increased in the COH group. The ultrastructures of ovarian tissues in the COH group showed that chromatin in granulosa cells was increased, agglutinated, aggregated or crescent-shaped. The focal cavitation and the typical apoptotic bodies could be seen in granulosa cells in the late stage of apoptosis. After the treatment with different doses of Bu-Shen-An-Tai recipe, the ultrastructural changes of ovarian granulosa cells apoptosis were dramatically improved and even disappeared under TEM. Visible mitochondria and mitochondrial cristae were increased and vacuoles were significantly reduced. The lipid dropltes were shown in a circluar or oval shape. The protein expression levels of Bax and cleaved-caspase-3 were decreased, and the expression of Bcl-2 protein was increased after treatment. It was concluded that Bu-Shen-An-Tai recipe can inhibit the apoptosis of ovarian granulosa cells, probably by up-regulating the protein expression of Bcl-2 and down-regulating Bax and cleaved-caspase-3, which contributes to the formation and maintenance of ovarian corpus luteum. It's helpful to promote the embryonic implantat...

Bushen Huoxue recipe attenuates early pregnancy loss via activating endometrial COX2-PGE2 angiogenic signaling in mice

“…The second possibility is that BSHXR directly facilitated embryo implantation. Based on the results of our previous study, there were no notable differences in the implantation sites on ED5 [22,23]. However, along with embryo development, the rate of fetal loss was higher in COH mice, and could be reversed by treatment using BSHXR [22,23].…”

“…Only mice with two successive regular estrous cycles were included in subsequent studies. Next, a modified COH model was established based on the protocols reported in our previous study [23]. Specifically, female mice in the model group and in all treatment groups were intraperitoneally injected with 0.4 IU/g pregnant mare serum gonadotrophin (PMSG) (Hangzhou Animal Medicine Factory, China) at diestrus.…”

“…In our previous studies, we established a mouse model using ovarian hyperstimulation to simulate disturbed steroid hormone levels and abnormal endometrial development during fresh embryo transfer. A higher postimplantation loss was observed in mice that underwent COH, and BSHXR prevented this loss by facilitating angiogenesis via fibroblast growth factor 2 (FGF2) in the endometrium from ED5 to ED8 [22,23]. In this study, we aimed to further explore the mechanism of BSHXR in regulating endometrial angiogenesis on ED4 midnight, a time point critical for the initiation of implantation.…”

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…”