Uterine Wet Weight Research Articles

The impact of heterotopic ovarian tissue transplantation with varied volumes on reproductive endocrine function in oophorectomized rats

Background This study aimed to investigate the effects of different volumes of ovarian tissue transplantation on the reproductive endocrine function of rats after oophorectomy. Methods Female rats were selected to establish a castration model and then underwent different volumes of ovarian tissue transplantation. Group I served as the sham operation group. The transplantation group was divided into five subgroups based on the calculated ratio of ovarian weight to body weight in normal female rats, δ = (2.52 ± 0.17) ×10−4: Group II: transplanted ovarian volume was δ; Group III: 0.75δ; Group IV: 0.5δ; Group V: 0.25δ; Group VI: without ovarian transplantation. The post-transplant oestrous cycle recovery was observed, and blood samples were collected every 2 weeks to measure serum hormone levels. Histological evaluation was performed at the end of the observation period. Results Rats in Group V exhibited disrupted oestrous cycles after transplantation, which were significantly longer than those in Group I. Rats in Groups II, III, and IV showed no cyclic changes. At 6 weeks post-transplantation, rats in Group V had lower E2 and AMH levels and higher FSH levels compared to Group I. The uterine wet weight and the number of normal follicles in Group V were significantly lower than those in Group I, but the number of atretic follicles was higher than in Group I. Conclusion The larger ovarian tissue transplantation resulted in a faster recovery with a higher survival rate of the uterus and normal follicles, compared to smaller ovarian tissue transplantation.

Subacute exposure to DEHP leads to impaired decidual reaction and exacerbates the risk of early miscarriage in mice.

To investigate the effect of subacute exposure of Di (2-ethylhexyl) phthalate (DEHP) on endometrial decidualization and early pregnancy miscarriage in mice. CD1 mice were orally administrated with 300 (low-dose group), 1000 (medium-dose group), or 3000 mg·kg－1·d－1 DEHP (1/10 LD50, high-dose group) for 28 days, respectively. An early natural pregnancy model and an artificially induced decidualization model were established. The uterine tissues were collected on D7 of natural pregnancy and D8 of artificially induced decidualization, respectively. The effects of a subacute exposure to DEHP on the decidualization of mice were detected by HE staining, Masson staining, TUNEL assay, and Western blotting. A model of spontaneous abortion was constructed in mice after subacute exposure to 300 mg·kg－1·d－1 DEHP, and the effect of impaired decidualization on pregnancy was investigated by observing the pregnancy outcome on the 10th day of gestation. Compared with the control group, the conception rate was significantly decreased in the high-dose DEHP subacute exposure group (P<0.05). HE staining showed that, compared with the control group, the decidual stromal cells in the low- and medium-dose exposure groups were disorganized, the nuclei of the cells were irregular, the cytoplasmic staining was uneven, and the number of polymorphonuclear cells was significantly reduced. Masson staining showed that compared with the control group, the collagen fibers in the decidua region of the DEHP low-dose group and the medium-dose group were more distributed, more abundant and more disorderly. TUNEL assay showed increased apoptosis in the decidua area compared to the control group. Western blotting showed that the expression of BMP2, a marker molecule for endometrial decidualization, was significantly reduced (P<0.05 or P<0.01). The abortion rate and embryo resorption rate were increased, and the number of embryos, uterine wet weight, uterine area and placenta wet weight were decreased in DEHP low-dose group compared to the control group stimulated by mifepristone, an abortifacient drug (P<0.05 or P<0.01). Subacute exposure to DEHP leads to impaired endometrial decidualization during early pregnancy and exacerbates the risk of adverse pregnancy outcomes in mice.

Maternal exposure to beta-Cypermethrin disrupts placental development by dysfunction of trophoblast cells from oxidative stress

As a broad-spectrum and efficient insecticide, beta-Cypermethrin (β-CYP) poses a health risk to pregnancy. It matters the mechanisms of maternal exposure to β-CYP for impacting reproductive health. The placenta, a transient organ pivotal for maternal-fetal communication during pregnancy, plays a crucial role in embryonic development. The effect of β-CYP exposure on the placenta and its underlying molecular mechanisms remain obscure. The objective of this study was to investigate the effect of β-CYP exposure on placental development and the function of trophoblast, as well as the underlying mechanisms through CD-1 mouse model (1, 10, 20 mg/kg.bw) and in vitro HTR-8/SVneo cell model (12.5, 25, 50, 100 μM). We found slower weight gain and reduced uterine wet weight in pregnant mice with maternal exposure to β-CYP during pregnancy, as well as adverse pregnancy outcomes such as uterine bleeding and embryo resorption. The abnormal placental development in response to β-CYP was noticed, including imbalanced placental structure and disrupted labyrinthine vascular development. Trophoblasts, pivotal in placental development and vascular remodeling, displayed abnormal differentiation under β-CYP exposure. This aberration was characterized by thickened trophoblast layers in the labyrinthine zone, accompanied by mitochondrial and endoplasmic reticulum swelling within trophoblasts. Further researches on human chorionic trophoblast cell lines revealed that β-CYP exposure induced apoptosis in HTR-8/SVneo cells. This induction resulted in a notable decrease in migration and invasion abilities, coupled with oxidative stress and the inhibition of the Notch signaling pathway. N-acetylcysteine (an antioxidant) partially restored the impaired Notch signaling pathway in HTR-8/SVneo cells, and mitigated cellular functional damage attributed to β-CYP exposure. Collectively, exposure to β-CYP induced oxidative stress and then led to inhibition of the Notch signaling pathway and dysfunction of trophoblast cells, ultimately resulted in abnormal placenta and pregnancy. These findings indicate Reactive Oxygen Species as potential intervention targets to mitigate β-CYP toxicity. The comprehensive elucidation contributes to our understanding of β-CYP biosafety and offers an experimental basis for preventing and managing its reproductive toxicity.

Dioscorea alata L. Tubers Ethanol Extract Improves Insulin Resistance and Estrogen Defectively in Type 2 Diabetic Ovariectomized Rats

Dioscorea alata L., commonly known as water yam, has attracted the attention of scientific researchers for its potential anti-diabetic and estrogenic properties. However, the existing literature lacks specific studies on the anti-diabetic and estrogenic effects of D. alata in the context of type 2 diabetic ovariectomized rats. Thus, Our research focuses on investigating the influence of D. alata on the antidiabetic and estrogenic properties in rats. These rats have developed diabetes through a high-fat diet and streptozotocin induction and have also undergone ovariectomy, making them type 2 diabetic ovariectomized rats (T2DM OVX). This comprehensive study both in vitro and in vivo methodologies. We conducted a phytochemical analysis, revealing the presence of key bioactive constituents, including flavonoids, alkaloids, terpenoids, tannins, saponins, cardiac glycosides, and steroids. The results of our phytochemical screening confirm the remarkable antidiabetic properties of the D. alata extract, as it effectively inhibits α-glucosidase activity, leading to a significant reduction in fasting blood glucose levels. Additionally, it enhances body weight, serum insulin levels, and pancreatic islet size. The extract also demonstrates notable estrogenic effects by significantly increasing uterine wet weight, endometrial and myometrium thickness, bone calcium, and overall bone density, while reducing bone porosity in the T2DM OVX rats. It is important to note that while the D. alata extract exhibits slightly less potent α-glucosidase inhibition compared to acarbose, it showcases potent antidiabetic activity at a dose of 1,000 mg/kg body weight, comparable to Metformin (250 mg/kg body weight). In terms of estrogenic activity, it is slightly less potent than 17β-estradiol (1 mg/kg body weight). These findings provide robust evidence supporting the traditional use of D. alata and its efficacy in the context of antidiabetic and estrogenic properties within the T2DM OVX model.

Effects of Dichlorodiphenyltrichloroethane on the Female Reproductive Tract Leading to Infertility and Cancer: Systematic Search and Review.

Persistent Organic Pollutants (POPs) such as dichlorodimethyltrichloroethane (DDT) are present and ubiquitous in the environment due to their resilient nature. DDT is a prevalent endocrine disruptor still found in detectable amounts in organisms and the environment even after its use was banned in the 1970s. Medline and Google Scholar were systematically searched to detect all relevant animal and human studies published in the last 20 years (January 2003 to February 2023) in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. In total, 38 studies were included for qualitative synthesis. This systematic search and review indicated that exposure to DDT is associated with female reproductive health issues, such as reduced fecundability; increased risk of preterm/premature deliveries; increased periods of gestation; alterations in the synthesis of crucial reproductive hormones (Progesterone and Oxytocin) through ion imbalances and changes in prostaglandin synthesis, myometrial and stromal hypertrophy, and edema; and variations in uterine contractions through increased uterine wet weight. There was also limited evidence indicating DDT as a carcinogen sufficient to instigate reproductive cancers. However, this review only takes into account the in vitro studies that have established a possible pathway to understand how DDT impacts female infertility and leads to reproductive cancers. Links between the pathways described in various studies have been developed in this review to produce a summarized picture of how one event might lead to another. Additionally, epidemiological studies that specifically targeted the exposure to DDT of females belonging to various ethnicities have been reviewed to develop an overall picture of prevailing female reproductive health concerns in different nations.

Structure-Activity Relationships of Triphenylethylene Derivatives and Their Evaluation as Anticancer and Antiviral Agents.

Tamoxifen (TAM) is a selective estrogen receptor modulator (SERM) that is used in the treatment of breast cancer, yet with the risk of developing uterine cancer. A perfect SERM would act as an estrogen activator on bones, the cardiovascular system, and the central nervous system while providing neutral or estrogen blocking effects on the breast and the uterus. Herein, we report on the design, synthesis, and evaluation of new rigid and flexible TAM analogues. Mainly, a chloro substituent is introduced at the para position of the TAM ring C blocking the CYP2D6 hydroxylation site. Most compounds showed estrogenic activity higher than TAM using the yeast estrogen screen assays, indicating the determinant role of the chloro substituent upon functional activity. Despite being estrogenic, compound 2B showed potent antiproliferative activity in the NCI 60 cell lines with mean GI50 = 3.67 μM, GI50 = 1.05 μM on MCF-7 cell lines, and GI50 = 1.30 μM on MDA-MB-231. The estrogenic activity of compound 2B was further confirmed by stimulating alkaline phosphatase in Ishikawa cells, and it showed no increase in relative uterine wet weight in ovariectomized rats. Compound 2F showed EC90 = 0.31 μg/mL and SI90 = 60 against Ebola virus; this is 200-fold more potent than the positive control favipiravir. This is the first time to report estrogenic triphenylethylenes as anti-EBOV agents. The anti-EBOV activity reported is a function of the substitution pattern of the scaffold rather than the functional activity. Moreover, compound 3D showed excellent PO pharmacokinetic properties in mice. In conclusion, for this class of TAM-like compounds, the blockage of the p-position of ring C is decisive for the functional activity; meanwhile, the triarylethylene substitution pattern is detrimental for the antiviral activity.

The phytoestrogenic potential of flavonoid glycosides from Selaginella moellendorffii via ERα-dependent signaling pathway.

Selaginella moellendorffii (SM) has been applied as an ethnic drug to treat conditions such as osteoporosis, idiopathic thrombocytopenic purpura, and chronic inflammation. It is known to be rich in flavonoids, including apigenin glycosides and unique elements of bioflavonoids. To investigate estrogen-like constituents of SM and the possible mechanism. We identified the main components in liquid chromatography and liquid chromatography-mass spectrometry. The estrogenic effects were examined using a recombinant yeast screening assay, an E-screen cell proliferation assay, and an in vivo uterotrophic assay. Flavonoid glycosides extract, some flavonoid glycosides, and apigenin showed estrogen agonistic activity in the yeast screening assay. They also induced cell proliferation in estrogen receptor-positive (ER+) cells but not in estrogen receptor-negative (ER-) cells. Consistently, the protein expression of ERα, phosphorylation protein kinase B (p-AKT), phosphatidylinositol 3 kinase (PI3K), phosphorylation mammalian target of rapamycin (p-mTOR), phosphorylation 38,000-Da protein (p-P38), and phosphorylation extracellular-regulated kinase 1/2 (p-ERK1/2) elevated following treatment with flavonoid glycoside extract (P<0.01 or P<0.05). These effects could be blocked by ER antagonist or ERα antagonist but not be blocked by ERβ antagonist. In vivo assay, flavonoid glycoside extract could significantly increase body weight, serum estradiol level, uterine wet weight, alter uterine morphology, and promote ERα protein expression (P<0.01 or P<0.05). ERα induction via mitogen-activated protein kinases (MAPK) and PI3K/Akt/mTOR pathways might be the possible mechanism underlying the phytoestrogen effect of SM, and the flavonoid glycosides might be the critical estrogenic constituents.

Pro-Sexual Effects of Aqueous and Methanol Extracts of Phyllanthus muellerianus (Kuntze) Exell (Phyllanthaceae) on a Model of Low Sexual Desire Disorder in Female Rats

Low sexual desire disorder is a recurrent deficiency in desire for sexual activity which impairs lifestyle. It is a frequent problem in women with limited therapeutic options. Phyllanthus muellerianus (Kuntze) Exell is a plant used by traditional healers to boost libido in women. This study aimed at evaluating the aphrodisiac properties of root barks extracts of this plant on a model of Low Sexual Desire Disorder (LSDD) in female rats. Low sexual desire was induced by ovariectomy plus low steroid hormones supplementations. Thirty ovariectomized rats were treated for 21 days with either distilled water, aqueous or methanol extracts (60 or 372 mg/kg) of P. muellerianus. All animals were supplemented with a low dose of 17β-estradiol and progesterone prior to sexual behavior test. Sexual behavior test was performed each week by pairing each female rat with a sexually experienced male. Sexual motivation (approach, ear wiggling, hops and darts, anogenital presentation and aggressive behavior frequency) and sexual receptivity parameters (lordosis frequency and quotient) were recorded. At the end of treatments, animals were sacrificed, uteri and vagina collected, weighed and conserved for total uterine proteins assay and histology. LSDD was characterized by significant inhibition of sexual motivation parameters and lordosis frequency compared to a normal control. Moreover, poor reproductive tissues growth characterized by low total uterine proteins, uterine wet weight and uterine epithelia size was noted in LSDD group compared to normal control. Interestingly, plant extracts significantly improved sexual motivation parameters (p &lt; 0.05-0.001) compared to LSDD group. In addition, moderate reproductive tissues growth was noted in plant treated groups as confirmed by amelioration of uterine cell integrity. Present results show that P. muellerianus exhibits pro-sexual effects through amelioration of sexual motivation and reproductive tissues growth on a model of LSDD.

Ethanol-Extracted Cameroonian Propolis Counteracts Tamoxifen-Induced Endometrial Hyperplasia by Modulating Apoptosis and Proliferation-Regulating Proteins in the Ovaries of Intact Wistar Rats.

Tamoxifen is the most common adjuvant that has been widely used in the treatment of positive estrogen receptor (ER+) breast cancer for over 20 years. However, long term exposure to tamoxifen doubles the risk of endometrial cancer. The association of tamoxifen with antiproliferative substances could abrogate its side effects on the endometrium. Recently, we demonstrated that ethanol-extracted Cameroonian propolis (EECP) has chemopreventive effects on ER+ breast cancer in rats. This study evaluated the capability of EECP to counteract tamoxifen-induced endometrial hyperplasia, without altering its effect on the breast. Thirty-six rats of ∼2 months were coadministered either EECP (16.5, 50, and 150 mg/kg BW) or fulvestrant (300 μg/kg BW) and tamoxifen (10 mg/kg BW) for 8 weeks. Afterward, the relative weights and histomorphometry of the uterus, vagina, ovaries, and mammary gland were assessed. The expression of some proteins of proliferation (PCNA), angiogenesis (VEGF), and apoptosis (Bax, Bcl-2, and caspase-3) was measured by immunohistochemistry. Rats that received only tamoxifen had endometrial hyperplasia compared to normal rats. EECP and fulvestrant protected the rats against tamoxifen-induced endometrial hyperplasia. A significant decrease in uterine wet weight (p < 0.01); endometrial height (p < 0.001); and expression of PCNA, Bcl-2, and VEGF proteins as well as a significant increase in the expression of Bax and caspase-3 proteins was observed in the EECP group compared to the Tamox group. EECP did not change the effects of tamoxifen on the breast. In summary, Cameroonian propolis which is efficacious in preventing breast cancer can also be a good complementary medicine to prevent tamoxifen-induced endometrial cancer in tamoxifen users.

Evaluation of Anti Estrogenic Activity and Anti-Osteoporotic Activity of Extracted Quercetin from Bambusa arundinacea Leaves on Ovariectomized Rats

Phytoestrogens has now become a emerging era of interest for the researchers. The mimicking effects of phytoestrogens had lead to its wide use in reproductive system. In this study it was aimed to study the estrogenic activity and anti-osteoporotic activity of isolated quercetin female Wistar rats. The estrogenic effect was analyzed by uterotropic assay, vaginal cytology and measurement of vaginal opening in female Wistar rats. The administration of isolated quercetin in ovariectomized immature and mature female Wistar rats in a dose of 30 mg/kg b.w. resulted in significant increase in the uterine wet weight when compared with ovariectomized control rats. The treated rats, showing only Cornified epithelial cells was an indication of the presence of the estrogen and also showed 100% vaginal opening. Thirty six female albino Wistar rats were randomly divided into six groups (n=6). After 60 days of Ovariectomy, animals were treated with of isolated quercetin for the next 45 days. Finally femur bone length, weight, bone ash calcium level, and bone mineral density (BMD) were estimated. The levels of serum alkaline phosphates (ALP), calcium, and phosphorous, and bone histopathology were also evaluated. OVX-induced increased serum ALP, calcium, and phosphorous levels were impaired in of isolated quercetin treated rats. The isolated quercetin which was evident by uterotropic assay, measurement of vaginal opening, and histopathological changes. It also exhibited a significant anti-osteoporotic effect in the experimental model of OVX-induced osteoporosis in rats, indicating advantageous effect in postmenopausal osteoporosis. Thus it concludes the Anti Estrogenic activity and Anti-Osteoporotic activity of extracted Quercetin from Bambusa arundinacea leaves on ovariectomized rats.

Effect of 50-Hertz Sinusoidal Vibration on the Uterus in Ovariectomy-Induced Osteoporotic Rats.

Objective To evaluate the influence of sinusoidal vibration (50-Hertz) stimulation on the uterus of osteoporotic rats. Methods We constructed an osteoporosis rat model by ovariectomy (OVX). 36 3-month-old Sprague Dawley rats were randomly divided into the control group, vibrating group, sham operation group, sham operation vibrating group, OVX group, and OVX vibrating group (n = 6 per group). Rats started to vibrate one week after the operation: one 10 minutes 50-Hertz sinusoidal vibration per day, except for Saturday and Sunday. In the second, 8, and 12 week after vibration stimulation, rats were sacrificed in batches. And then, the uteruses were taken out to measure the wet weight and calculate uterus relative wet weight. Results Compared with the control group, OVA induced a significant increase in wet weight and relative wet weight in rat uterus. The vibration was to the uterus wet weight and the uterus relative wet weight in ovariectomized rats and at the same time had no significant effect, but the 12-week prolonged vibration can significantly reduce the uterus wet weight and the uterus relative wet weight in ovariectomized rats than 2 weeks. Conclusions The uterus wet weight and the uterus relative wet weight were increased in the OVA-induced osteoporosis rats. The 50-Hertz sinusoidal vibration had no significant effect on the uterus wet weight and the uterus relative wet weight in the ovariectomized rats at the same time, but 12 weeks of vibration can significantly reduce the uterine wet weight and uterine relative wet weight of ovariectomized rats. And the uterus relative wet weight can be used as a new indicator of stimulating the uterus.

Estrogenic Effect of Bambusa arundinacea, Trichosanthes dioica and Punica granatum on Rats

Background: Phytoestrogens have recently become a hot topic among scientists. Phytoestrogens’ estrogen-like properties have led to their widespread use in the reproductive system. The aim of this research was to see whether the ethanolic extract of Bambusa arundinaceae, Trichosanthes dioica and Punica granatum had any estrogenic activity in female wistar rats. Methods: In female wistar rats, the estrogenic effect was studied using a uterotropic assay, vaginal cytology and vaginal opening. In ovariectomized immature and mature female wistar rats, a 400 mg/kg body weight (b.w.) dose of ethanolic extract of Bambusa arundinaceae, Trichosanthes dioica and Punica granatum was given. Result: When compared to ovariectomized control rats, the uterine wet weight increased significantly. The estrogen-treated rats had only cornified epithelial cells, indicating the existence of oestrogen, as well as 100% vaginal opening. At 400 mg/kg b.w., the ethanolic extract of Bambusa arundinaceae, Trichosanthes dioica and Punica granatum demonstrated promising estrogenic activity, as evidenced by uterotropic assays, vaginal opening measurements and histopathological changes. As a result of this research, it’s possible to infer that the ethanolic extract of Bambusa arundinaceae, Trichosanthes dioica and Punica granatum play an important role in estrogenic activity in female rats.

Manipulating Estrogenic/Anti-Estrogenic Activity of Triphenylethylenes towards Development of Novel Anti-Neoplastic SERMs.

Selective estrogen receptor modulators (SERMs) act as estrogen receptor (ERα) agonists or antagonists depending on the target issue. Tamoxifen (TAM) (a non-steroidal triphenylethylene derivative) was the first SERM approved as anti-estrogen for the treatment of metastatic breast cancer. On the hunt for novel SERMs with potential growth inhibitory activity on breast cancer cell lines yet no potential to induce endometrial carcinoma, we designed and synthesized 28 novel TAM analogs. The novel analogs bear a triphenylethylene scaffold. Modifications on rings A, B, and C aim to attenuate estrogenic/anti-estrogenic activities of the novel compounds so they can potentially inhibit breast cancer and provide positive, beneficial estrogenic effects on other tissues with no risk of developing endometrial hyperplasia. Compound 12 (E/Z-1-(2-{4-[1-(4-Chloro-phenyl)-2-(4-methoxy-phenyl)-propenyl]-phenoxy}-ethyl)-piperidine) showed an appreciable relative ERα agonistic activity in a yeast estrogen screen (YES) assay. It successfully inhibited the growth of the MCF-7 cell line with GI50 = 0.6 µM, and it was approximately three times more potent than TAM. It showed no potential estrogenicity on Ishikawa endometrial adenocarcinoma cell line via assaying alkaline phosphatase (AlkP) activity. Compound 12 was tested in vivo to assess its estrogenic properties in an uterotrophic assay in an ovariectomized rat model. Compared to TAM, it induced less increase in wet uterine wet weight and showed no uterotrophic effect. Compound 12 is a promising candidate for further development due to its inhibition activity on MCF-7 proliferation with moderate AlkP activity and no potential uterotrophic effects. The in vitro estrogenic activity encourages further investigations toward potential beneficial properties in cardiovascular, bone, and brain tissues.

Abstract 1236: Preclinical characterization of LY3484356, a novel, potent and orally bioavailable selective estrogen receptor degrader (SERD)

Abstract Nearly 70% of newly diagnosed breast cancers are estrogen receptor alpha (ERα) positive, for which endocrine therapy is a primary treatment. However, approximately 40% of those patients on endocrine therapy develop resistance, which includes mutations in ERα (ESR1) that drive constitutive activation of the receptor. One approach to overcome this resistance is to develop a potent degrader and pure antagonist of the estrogen receptor, which can effectively suppress estrogen receptor signaling. Fulvestrant, a pure antagonist and selective estrogen receptor degrader (SERD) is approved for the treatment of HR+/HER2- metastatic breast cancer. However, fulvestrant is limited by poor pharmacokinetics properties, limited exposure and sub-optimal in vivo estrogen receptor degradation. Here we describe the preclinical profile of LY3484356, an oral SERD and pure estrogen receptor antagonist, with potent activity against the wild type and mutant estrogen receptor. LY3484356 has Ki values of 0.64 nM and 2.8 nM against wild type ERα and Y537S mutant ERα proteins, respectively. It is a potent and highly efficient degrader of wild type ERα and Y537N mutant ERα proteins in cells, with IC50 values 3.0 nM and 9.6 nM, respectively. LY3484356 is also a potent inhibitor of ERα-mediated transcription in vitro and in vivo. It inhibits cell proliferation in wild type ERα and ESR1 Y537N mutant breast cancer cell lines, with average IC50 values of 3 nM and 17 nM, respectively. In a panel of breast cancer cell lines, 11 out of 12 ER+ breast cancer cell lines were sensitive to LY3484356 (IC50 less than 100 nM), whereas all ER- cell lines tested were insensitive. LY3484356 has demonstrated sustained and prolonged target inhibition (&amp;gt;75% inhibition of PGR transcription up to 96h after last dose) in ESR1 wild type (MCF7) and ESR1 Y537S mutant (ST941/C) xenograft tumors. Consistent with its profile as a pure antagonist, immature rat studies demonstrated no significant effect on uterine wet weight. LY3484356 demonstrated significant tumor growth inhibition and tumor regressions in wild type ESR1 breast cancer xenograft models such as MCF7, T47D and ZR-75-1, as well as ESR1 mutant breast cancer PDX models. LY3484356 has shown synergy or additivity in combination with CDK4/6 inhibitor abemaciclib, mTOR inhibitor everolimus and PIK3CA inhibitor alpelisib in inhibiting cell proliferation in ER+ breast cancer cell lines in vitro and tumor growth inhibition in relevant xenograft or PDX models in vivo. The first-in-human Phase 1/2 clinical trial of LY3484356 (EMBER, ClinicalTrials.gov NCT04188548) is currently ongoing and a window-of-opportunity study evaluating the pharmacodynamic effects of LY3484356 in early stage breast cancer is expected to begin early 2021. Citation Format: Shripad V. Bhagwat, Baohui Zhao, Weihua Shen, Cecilia Mur, Robert Barr, Lisa J. Kindler, Almudena Rubio, Jolie A. Bastian, Jeffrey D. Cohen, Brian E. Mattioni, Eunice Yuen, Thomas K. Baker, Mark A. Castanares, Dongling Fei, Jason R. Manro, Maria Jose Lallena, Sheng-Bin Peng, Alfonso de Dios. Preclinical characterization of LY3484356, a novel, potent and orally bioavailable selective estrogen receptor degrader (SERD) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1236.

Does 2’-hydroxygenistein inhibit the endometrial proliferation? A preliminary study

Isoflavones, due to their claimed or demonstrated beneficial biological activities, have attracted great interest and gained popularity among the public. On the risk of endometrial hyperplasia, conflicting findings for such compounds have been reported and genistein has been intensively studied. Studies also indicated that 2’-hydroxylation of isoflavones can lead to beneficial components with superior bioactivity compared to isoflavones lacking this substituent. Till now there is no study evaluating the effect of 2’-hydroxygenistein on endometrial hyperplasia in vivo. In line with this, a 3-day uterotrophic assay was carried out to evaluate the effect of 2’-hydroxygenistein on the uterus as its endometrial hyperplasia is of significant clinical concern. Daily subcutaneous administration of 2’-hydroxygenistein significantly (p ≤ 0.05) increased uterine wet weight at 2 and 8 mg/kg/day, while it reduced (p ≤ 0.05) uterine epithelial height at all tested doses. In contrast, no significant variation was observed on vaginal epithelial height. As global result, it appears that 2’-hydroxygenistein might exhibit anti-proliferative effects in the uterus, while having no effect on the vagina. However, this aspect needs to be further investigated.

Assessment of reproductive impact of the aerial parts of Caralluma dalzielii N. E. Br in female Wistar rats

Caralluma dalzielii N. E. Brown belonging to family Asclepiadaceae, is a popular cactus-shaped plant native to East Africa. The aerial parts are used traditionally for treating various diseases including infertility. The present study evaluated the effects of the aqueous extract of the aerial parts of Caralluma dalzielii (AECD) on reproductive performance of female Wistar rats. Adult female virgin rats were allotted into four major groups namely pre-conception, post-conception, implantation site and ovariectomized rats' groups. Each group was subdivided into 4 groups and treated orally with 125, 250, 500 mg/kg of AECD or distilled water (vehicle). In the pre-conception and post-conception groups, litter sizes, pups' weights, deformities, gestation length and reproductive indices were determined. Number of implantation sites and weights of embryos were assessed in the implantation site group while in the ovariectomised rats' group, uterine weights were determined. AECD produced no difference in litter size and reproductive indices in pre-conception group while in post-conception group the litter size at 500 mg/kg was significantly (p < 0.05) reduced compared to the control. Post-implantation loss index was high, and the other reproductive indices were reduced at 500 mg/kg. Whereas at the dose of 125 mg/kg, post-implantation loss index was reduced, and litter size was increased when compared to the control group. At 500 mg/kg, AECD caused a significant (p < 0.05) decrease in the number of implantation sites and weight of embryos while at 125 mg/kg the implantation sites increased. A significant (p < 0.05) increase in the uterine weight in the ovariectomised rats' group was observed at all dose levels. Our study provides scientific evidence that supports the traditional use of AECD in the treatment of infertility. At a lower dose, AECD acts by increasing the number of implantation sites and litter size of animals but at a higher dose, it may be embryotoxic. AECD increases uterine wet weight in ovariectomised rats suggesting that the plant may be oestrogen-like.

Impaired Female Reproductive Function in A Novel Estrogen Receptor Alpha Hypomorphic Mouse Model

IntroductionEstrogen signaling plays an important role in the regulation of both energy homeostasis and reproductive function. Human and animal studies indicate that the metabolic actions of estrogen signaling are primarily mediated by estrogen receptor alpha (ERα). Both human and mouse ERα genes show remarkable heterogeneity due to complicated alternative splicing, which results in a wide variety of variant transcripts with different protein products or a common protein. However, functional significance of this ERα genomic complexity has not been well documented. Here we generated a novel mouse model in which ERα expression was differentially affected in various tissues, and examined their body weight change and reproduction.MethodUsing CRISPR/Cas9 technology, we introduced a loxP‐flanked sequence containing En2 splice acceptor, a green fluorescent marker with mitochondrial localization signal and DNA translation stop signals before the first coding exon of mouse ERα gene for the potential restoration of the ERα expression.ResultsERα protein expressions were significantly reduced in pituitary (~37%) and ovary (~67%), but not in brain, of the homozygous mutant female compared to those of controls, indicating that unknown elements important for ERα tissue‐specific expression were affected. In contrast to body weight gain observed in previous studies of ERα knockout female mice, our hypomorphic mutant females did not show significant body weight gain. On the other hand, virginal smears from the mutant females rarely showed typical estrus stage. Uterine wet weight was significantly lower in the mutant female mice compared to the controls, indicating abnormal reproductive system. Continuous mating study further confirmed delayed pregnancy and inability to have alive newborns in the mutant female mice.ConclusionEstrogen receptor alpha protein expression varies among tissues in terms of the level and pattern of splicing variants. Splicing variant‐specific ERα functions are not fully elucidated. Our results suggest that a particular variant affected in our mouse model is less critical for proper functioning of systems involved in estrogen‐regulated energy metabolism, but indispensable for some aspects of female reproductive function.

Uterine Patterning, Endometrial Gland Development, and Implantation Failure in Mice Exposed Neonatally to Genistein.

Background:Embryo implantation relies on precise hormonal regulation, associated gene expression changes, and appropriate female reproductive tract tissue architecture. Female mice exposed neonatally to the phytoestrogen genistein (GEN) at doses similar to those in infants consuming soy-based infant formulas are infertile due in part to uterine implantation defects.Objectives:Our goal was to determine the mechanisms by which neonatal GEN exposure causes implantation defects.Methods:Female mice were exposed to GEN on postnatal days (PND)1–5 and uterine tissues collected on PND5, PND22–26, and during pregnancy. Analysis of tissue weights, morphology, and gene expression was performed using standard histology, confocal imaging with three-dimensional analysis, real-time reverse transcription polymerase chain reaction (real-time RT-PCR), and microarrays. The response of ovariectomized adults to (E2) and artificial decidualization were measured. Leukemia inhibitory factor (LIF) injections were given intraperitoneally and implantation sites visualized. Gene expression patterns were compared with curated data sets to identify upstream regulators.Results:GEN-exposed mice exhibited reduced uterine weight gain in response to E2 treatment or artificial decidualization compared with controls; however, expression of select hormone responsive genes remained similar between the two groups. Uteri from pregnant GEN-exposed mice were posteriorized and had reduced glandular epithelium. Implantation failure was not rescued by LIF administration. Microarray analysis of GEN-exposed uteri during early pregnancy revealed significant overlap with several conditional uterine knockout mouse models, including Foxa2, Wnt4, and Sox17. These models exhibit reduced endometrial glands, features of posteriorization and implantation failure. Expression of Foxa2, Wnt4, and Sox17, as well as genes important for neonatal uterine differentiation (Wnt7a, Hoxa10, and Msx2), were severely disrupted on PND5 in GEN-exposed mice.Discussion:Our findings suggest that neonatal GEN exposure in mice disrupts expression of genes important for uterine development, causing posteriorization and diminished gland function during pregnancy that contribute to implantation failure. These findings could have implications for women who consumed soy-based formulas as infants. https://doi.org/10.1289/EHP6336

Medial preoptic estrogen receptor-beta blunts the estrogen receptor-alpha mediated increases in wheel-running behavior of female rats

Estrogens are believed to enhance rodent voluntary wheel-running through medial preoptic (mPOA) estrogen receptor α (ERα) signaling, with little role attributed to estrogen receptor β (ERβ). Systemic ERβ activation has been shown to mitigate ERα driven increases in wheel-running. Therefore, the present goal was to determine whether ERβ signaling in the mPOA plays a similar modulatory role over ERα. We utilized outbred wild-type (WT) and rats selectively bred for low voluntary running (LVR) behavior to address whether mPOA ERβ signaling blunts ERα driven wheel-running behavior and immediate-early gene (Fos, Zif268, and Homer1) mRNA induction. Further, we addressed baseline mPOA mRNA expressions and circulating 17β-estradiol levels between female WT and LVR rats. Following ovariectomy, WT rats reduced running behavior ∼40 %, with no effect in LVR rats. Intra-medial preoptic injection of the ERα-agonist propylpyrazoletriol (PPT) increased wheel-running ∼3.5-fold in WT rats, while injections of the ERβ-agonist diarylpropionitrile (DPN) or a combination of the two agonists had no effect. Similarly, ERα-agonism (PPT) increased Fos and Homer1 induction ∼3-fold in WT and LVR isolated mPOA neurons, with no effect of the ERβ-agonist DPN alone or in combination with PPT, suggesting medial-preoptic ERβ activity may blunt ERα signaling. LVR rats exhibited higher mPOA mRNA expressions of Esr1, Esr2 and Cyp19a1, lower normalized uterine wet weights and lower 17β-estradiol plasma levels compared to WT, suggesting their low running may be due to low circulating estrogen levels. Collectively, these findings highlight mPOA ERβ as a potential neuro-molecular modulator of the estrogenic control of wheel-running behavior.

Trichostatin A inhibits uterine histomorphology alterations induced by cigarette smoke exposure in mice

AimsCigarette smoking results in well-known negative reproductive consequences. However, the role of histone deacetylase 1 and 2 (HDAC1/2) in the structural changes of uterine tissues induced by cigarette smoke (CS) exposure and the therapeutic potential of trichostatin A (TSA), a HDAC inhibitor, have not been investigated. Main methodsFemale mice were exposed to CS twice daily for 30 days and TSA was injected intraperitoneally into CS-exposed mice on alternate days in the TSA-treated group. Uteri in the estrus phase were weighed and uterine histomorphology and HDAC1 cell distribution were examined by HE and immunohistochemistry. Markers associated with macro-autophagy (Beclin-1), autophagic flux (increased LC3-II and a lack of p62 accumulation), autophagy inhibiting factor (mTOR, phosphorylated mTOR and its upstream IRS, phosphorylated IRS), HDAC1/2, FOXO1 and FOXO3 were assessed by Western blot. Key findingsCS exposure decreased body weight and triggered uterine histomorphologic alterations, including a thinner myometrium and a reduced number of glandular and interstitial cells. HDAC1/2 were activated in uterine tissues after CS exposure and TSA effectively inhibited HDAC1/2 activation and attenuated the loss of body weight and uterine wet weight induced by CS exposure. TSA effectively restored the thickness of the myometrium and number of glandular and interstitial cells. TSA also restored the expression of markers of macro-autophagy (LC3-II and Beclin-1) and reduced phosphorylated mTOR, phosphorylated IRS, FOXO1 and FOXO3 activation. SignificanceTSA inhibited uterine histomorphologic alterations induced by CS exposure. The TSA effect might be associated with resumption of macro-autophagy via HDAC1/2 inhibition.