Estrogen receptor alpha regulates the expression of adipogenic genes genetically and epigenetically in rat bone marrow-derived mesenchymal stem cells

Regulation of the efficacy of epigenetic modifiers is regarded as an important control mechanism in the determination and differentiation of stem cell fate. Studies are showing that the effect of estrogen is important in the differentiation of mesenchymal stem cells (MSCs) into adipocytes, osteocytes, and chondrocytes. Activation of certain transcription factors and epigenetic modifications in related genes play an active role in the initiation and completion of adipogenic differentiation. Understanding the role of estrogen in diseases such as obesity, which increases with the onset of menopause, will pave the way for more effective use of estrogen as a therapeutic option. Demonstration of the differentiation tendencies of MSCs change in the presence/absence of estrogen, especially the evaluation of reversible epigenetic changes, will provide valuable information for clinical applications. In this study, the effect of estrogen on the expression of genes involved in adipogenic differentiation of MSCs and accompanying epigenetic modifications was investigated. Our results showed that estrogen affects the expression of adipogenesis-related transcription factors such as PPARy, C/EBPα and Adipsin. In addition, after estrogen treatment, increased accumulation of estrogen receptor alpha (ERα) and repressive epigenetic markers such as H3K27me2 and H3K27me3 were observed on the promoter of given transcription factors. By using co-immunoprecipitation experiments we were also able to show that ERα physically interacts with the zeste homolog 2 (EZH2) H3K27 methyltransferase in MSCs. We propose that the increase of H3K27me2 and H3K27me3 markers on adipogenic genes upon estrogen treatment may be mediated by the direct interaction of ERα and EZH2. Taken together, these findings suggest that estrogen has a role as an epigenetic switcher in the regulation of H3K27 methylation leading to suppression of adipogenic differentiation of MSC.

The duration of estrogen treatment before progesterone application does not affect neonatal and perinatal outcomes in single frozen blastocyst transfer cycles

Background: The number of frozen embryo transfer (FET) cycles has substantially increased in the past decade. Preparing the endometrium in artificial cycles is widely used in clinical practice. Therefore, how to optimize this program, improve the clinical outcome and ensure the safety of the perinatal period is the focus of our attention. The purpose of this study was to explore whether the duration of estrogen treatment before progesterone application affects neonatal and perinatal outcomes in single frozen blastocyst transfer cycles.Methods: It was a retrospective cohort study. Patients receiving single frozen blastocyst transfer and delivering a single live birth between January 2015 and December 2019 were included. Primary outcome was small for gestational age (SGA). Secondary outcomes were neonatal birthweight, gestational weeks at delivery, preterm birth, low birth weight (LBW), macrosomia, large for gestational age (LGA), neonatal malformation and rate of pregnancy-related complications.Cycles were allocated to four groups according to the estrogen-treatment duration before single frozen blastocyst transfer ①≤12 days (n=306), ②13-15 days (n=620), ③16-18 days (n=471), ④≥19 days (n=275).Results: In total, 1672 cycles were analyzed. Cycles were allocated to four groups according to the estrogen-treatment duration before single frozen blastocyst transfer ①≤12 days (n=306), ②13-15 days (n=620), ③16-18 days (n=471), ④≥19 days (n=275). The rates of SGA among the four groups were 7.8% (24/306), 4.8% (30/620), 5.7% (27/471), and 7.6% (21/275), with no statistical significance (P=0.20). Other neonatal outcomes, including mean neonatal birth weight, gestational weeks at delivery, preterm birth rate, LBW, macrosomia, LGA and neonatal malformation, were comparable among the groups (P=0.38, P=0.16, P=0.20, P=0.58, P=0.20, P=0.34, P=0.96). The rate of pregnancy-related complications was similar among the groups. Multiple logistics regression showed that the duration of estrogen treatment did not affect the rate of singleton SGA (13-15 days, AOR=1.37, 95% CI= 0.70-2.70, P=0.36; 16-18 days, AOR=0.74, 95% CI= 0.40-1.36, P=0.34; ≥19 days, AOR=0.81, 95% CI= 0.44-1.49, P=0.50).Conclusion: The estrogen-treatment duration before progesterone application does not affect neonatal and perinatal outcomes in single frozen blastocyst transfer cycles.

Influence of Estrogen Treatment On ESR1+ and ESR1- Cells In ER+ Breast Cancer: Insights From Single-Cell Analysis of Patient-Derived Xenograft Models

Background: Estrogen typically promotes the progression of hormone-dependent breast cancer through activation of estrogen receptor (ER)-α encoded by ESR1. While estrogen-induced tumor suppression in ER+ breast cancer has been clinically observed as an unexpected outcome of aromatase inhibitor (AI)-resistance, the molecular mechanisms have not yet been fully defined. Characterization of estrogen regulation in two ER+ breast cancer patient-derived xenograft (PDX) models with opposite responses to estrogen offered us an unprecedented opportunity to assess how 17β-estradiol (E2) modulates ER+ cancer.Methods: We established two PDX breast cancer models in mice using ER+ tumors from patients that responded (SC31) or were suppressed (GS3) by exogenous estrogen. In vivo tumor promotion or suppression by estrogen were confirmed through experiments by implanting E2 pellets in mice carrying SC31 or GS3, and then single-cell analysis was performed.Results: E2 promoted SC31 tumor growth but suppressed growth of GS3 in vivo. The E2-mediated suppression of GS3 involves ERα, which was wild-type and not amplified. Single-cell RNA sequencing analysis showed that E2 treatment induced cell cycle promotion in SC31, while E2 induced cell cycle arrest in GS3. However, E2 treatment upregulated the expression of estrogen-regulated genes in both tumors. These gene-expression changes by E2 occurred in both ESR1+ cells and ESR1– cells within the same tumor, demonstrating for the first time the influence of estrogen on ESR1– cells in ER+ breast tumors. E2 also upregulated a tumor suppressor gene, IL24, only in GS3, and lower levels of IL24 were linked to estrogen independence, after three rounds of intermittent E2 treatment. More IL24+ cells were ESR1+ and in G1 phase than IL24– cells. Hallmark apoptosis gene sets were upregulated and the hallmark G2M checkpoint gene set was downregulated in IL24+ cells after E2 treatment.Conclusions: Our study has revealed the effects of estrogen treatment on both ESR1+ and ESR1– cells in ER+ tumors, but not all ER+ cancers respond the same manner to estrogen. SC31 is a tumor that is stimulated by E2, while GS3 is suppressed by E2 via cell cycle arrest. Our results indicate a potential role of IL24 in estrogen-suppressive tumors.

P–696 The duration of estrogen treatment prior to frozen-blastocyst transfer does not impact live birth rate

Study question Does the prolonged duration of oestrogen treatment prior to frozen-blastocyst transfer (FET) affect live birth rate? Summary answer Variation in the duration of estrogen treatment prior to frozen-blastocyst transfer does not impact live birth rate. What is known already With improvements in cryopreservation techniques and fertility preservation, single embryo transfer policy and the increase in freeze-all cycles, frozen blastocyst transfer (FET) has strongly risen over the last years. Artificial endometrial preparation (AEP) is often used prior to FET. The endometrium is prepared by a sequentially treatment of estrogen and progesterone in order to synchronize endometrium and the embryo development. Whether the duration of progesterone administration before FET is well established, the optimal estrogen treatment duration remains controversial. Study design, size, duration All consecutive frozen thawed autologous blastocyst transfer cycles conducted between January 1, 2012 and July 1, 2019 in our University IVF center were included in this retrospective cohort study. We included 2235 single blastocyst FET cycles prepared with hormonal replacement therapy using oral E2 and vaginal progesterone administration in 1376 patients aged from 18 to 43 years. Participants/materials, setting, methods Patient’s characteristics, stimulation characteristics, FET cycles characteristics and cycles outcomes were anonymously recorded and analyzed. Univariate and multivariate analysis were performed. At first, each FET cycle was analyzed individually and secondly taking into account that some of the patients had undergone several FET, the model considered the number of implanting attempts for each woman. Main results and the role of chance We found no significant difference in the mean duration of estradiol administration before frozen embryo transfer between the group live birth versus non-live birth (27.0 ± 5.4 days versus 26.6 ± 5.0 days ; p=0.11). Endometrial thickness was not significantly different between the 2 groups (8.3 ± 1.7 mm versus 8,2 ± 1,7 mm ; p = 0.21). When the duration of estradiol exposure was analyzed in weeks, we observed no difference for the £ 21 days group (OR = 0.97 ; IC 0.64–1.47 ; p = 0.88), 29–35 days group (OR = 0.89 ; IC 0.68–1.16 ; p = 0.37) and > 35 days group (OR = 0.75 ; IC 0.50–1.15 ; p = 0.10) compared to the reference group (22–28 days). After multivariate analysis, the duration of estradiol treatment before frozen embryo transfer did not affect live birth. Limitations, reasons for caution The relatively limited numbers of cycles with more than 35 days or less than 21 days as well as the retrospective design of the study are significant limitations. Wider implications of the findings: Variation in the duration of estradiol supplementation before progesterone initiation does not impact FET outcomes. We therefore can be reassuring with our patients when E2 treatments need to be extended, allowing flexibility in scheduling the day of transfer. Trial registration number Not applicable

Systemic chemotherapy in patients with brain only metastatic breast cancer: A retrospective analysis.

1093 Background: The treatment of patients with brain only metastatic breast cancer (BO-MBC) remains very challenging. There is also very limited literature informing on appropriate treatment or natural history of this entity. Systemic chemotherapy in addition to targeted therapy and/or anti-estrogen treatment is often used, but little is known if it adds to the overall or disease free survival. In this retrospective study, we examine this, as well as other factors which may be associated with increased risk of CNS or systemic recurrence in these patients. Methods: A database search at a single institution identified 178 patients with brain metastases (BM) from breast cancer out of which 45 patients had BO-MBC between 2007-2020. We collected demographic, clinical, radiographic and other treatment data. Leptomeningeal disease (LMD) was diagnosed by cerebrospinal fluid (CSF) cytology, neuroimaging, or both. We used the Brookmeyer and Crowley method. Results: The patients were followed for a median of 17.9 months; 36 out of 45 patients (80%) received local treatment for BM (surgery/radiation/both) and HER2 directed antibodies or tyrosine kinase inhibitors and/or anti-estrogen treatment, whereas 9 out of 45 patients (20%) received systemic chemotherapy in addition. There were 22 out of 45 (49%) HER2 +, 5 out of 45 (11%) HR + and 18 out of 45 (40%) triple negative breast cancer (TNBC) patients. There were 17 out of 45 patients (38%) who were deemed to have low burden of BM (defined as one to three BM and largest being ≤3 cm) whereas there were 24 out of 45 patients (53%) who had high burden of BM (defined as four or more BM or largest being > 3 cm). Conclusions: Patients with BO-MBC represent a distinct entity. Despite having better survival than patients with BM and extra CNS disease these patients have a high risk of developing LMD, CNS and systemic recurrences. The addition of chemotherapy to targeted therapy and/or anti-estrogens does not decrease the rates of systemic or CNS recurrence. The ER+ subset have a lower rate of development of systemic disease, as expected due to their relatively indolent biology. The CNS and systemic recurrence seem to be higher in patients with HER2+ cancers and counterintuitively even in those with low burden of BM; albeit these were statistically insignificant.[Table: see text]

Blood pressure dynamics after pubertal suppression with gonadotropin-releasing hormone analogs followed by estradiol treatment in transgender female adolescents: a pilot study

Objectives The benefits of gonadotropin-releasing hormone analogues (GnRHa) in the treatment of central precocious puberty are well established, and their use is regarded as both safe and effective. Possible adverse effects on blood pressure (BP) and cardiac outcomes, body composition, bone health and brain development, however, continue to be of some concern. The aim of this study was to analyze BP changes in transgender female adolescents before and after receiving GnRHa and after adding estrogen treatment. Methods This was a retrospective pilot study. We analyzed systolic BP (SBP) and diastolic BP (DBP) before and after GnRHa initiation and after adding estrogen. Results Nineteen transgender female adolescents received GnRHa and 15 continued to estrogen treatment. Their baseline SBP and DBP percentiles did not change significantly after either GnRHa or the addition of estrogen treatment. Conclusions Blood pressure is apparently not affected by GnRHa or GnRHa + estrogen treatment in transgender female adolescents. Further larger studies are indicated to confirm these findings.