Polycystic ovary syndrome (PCOS) is a complex disorder associated with multiple metabolic disturbance, including defective glucose metabolism and insulin resistance. The altered metabolites caused by the related metabolic disturbance may affect ovarian follicles, which can be reflected in follicular fluid composition. The aim of this study is to investigate follicular fluid metabolic profiles in women with PCOS using an advanced sequential window acquisition of all theoretical fragment-ion spectra (SWATH) mass spectrometry. Nineteen women with PCOS and twenty-one healthy controls undergoing IVF/ET were recruited, and their follicular fluid samples were collected for metabolomic study. Follicular fluid metabolic profiles, including steroid hormones, free fatty acids, bioactive lipids, and amino acids were analyzed using the principal component analysis (PCA) and partial least squares to latent structure-discriminant analysis (PLS-DA) model. Levels of free fatty acids, 3-hydroxynonanoyl carnitine and eicosapentaenoic acid were significantly increased (P < 0.05), whereas those of bioactive lipids, lysophosphatidylcholines (LysoPC) (16:0), phytosphingosine, LysoPC (14:0) and LysoPC (18:0) were significantly decreased in women with PCOS (P < 0.05). Additionally, levels of steroid hormone deoxycorticosterone and two amino acids, phenylalanine and leucine were higher in the PCOS patients (P < 0.05). Women with PCOS display unique metabolic profiles in their follicular fluid, and this data may provide us with important biochemical information and metabolic signatures that enable a better understanding of the pathogenesis of PCOS.

Objective To evaluate the effect of electro-acupuncture (EA) in infertile patients with phlegm-dampness polycystic ovary syndrome-insulin resistance (PCOS-IR). Methods Seventy-six PCOS-IR patients who underwnet in vitro fertilization and embryo transfer (IVF-ET) were equally assigned to two groups according to a random digital table: the EA group and the control group, with 38 cases in each group. Before undergoing IVF, the two groups were treated with EA or pseudo-acupuncture, respectively, for 3 menstrual cycles. The intervention was 25 min twice a week until the day of oocyte collection. The selected acupoints were Zhongwan (RN 12), Tianshu (ST 25), Daheng (SP 15), Daimai (GB 26), Qihai (CV 6), Guanyuan (CV 4), and bilateral points including Xuehai (SP 10), Fenglong (ST 40), Zusanli (ST 36), and Yinlingquan (SP 9). Evaluation of phlegm-dampness syndrome score and IR score were carried out before and after treatment. Additionally, the number of oocytes retrieved, transplantable embryo rate, high-quality embryo rate, clinical pregnancy rate and live birth rate were compared between the two groups. Real-time polymerase chain reaction analysis was used to monitor the mRNA expression of the insulin receptor substrate (IRS-1), phosphatidylinositiol 3-kinase (PI3K) and glucose transport factor 4 (GLUT4) in ovarian granulosa cells. Results EA treatment reduced the phlegm-dampness syndrome score as well as the IR scores compared with the control group (P0.05). Moreover, the transplantable embryo rate [49.0% (284/580) vs. 41.9% (273/652)], high-quality embryo rate [36.6% (104/284) vs. 27.8% (76/273)], and live birth rate [50% (19/38) vs. 26.3% (10/38)] in the EA group were significantly higher than in the control group (P<0.05). Gene expression analyses revealed significantly elevated IRS-1, PI3K and GLUT4 mRNA in ovarian granulosa cells of the EA group compared with the control group (P<0.05). Conclusions EA may ameliorate the effects of phlegm-dampness syndrome and ovarian IR in PCOS-IR patients. Mechanistically, this effect might be through an upregulation of the IRS-1/PI3K/GLUT4 signaling pathway, which may result in improved oocyte quality and embryonic development potential. (Registration No. ChiCTR1800015453).

Background: Follicular fluid is an important external environment for the growth and development of oocytes. A thorough identification of specific components in follicular fluid can better the existing understand of intracellular signal transduction and reveal potential biomarkers of oocyte health in women undergoing assisted reproductive therapy. To study on follicular fluid metabolomics components at different ages based on lipid metabolism, we have adopted a new method of SWATH to MRM(the sequential window acquisition of all theoretical fragment-ion spectra to multiple reaction monitor)metabolomics to provide extensive coverage and excellent quantitative data. This was done to investigate the differences in follicular fluid of patients undergoing in vitro fertilization (IVF) and embryo transfer in different age groups and to further explore the relationship between follicular fluid, age and reproductive function.Method: A combination of Ultra-high-performance liquid chromatography and high resolution mass spectrometry techniques were used to analyze the follicular fluid of 230 patients enrolled for the IVF cycle. The patients were of different ages grouped into two groups:the younger and older patients.The obtained multidimensional chromatographic data were processed by principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA). The charge ratios and mass numbers enabled for the identification of different fragments in the samples. Matching information obtained through database search and the fragment information obtained by fragment ion scan structurally identified substances in the samples. This was used to determine the differential compounds.Results: The quality of oocytes decline with age,and the lipid composition in follicular fluid also changes,The lipid metabolism that changes with age may be related to the quality of oocytes.The main differences were in lipid metabolites. Some were up-regulated: Arachidonate, LysoPC(16:1), LysoPC(20:4) and LysoPC(20:3) while others were down-regulated: LysoPC(18:3) and LysoPC(18:1).Conclusions: Metabolomic analysis of follicular fluid revealed that with the increase in age, several differential metabolites are at play. Among these metabolites, lipid metabolism undergoes significant changes that affect the development of oocytes thus causing reduced fertility in older women. These differential metabolites related to follicular development may provide possible detection and treatment targets for promoting oocyte health, and provide scientific basis for understanding the environment of oocyte development.

In our earlier study, we showed that the expression of microRNA‐221‐3p (miR‐221‐3p) was significantly lower in women of advanced age with diminished ovarian reserve (DOR) compared with young women with normal ovarian reserve (NOR). Therefore, in this study, we aimed to explore how miR‐221‐3p regulates apoptosis of granulosa cells and the pathogenesis of DOR. Bioinformatics prediction and dual‐luciferase reporter assay were conducted to identify the target gene of miR‐221‐3p. miR‐221‐3p expression was manipulated by transfecting KGN cells with miR‐221‐3p mimics, inhibitor, and negative control. Following transfection, apoptosis of granulosa cells was determined by flow cytometry, and the expression of the target gene was measured by quantitative real‐time polymerase chain reaction (qRT‐PCR) and western blot analysis (WB). In addition, the expression of the target gene in granulosa cells of DOR patients and NOR patients was measured. miR‐221‐3p were found to directly bind the 3ʹ untranslated region of Forkhead box O1 (FOXO1). Transfection with miR‐221‐3p mimics significantly decreased the apoptosis rate of KGN cells compared with transfection with miR‐221‐3p inhibitors. The expression level of miR‐221‐3p was negatively correlated with the messenger RNA and protein levels of the FOXO1 gene. Besides, FOXO1 expression was upregulated in DOR patients. In conclusion, these results provide evidence that downregulation of miR‐221‐3p expression promotes apoptosis of granulosa cells by upregulating FOXO1 expression, thus serving an important role in DOR pathogenesis.

The complexity of follicular fluid metabolome presents a significant challenge for qualitative and quantitative metabolite profiling, and for discovering the comprehensive biomarkers. In order to address this challenge, a novel SWATHtoMRM metabolomics method was used for providing broad coverage and excellent quantitative capability to discover the human follicular fluid metabolites related to recurrent spontaneous abortion (RSA) after in vitro fertilization and embryo transfer, and to evaluate their relationship with pregnancy outcome. The follicular fluid samples from the spontaneous abortion group (n = 22) and the control group (n = 22) were analyzed using ultra-performance liquid chromatography high-resolution mass spectrometry. A novel, high-coverage, targeted metabolomics method (SWATH to MRM) and a targeted metabolomics method were used to find and validate the differential metabolites between the two groups. A total of 18 follicular fluid metabolites, including amino acids, cholesterol, vitamins, fatty acids, cholic acid, lysophosphatidylcholine and other metabolites, were identified. In the RSA group, 8 metabolites, namely dehydroepiandrosterone, lysoPC(16:0), lysoPC(18:2), lysoPC(18:1), lysoPC(18:0), lysoPC(20:5), lysoPC(20:4), and lysoPC(20:3), were up-regulated, and 10 metabolites, namely phenylalanine, linoleate, oleic acid, docosahexaenoic acid, lithocholic acid, 25-hydroxyvitamin D3, hydroxycholesterol, 13-hydroxy-alpha-tocopherol, leucine, and tryptophan, were down-regulated. These differential metabolites related to RSA may provide a possible diagnostic basis and therapeutic target for RSA, as well as a scientific basis for elucidating the mechanism of RSA.

Thus far, all clinical trials evaluating the efficacy of embryo transfer strategies have selectively delayed the first frozen embryo transfer (FET) by at least 1 menstrual cycle. Nevertheless, this approach, which is based solely on clinical experience, may create unnecessary psychological stress on infertile patients who are anxious to conceive as soon as possible. This study aimed to investigate whether the time interval between oocyte retrieval and subsequent FET affects reproductive outcomes. We implemented a large retrospective cohort study in a single assisted reproductive technology (ART) unit at a university-based hospital, including 1540 autologous FET cycles performed in freeze-all cycles. The beginning of the FET was classified as either ‘cycle 1’ (performing FET within the first menstrual cycle) or ‘cycle ≥2’ (performing FET after one or more menstrual cycles). Live birth rate (LBR) was the primary outcome of our study. The mean interval for ‘cycle 1’ and ‘cycle ≥2’ FETs was 25.72 ± 5.10 days and 75.33 ± 24.85 days, respectively ( P < .001). The type of controlled ovarian hyperstimulation (COH) and endometrial preparation protocols differed significantly between groups ( P = .008 and P = .004, respectively). However, FET groups were similar in many ways. Univariate analysis showed that there was no significant difference in LBR between the different cycles (33.1% after ‘cycle 1’ FET vs 34.2% after ‘cycle ≥2’ FET, P = .68). To evaluate whether LBR remained unchanged after adjustment for potential confounders, we performed multivariate logistic regression. FET timing had no significant impact on LBR in the first FET (odds ratio [OR]: 1.06, 95% confidence interval [CI]: 0.80–1.39). In accordance with the present study, it might not be necessary for clinicians to wait more than 1 menstrual cycle before performing FET. This allows us to reduce otiose deferment in FET, without adversely affecting reproductive outcomes.