Immediate Effect of Electro-acupuncture on Endometrial Blood Flow in Patients with Recurrent Implantation Failure: A Randomized Controlled Trial.

OBJECTIVE: To investigate the immediate effects of electro-acupuncture (EA) on endometrial blood flow among recurrent implantation failure (RIF) patients. METHODS: Eighty RIF patients, enrolled from March 2022 to December 2022, were randomly allocated into either the EA group (40 cases) or the waiting-list (WL) group (40 cases) by using a random number table. The EA group underwent acupuncture at points of Shenting (GV 24), Baihui (GV 4), Benshen (GB 13), bilateral Zigong (EX-CA 1), Huangshu (KI 16), Sanyinjiao (SP 6) and Xuehai (SP10), and electric acupuncture apparatus was connected to EX-CA 1, KI 16, SP 6, and SP 10 with disperse-dense waves at 4/20 Hz frequencies for 30 min after transvaginal ultrasound, while the WL group received no intervention. The primary outcome measured was the endometrial volume blood flow. The secondary outcomes included the bilateral uterine artery index, endometrial volume, endometrial blood flow type, vascular distribution index (VIMV) for endometrial and ovary, clinical pregnancy rate, and embryo implantation rate. RESULTS: In the EA group, there was a notable decrease in the bilateral pulsatility index and a significant improvement in the endometrial blood flow type post-EA (P<0.05). Both the endometrial blood flow type and VIMV for the endometrium and right ovary were markedly higher in the EA group compared to the WL group post-treatment (P<0.05). Conversely, no significant disparities were observed in vascular index, flow index, vascular blood flow index, uterine arterial blood flow indices, endometrial volume, clinical pregnancy rate and embryo implantation rate between the two groups after treatment (P>0.05). Besides, no adverse events related to EA were observed. CONCLUSIONS: EA can promptly ameliorate VIMV for the endometrial and right ovary, and endometrial blood flow type. Future randomized controlled trials are warranted to investigate the long-term effects of EA on blood flow of RIF patients and its implications for pregnancy outcomes. (Trial registration No. ChiCTR2200057377).

Bushen Huoxue Recipe inhibits endometrial epithelial-mesenchymal transition through the transforming growth factor-ß/nuclear factor kappa-B pathway to improve polycystic ovary syndrome-mediated infertility.

OBJECTIVE: To investigate the target and mechanism of action of Bushen Huoxue Recipe (BSHX) for the treatment of infertility in polycystic ovary syndrome (PCOS), to provide a basis for the development and clinical application of herbal compounds. METHODS: Prediction and validation of active ingredients and targets of BSHX for the treatment of PCOS by using network pharmacology-molecular docking technology. In an animal experiment, the rats were randomly divided into four groups (control group, model group, BSHX group, metformin group, n = 16 in each group), and letrozole combined with high-fat emulsion gavage was used to establish a PCOS rat model. Body weight, vaginal smears, and number of embryos were recorded for each group of rats. Hematoxylin-eosin (HE) staining was used to observe the morphological changes of ovarian and endometrial tissues, and an enzyme-linked immunosorbent assay (ELISA) was used to detect the serum inflammatory factor levels. Expression levels of transforming growth factor-ß (TGF-ß), transforming growth factor beta activated kinase 1 (TAK1), nuclear factor kappa-B (NF-κB), Vimentin, and E-cadherin proteins were measured by western blot (WB). RESULTS: Ninety active pharmaceutical ingredients were obtained from BSHX, involving 201 protein targets, of which 160 were potential therapeutic targets. The active ingredients of BSHX exhibited lower binding energy with tumor necrosis factor-α (TNF-α), TGF-ß, TAK1, and NF-κB protein receptors (< -5.0 kcal/mol). BSHX significantly reduced serum TNF-α levels in PCOS rats (p < .01), effectively regulated the estrous cycle, restored the pathological changes in the ovary and endometrium, improved the pregnancy rate, and increased the number of embryos. The results of WB suggested that BSHX can down-regulate protein expression levels of TGF-ß and NF-κB in endometrial tissue (p < .05), promote the expression level of E-cadherin protein (p < .001), intervene in the endometrial epithelial-mesenchymal transition (EMT) process. CONCLUSIONS: TGF-ß, TAK1, NF-κB, and TNF-α are important targets of BSHX for treating infertility in PCOS. BSHX improves the inflammatory state of PCOS, intervenes in the endometrial EMT process through the TGF-ß/NF-κB pathway, and restores endometrial pathological changes, further improving the pregnancy outcome in PCOS.

Regulation of iron metabolism in HEC-1A endometrium cells by macrophage-derived factors and fractalkine.

Macrophages in the endometrium promote receptivity and implantation by secreting proinflammatory cytokines and other factors like fractalkine (FKN). Macrophages are closely linked to regulating iron homeostasis and can modulate iron availability in the tissue microenvironment. It has been revealed that the iron metabolism of the mother is crucial in fertility. Iron metabolism is strictly controlled by hepcidin, the principal iron regulatory protein. The inflammatory cytokines can modulate hepcidin synthesis and, therefore, the iron metabolism of the endometrium. It was proven recently that FKN, a unique chemokine, is implicated in maternal-fetal communication and may contribute to endometrial receptivity and implantation. In the present study, we investigated the effect of activated THP-1 macrophages and FKN on the iron metabolism of the HEC-1A endometrial cells. We established a noncontact coculture with or without recombinant human FKN supplementation to study the impact of the macrophage-derived factors and FKN on the regulation of hepcidin synthesis and iron release and storage of endometrial cells. Based on our findings, the conditioned medium of the activated macrophages could modify hepcidin synthesis via the nuclear factor kappa-light-chain-enhancer of activated B cells, the signal transducer and activator of transcription 3, and the transferrin receptor 2/bone morphogenetic protein 6/suppressor of mothers against decapentaplegic 1/5/8 signaling pathways, and FKN could alter this effect on the endometrial cells. It was also revealed that the conditioned macrophage medium and FKN modulated the iron release and storage of HEC-1A cells. FKN signaling may be involved in the management of iron trafficking of the endometrium by the regulation of hepcidin. It can contribute to the iron supply for fetal development at the early stage of the pregnancy.

Mechanisms of Bushen Tiaoxue Granules against controlled ovarian hyperstimulation-induced abnormal morphology of endometrium based on network pharmacology.

Bushen Tiaoxue Granules (BTG) is an empirical Chinese herbal formula that has been used for the treatment of subfertility. The protective effect of BTG on controlled ovarian hyperstimulation (COH)-induced impaired endometrial receptivity has been reported in our previous study. This study aims to explore the mechanisms of BTG on ameliorating abnormal morphology of endometrium based on network pharmacology. Active compounds of BTG were identified via the traditional Chinese medicine systems pharmacology and UPLC-MS technology. The SwissTargetPrediction platform and HERB database were used to screen out the putative targets of BTG. Potential targets of endometrial dysfunction caused by COH were obtained from three GEO databases. Through the STRING database, the protein-protein interaction was carried out according to the cross-common targets of diseases and drugs. GO terms and KEGG pathways enrichment analyses were conducted via the Metascape database. AutoDock Vina was used for docking validation of the affinity between active compounds and potential targets. Finally, in vivo experiments were used to verify the potential mechanisms derived from network pharmacology study. A total of 141 effective ingredients were obtained from TCMSP and nine of which were verified in UPLC-MS. Six genes were selected through the intersection of 534 disease related genes and 165 drug potential targets. Enrichment analyses showed that BTG might reverse endometrial dysfunction by regulating adherens junction and arachidonic acid metabolism. Hematoxylin-eosin staining revealed that BTG ameliorated the loose and edematous status of endometrial epithelium caused by COH. The protein expression of FOXO1A, ß-Catenin and COX-2 was decreased in the COH group, and was up-regulated by BTG. BTG significantly alleviates the edema of endometrial epithelium caused by COH. The mechanisms may be related to adheren junctions and activation of arachidonic acid metabolism. The potential active compounds quercetin, taxifolin, kaempferol, eriodictyol, and isorhamnetin identified from the BTG exhibit marginal cytotoxicity. Both high and low concentrations of kaempferol, eriodictyol, and taxifolin are capable of effectively ameliorating impaired hESC cellular activity.

Effect of Acupuncture on Endometrial Blood Flow in Women Undergoing in vitro Fertilization Embryo Transfer: A Single Blind, Randomized Controlled Trial.

OBJECTIVE: To evaluate the effect of manual acupuncture on endometrial blood flow parameters by three-dimensional (3D) power Doppler ultrasound in women undergoing in vitro fertilization embryo transfer (IVF-ET). METHODS: Seventy patients undergoing IVF-ET were equally randomized into traditional or sham acupuncture treatment group for totally 4 days (from the day of oocyte aspiration to the day of embryo transfer) of treatment by random envelope method at the Reproductive Medicine Center and Outpatient Department of Integrated Traditional Chinese and Western Medicine, Tongji Hospital, Tongji Medicine College, Huazhong University of Science and Technology from January 2013 to December 2015. Patients in the traditional acupuncture group accepted traditional acupuncture methods with manual acupuncture, and Zhongji (CV3), Qihai (CV 6), Sanyinjiao (SP6), Taichong (LR 3), Tianshu (ST 25), Guilai (ST 29) and Zusanli (ST 36) were chosen. Patients at the sham acupuncture group accepted shallow acupuncture methods at 4 non-meridian points at each shoulder and upper arm. Outcome measures included endometrial ultrasonic indices such as vascularization index (VI), flow index (FI) and vascularization flow index (VFI), endometrial thickness and volume, subendometrial VI (sVI), subendometrial FI (sFI), subendometrial VFI (sVFI), implantation rate, clinical pregnancy rate, abortion rate, live birth rate and number of live births. RESULTS: Finally, 34 patients in the traditional acupuncture group and 35 in the sham acupuncture group completed this trial. VI, FI and VFI of the traditional acupuncture group were significantly higher than those in the sham acupuncture group (P<0.05). No significant differences were found in endometrial thickness, endometrial volume, sVI, sFI, sVFI, implantation rate, clinical pregnancy rate, abortion rate, live birth rate and number of live births (P>0.05). CONCLUSIONS: Manual acupuncture performed after oocyte aspiration and before transplantation improved the endometrial blood flow parameters VI, RI and VFI in women who underwent IVF-ET, instead of sVI, sFI and sVFI. Therefore, acupuncture might be beneficial in women undergoing IVF-ET by increasing endometrial blood flow and endometrial receptivity. (Registration No. ChiCTR2100053354).

Photobiomodulation therapy at 632 nm wavelength ameliorates intrauterine adhesion via activation of cAMP/PKA/CREB pathway.

Intrauterine adhesion (IUA), a major cause of uterine infertility, is pathologically characterized by endometrial fibrosis. Current treatments for IUA have poor efficacy with high recurrence rate, and restoring uterine functions is difficult. We aimed to determine the therapeutic efficacy of photobiomodulation (PBM) therapy on IUA and elucidate its underlying mechanisms. A rat IUA model was established via mechanical injury, and PBM was applied intrauterinely. The uterine structure and function were evaluated using ultrasonography, histology, and fertility tests. PBM therapy induced a thicker, more intact, and less fibrotic endometrium. PBM also partly recovered endometrial receptivity and fertility in IUA rats. A cellular fibrosis model was then established with human endometrial stromal cells (ESCs) cultured in the presence of TGF-ß1. PBM alleviated TGF-ß1-induced fibrosis and triggered cAMP/PKA/CREB signaling in ESCs. Pretreatment with the inhibitors targeting this pathway weakened PBM's protective efficacy in the IUA rats and ESCs. Therefore, we conclude that PBM improved endometrial fibrosis and fertility via activating cAMP/PKA/CREB signaling in IUA uterus. This study sheds more lights on the efficacy of PBM as a potential treatment for IUA.

Ligustilide enhances pregnancy outcomes via improvement of endometrial receptivity and promotion of endometrial angiogenesis in rats.

Ligustilide (LIG) is the main active ingredient of Angelica sinensis (Oliv.) Diels, which could promote focal angiogenesis to exert neuroprotection. However, there was no report that verified the exact effects of LIG on endometrial angiogenesis and the pregnancy outcomes. To explore the effects of LIG on low endometrial receptivity (LER) and angiogenesis, pregnancy rats were assigned into Control (saline treatment), LER (hydroxyurea-adrenaline treatment), LIG 20 mg/kg and LIG 40 mg/kg groups. Hematoxylin and eosin (H&E) staining was performed to evaluate endometrial morphology. Quantitative real-time PCR, immunofluorescence staining, western blot and immunohistochemistry staining were employed to assess the expression of endometrial receptivity factors and angiogenesis-related gene/protein, respectively. RNA sequencing was used to analyze the effects of LIG on LER caused by Kidney deficiency and blood stasis. We found that endometrial thickness and the implanted embryo number were substantially reduced in the hydroxyurea-adrenaline-treated pregnancy rats. At the same time, the gene and protein expressions of ERα, LIF, VEGFA and CD31 in the endometrium were markedly reduced, while the expressions of MUC1, E-cadherin were increased in the LER group. Administration of LIG raised the endometrial thickness and implanted embryos, as well as reversed the expressions of these factors. Collectively, our findings revealed that LIG could facilitate embryo implantation via recovery of the endometrium receptivity and promotion of endometrial angiogenesis.

The Efficacy of Vitamin D Supplement in the Expression and Protein Levels of Endometrial Decidualization Factors in Women with Recurrent Implantation Failure.

Recurrent implantation failure (RIF) is a challenging situation for infertility specialists, and its treatment is introduced as a difficult case in the field of assisted reproductive technology (ART). Vitamin D (VD) is one of the supplements that have been suggested to improve the implantation process. In the present study, the effect of VD on the expression and protein levels of VD receptor (VDR), progesterone receptor (PR), prolactin (PRL), insulin-like growth factor binding protein-1 (IGFBP-1), and homeobox protein A10 (HOXA10) in the endometrial cells of unknown RIF women with and without VD deficiency were assessed by qRT-PCR and immunohistochemistry. Twelve women with unknown RIF and VD deficiency (≤ 20 ng/ml) and twelve women with unknown RIF without VD deficiency (≥ 30 ng/ml) from 2021 to 2022 were identified. Endometrial specimens were collected in the mid-luteal stage before treatment or pregnancy. In the group with VD deficiency, oral medication of VD 50,000 units was prescribed for 2 to 3 months and their serum levels of VD were re-measured, then an endometrial biopsy at the same stage of the menstrual cycle was performed. The expression and protein levels of VDR, PR, PRL, IGFBP1, and HOXA10 in RIF patients with VD deficiency were lower than the RIF patients without VD deficiency (P value < 0.05). Our findings suggest that VD can play a key role in the pregnancy process, especially during embryo implantation and decidualization of the endometrial cells.IRCT registration number: IRCT20220528055006N1, Registration date: 2022-10-15, Registration timing: retrospective.

E2-Loaded Microcapsules and Bone Marrow-Derived Mesenchymal Stem Cells with Injectable Scaffolds for Endometrial Regeneration Application.

Bone marrow-derived mesenchymal stem cells (BMSCs) have been recognized as new candidates for the treatment of serious endometrial injuries. However, owing to the local microenvironment of damaged endometrium, transplantation of BMSCs yielded disappointing results. In this study, Pectin-Pluronic® F-127 hydrogel as scaffolds were fabricated to provide three-dimensional architecture for the attachment, growth, and migration of BMSCs. E2 was encapsulated into the W/O/W microspheres to construct pectin-based E2-loaded microcapsules (E2 MPs), which has the potential to serve as a long-term reliable source of E2 for endometrial regeneration. Then, the BMSCs/E2 MPs/scaffolds system was injected into the uterine cavity of mouse endometrial injury model for treatment. At 4 weeks after transplantation, the system increased proliferative abilities of uterine endometrial cells, facilitated microvasculature regeneration, and restored the ability of endometrium to receive an embryo, suggesting that the BMSCs/E2 MPs/scaffolds system is a promising treatment option for endometrial regeneration. Furthermore, the mechanism of E2 in promoting the repair of endometrial injury was also investigated. Exosomes are critical paracrine mediators that act as biochemical cues to direct stem cell differentiation. In this study, it was found that the expression of endometrial epithelial cell (EEC) markers was upregulated in BMSCs treated by exosomes secreted from endometrial stromal cells (ESCs-Exos). Exosomes derived from E2-stimulated ESCs further promoted the expression level of EECs markers in BMSCs, suggesting exosomes released from ESCs by E2 stimulation could enhance the differentiation efficiency of BMSCs. Therefore, exosomes derived from ESCs play paracrine roles in endometrial regeneration stimulated by E2 and provide optimal estrogenic response.

Mechanism of cryptotanshinone to improve endocrine and metabolic functions in the endometrium of PCOS rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Cryptotanshinone is the main bioactive component of Salvia miltiorrhiza, with various mechanisms of action, including antioxidant, anti-inflammatory, cardiovascular protection, neuroprotection, and hepatoprotection. Salvia miltiorrhiza is used clinically by gynecologists in China. AIM OF THE STUDY: Polycystic ovary syndrome (PCOS) has a significant impact on women's quality of life, leading to infertility and reproductive disorders. Hence, this study aims to assess the pharmacological activity of cryptotanshinone in the treatment of PCOS and investigate its therapeutic mechanism. MATERIALS AND METHODS: Human chorionic gonadotropin (HCG) combined with insulin is used to simulate a PCOS-like rat model and attempt to discover the abnormal changes that occur and the means by which the pathway acts in this model. RESULTS: The transcriptome sequencing method is used to identify 292 differential genes that undergo significant changes, of which 219 were upregulated and 73 were downregulated. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the signaling pathways reveals that differential expressed genes are significantly enriched in 23 typical pathways. Estrogen signaling pathways are screened in the cryptotanshinone and model groups, and significant differential changes in Fos, ALOX12, and AQP8 are found. This suggests that these signaling pathways and molecules may be the main signaling targets for regulating the differences in endometrial tissue. CONCLUSION: These results indicate that cryptotanshinone has targets for regulating the proliferation of endometrial tissue via estrogen signaling pathways in PCOS-like rats, providing an experimental basis for the clinical application of cryptotanshinone in the treatment of PCOS.

Oral dydrogesterone versus micronized vaginal progesterone for luteal phase support: a double-blind crossover study investigating pharmacokinetics and impact on the endometrium.

STUDY QUESTION: How do plasma progesterone (P) and dydrogesterone (D) concentrations together with endometrial histology, transcriptomic signatures, and immune cell composition differ when oral dydrogesterone (O-DYD) or micronized vaginal progesterone (MVP) is used for luteal phase support (LPS)? SUMMARY ANSWER: Although after O-DYD intake, even at steady-state, plasma D and 20αdihydrodydrogesterone (DHD) concentrations spiked in comparison to P concentrations, a similar endometrial signature was observed by histological and transcriptomic analysis of the endometrium. WHAT IS KNOWN ALREADY: O-DYD for LPS has been proven to be noninferior compared to MVP in two phase III randomized controlled trials. Additionally, a combined individual participant data and aggregate data meta-analysis indicated that a higher pregnancy rate and live birth rate may be obtained in women receiving O-DYD versus MVP for LPS in fresh IVF/ICSI cycles. Little data are available on the pharmacokinetic (PK) profiles of O-DYD versus MVP and their potential molecular differences at the level of the reproductive organs, particularly at the endometrial level. STUDY DESIGN, SIZE, DURATION: Thirty oocyte donors were planned to undergo two ovarian stimulation (OS) cycles with dual triggering (1.000 IU hCG + 0.2 mg triptorelin), each followed by 1 week of LPS: O-DYD or MVP, in a randomized, cross-over, double-blind, double-dummy fashion. On both the first and eighth days of LPS, serial blood samples upon first dosing were harvested for plasma D, DHD, and P concentration analyses. On Day 8 of LPS, an endometrial biopsy was collected for histologic examination, transcriptomics, and immune cell analysis. PARTICIPANTS/MATERIALS, SETTING, METHODS: All oocyte donors were <35 years old, had regular menstrual cycles, no intrauterine contraceptive device, anti-Müllerian hormone within normal range and a BMI ≤29 kg/m2. OS was performed on a GnRH antagonist protocol followed by dual triggering (1.000 IU hCG + 0.2 mg triptorelin) as soon as ≥3 follicles of 20 mm were present. Following oocyte retrieval, subjects initiated LPS consisting of MVP 200 mg or O-DYD 10 mg, both three times daily. D, DHD, and P plasma levels were measured using liquid chromatography-tandem mass spectrometry. Histological assessment was carried out using the Noyes criteria. Endometrial RNA-sequencing was performed for individual biopsies and differential gene expression was analyzed. Endometrial single-cell suspensions were created followed by flow cytometry for immune cell typing. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 21 women completed the entire study protocol. Subjects and stimulation characteristics were found to be similar between groups. Following the first dose of O-DYD, the average observed maximal plasma concentrations (Cmax) for D and DHD were 2.9 and 77 ng/ml, respectively. The Cmax for D and DHD was reached after 1.5 and 1.6 h (=Tmax), respectively. On the eighth day of LPS, the first administration of that day gave rise to a Cmax of 3.6 and 88 ng/ml for D and DHD, respectively. For both, the observed Tmax was 1.5 h. Following the first dose of MVP, the Cmax for P was 16 ng/ml with a Tmax of 4.2 h. On the eighth day of LPS, the first administration of that day showed a Cmax for P of 21 ng/ml with a Tmax of 7.3 h. All 42 biopsies showed endometrium in the secretory phase. The mean cycle day was 23.9 (±1.2) in the O-DYD group versus 24.0 (±1.3) in the MVP group. RNA-sequencing did not reveal significantly differentially expressed genes between samples of both study groups. The average Euclidean distance between samples following O-DYD was significantly lower than following MVP (respectively 12.1 versus 18.8, Mann-Whitney P = 6.98e-14). Immune cell profiling showed a decrease of CD3 T-cell, γδ T-cell, and B-cell frequencies after MVP treatment compared to O-DYD, while the frequency of natural killer (NK) cells was significantly increased. LIMITATIONS, REASONS FOR CAUTION: The main reason for caution is the small sample size, given the basic research nature of the project. The plasma concentrations are best estimates as this was not a formal PK study. Whole tissue bulk RNA-sequencing has been performed not correcting for bias caused by different tissue compositions across biopsies. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study comparing O-DYD/MVP, head-to-head, in a randomized design on a molecular level in IVF/ICSI. Plasma serum concentrations suggest that administration frequency is important, in addition to dose, specifically for O-DYD showing a rapid clearance. The molecular endometrial data are overall comparable and thus support the previously reported noninferior reproductive outcomes for O-DYD as compared to MVP. Further research is needed to explore the smaller intersample distance following O-DYD and the subtle changes detected in endometrial immune cells. STUDY FUNDING/COMPETING INTEREST(S): Not related to this work, C.Bl. has received honoraria for lectures, presentations, manuscript writing, educational events, or scientific advice from Abbott, Ferring, Organon, Cooper Surgical, Gedeon-Richter, IBSA, and Merck. H.T. has received honoraria for lectures, presentations, manuscript writing, educational events, or scientific advice from Abbott, Ferring, Cooper Surgical, Gedeon-Richter, Cook, and Goodlife. S.M. has received honoraria for lectures, presentations, educational events, or scientific advice from Abbott, Cooper Surgical, Gedeon-Richter, IBSA, and Merck and Oxolife. G.G. has received honoraria for lectures, presentations, educational events, or scientific advice from Merck, MSD, Organon, Ferring, Theramex, Gedeon-Richter, Abbott, Biosilu, ReprodWissen, Obseva, PregLem, Guerbet, Cooper, Igyxos, and OxoLife. S.V.-S. is listed as inventor on two patents (WO2019115755A1 and WO2022073973A1), which are not related to this work. TRIAL REGISTRATION NUMBER: EUDRACT 2018-000105-23.

Transcriptomic Changes in Response to Form of Selenium on the Interferon-Tau Signaling Mechanism in the Caruncular Tissue of Beef Heifers at Maternal Recognition of Pregnancy.

We have reported that selenium (Se) provided to grazing beef cattle in an inorganic (ISe) form versus a 1:1 mixture (MIX) of inorganic and organic (OSe) forms affects cholesterol biosynthesis in the corpus luteum (CL), the abundance of interferon tau (IFNτ) and progesterone (P4)-induced mRNAs in the caruncular (CAR) tissue of the endometrium, and conceptus length at maternal recognition of pregnancy (MRP). In this study, beef heifers were supplemented with a vitamin-mineral mix containing 35 ppm Se as ISe or MIX to achieve a Se-adequate status. Inseminated heifers were killed at MRP (d 17, n = 6 per treatment) for tissue collection. In CAR samples from MIX versus ISe heifers, qPCR revealed that mRNA encoding the thyroid regulating DIO2 and DIO3 was decreased (p < 0.05) and a complete transcriptomic analysis revealed effects on the interferon JAK-STAT1/2 pathway, including decreased expression of mRNAs encoding the classical interferon stimulated genes IFIT1, IFIT2, IFIT3, IRF1, IRF9, ISG15, OAS2, and RSAD2 (p < 0.05). Treatment also affected the abundance of mRNAs contributing to the immunotolerant environment (p < 0.05). In combination, these findings suggest more advanced preparation of the CAR and developing conceptus for implantation and to evade immune rejection by the maternal system in MIX- vs. ISe-treated heifers.

Synergistic effects of electroacupuncture and bone marrow mesenchymal stem cell transplantation on rat intrauterine adhesion： an observation. / 电针联合骨髓间å è´¨å¹²ç»†èƒžç§»æ¤æ²»ç–—å¤§é¼ å®«è ”ç²˜è¿žçš„ååŒæ•ˆåº”è§‚å¯Ÿ.

OBJECTIVES: To investigate the effects of electroacupuncture(EA) combined with bone marrow mesen-chymal stem cells(BMSCs) transplantation on the endometrium of rats with intrauterine adhesions(IUA), so as to explore the possible mechanisms underlying their combined therapeutic effects. METHODS: Forty adult female SD rats were randomly divided into control, model, cell, and combined groups. The IUA rat model was established using a dual injury method of mechanical scratching and lipopolysaccharide infection. After successful modeling, on days 1, 3, and 7, rats in the model group received tail vein injection of phosphate buffered solution, while rats in the cell group received tail vein injection of BMSCs suspension for BMSCs transplantation；and rats in the combined group received BMSCs transplantation combined with EA treatment (2 Hz/15 Hz, 1-2 mA), targeting the "Guanyuan"(CV4), bilateral "Zusanli"(ST36) and "Sanyinjiao"(SP6) for 20 min daily for 3 consecutive estrous cycles. After intervention, uterine tissue was collected from 5 rats in each group. Histological analysis was performed using hematoxylin and eosin staining to evaluate endometrial thickness and glandular number. Masson staining was used to assess endometrial fibrosis area. Immunohistochemistry was performed to detect the positive expressions of vascular endothelial growth factor(VEGF), proliferating cell nuclear antigen(PCNA), and estrogen receptor(ER). Western blot analysis was conducted to determine the protein expressions of homeobox A10(HoxA10) and leukemia inhibitory factor(LIF), both key regulators of endometrial receptivity. The remaining 5 rats in each group were co-housed with male rats, and the uterine function recovery was evaluated by assessing the number of embryo implantations. RESULTS: Compared with the control group, the model group showed thinning endometrium(P<0.001), decreased glandular number(P<0.001), increased endometrial fibrosis area(P<0.001), reduced positive expressions of VEGF, PCNA, ER, expressions of HoxA10 and LIF, and decreased embryo implantation number (P<0.001) on the injured side of the uterus. Compared with the model group, the combined group showed a reversal of the aforementioned indicators(P<0.001, P<0.01)；the cell group exhibited thicker endometrium(P<0.001) and reduced endometrial fibrosis area(P<0.001). Compared with the cell group, the combined group showed increased endometrial thickness(P<0.01), elevated glandular number(P<0.05), significantly decreased endometrial fibrosis area(P<0.05), enhanced positive expressions of VEGF, PCNA and ER, expressions of HoxA10 and LIF in the endometrium, and a significant increase in embryo implantation number (P<0.001, P<0.05, P<0.01) on the injured side of the uterus, indicating better results than the cell group. CONCLUSIONS: The combination of EA and BMSCs synergistically promotes the repair of damaged endometrium, improves endometrial morphology, reduces fibrosis levels, enhances vascular regeneration and matrix cell proliferation, improves endometrial receptivity, which ultimately facilitates embryo implantation.

Traditional Chinese medicine Dingkun pill to increase fertility in women with a thin endometrium-a prospective randomized study.

Objective: The aim of this study is to optimize the treatment methods of infertility, which is suggested to be mainly caused by thin endometrium, using a special form of traditional Chinese medicine, the Dingkun pill (DKP), to increase the beneficial endometrial effect of conventional hormone/progestogen therapy. Methods: A total of 307 patients visiting our specialized gynecological endocrinology department because of infertility, which we suggested to be caused by thin endometrium [endometrial thickness (EMT) < 7 mm], were randomly assigned to the experimental group and the control group. The experimental group was treated with estradiol + sequential dydrogesterone + DKP (every day); the control group received hormonal treatment without the Chinese medicine. All patients were monitored in terms of follicle diameter, EMT, and endometrial type every 2 days from the 8th to the 10th day of the menstrual cycle until ovulation day during three menstrual cycles. Serum progesterone levels on 7-8 days after ovulation were measured, and the cumulative pregnancy rate during three menstrual cycles between the two groups was compared. Results: EMT on ovulation day in the experimental group was significantly higher than that in the control group (7.88 vs. 7.15 mm; p < 0.001). The proportion of type A and type B endometrium in total was significantly higher in the experimental group than that in the control group (83.2% vs. 77.7%; p < 0.05). Progesterone levels were significantly higher in the experimental group than those in the control group (10.874 vs. 10.074 ng/mL; p < 0.001). The cumulative pregnancy rate, the main outcome of the study, was significantly higher in the experimental group than that in the control group (29.2% vs. 15.7%; p < 0.05). Conclusion: DKP added to conventional estrogen/progestogen therapy can significantly improve EMT and luteal function in patients attending due to infertility. Because this regimen increased the cumulative pregnancy rate in our study, we conclude that DKP can be used to increase the so-called "thin endometrium infertility".

[Electroacupuncture combined with bone marrow mesenchymal stem cell transplantation promotes repair of thin endometrium by regulating SDF-1/CXCR4 signaling].

OBJECTIVE: It is to explore, based on stromal cell derived factor 1 (SDF-1)/CXC chemokine receptor 4 (CXCR4) signal axis, whether the electroacupuncture (EA) combined with bone marrow mesenchymal stem cells (BMSCs) transplantation can promote thin endometrium regeneration and improve endometrial receptivity, so as to further study its mechanisms underlying improvement of promoting BMSCs homing to repair thin endometrium. METHODS: Thirty matured female SD rats were randomly divided into normal control , model , BMSCs transplantation (BMSCs), BMSCs+AMD3100 (a specific antagonist of CXCR4, BMSCs+AMD3100), BMSCs+EA, and BMSCs+EA+AMD3100 groups, with 5 rats in each group. The thin endometrial model was established by intrauterine injection of 95% ethanol during the period of estrus. Rats of the model group received intravenous injection of PBS solution (tail vein) on day 1, 3 and 7 of modeling and intraperitoneal injection of normal saline once daily for 3 estrous cycles. Rats of the BMSCs group received intravenous injection of BMSCs suspension on day 1,3 and 7 of modeling, and those of the BMSCs+EA group received BMSCs transplantation and EA stimulation. EA (2 Hz/15 Hz, 1 mA) was applied to "Guanyuan" (CV4) and bilateral "Sanyinjiao"(SP9), "Zigong" (EX-CA1) for 15 min, once daily for 3 estrous cycles. Rats of the BMSCs+AMD3100 group received intravenous injection of BMSCs suspension (1×106/mL) and intraperitoneal injection of AMD3100 (5 mg/kg), and those of the BMSCs+EA+AMD3100 group received administration of BMSCs, AMD3100 and EA, with both groups being once daily for 3 estrous cycles. H.E. staining was used to observe histopathological changes of endometrium tissues, and immunohistochemistry was used to detect the expressions of cytokeratin (CK19) and vimentin in endometrium (for evaluating the damage and repair of endometrium). The expression levels of homeobox A10 (HOXA10), leukemia inhibitory factor (LIF), SDF-1 and CXCR4 proteins were detected by Western blot, and those of SDF-1 and CXCR4 mRNAs in the endometrium detected by real-time PCR. RESULTS: In comparison with the normal control group, the number of endometrial glands, the immunoactivity of CK19 and vimentin, the expression leve-ls of HOXA10, LIF and CXCR4 proteins and CXCR4 mRNA were significantly down-regulated (P<0.01), and the expression levels of SDF-1 protein and mRNA significantly up-regulated (P<0.05) in the model group. Compared with the model group, the number of endometrial glands, the immunoactivity of CK19 and vimentin, and the expression levels of HOXA10, LIF, CXCR4 proteins and CXCR4 mRNA in the BMSCs group, and the number of endometrial glands, the immunoactivity of CK19 and vimentin, the expression levels of HOXA10, LIF, CXCR4 proteins and CXCR4 mRNA, and SDF-1 protein and mRNA in the BMSCs+EA group were significantly up-regulated (P<0.05, P<0.01). Compared to the BMSCs group, the number of endometrial glands, and the expression levels of LIF, CXCR4 proteins and CXCR4 mRNA in the BMSCs+EA group were up-regulated (P<0.01, P<0.05)； the number of endometrial glands, the immunoactivity of CK19 and vimentin, the expression levels of HOXA10, LIF, CXCR4 proteins and CXCR4 mRNA in the BMSCs+AMD3100 group were down-regulated (P<0.01). Compared to the BMSCs+EA group, the number of endometrial glands, the immunoactivity of CK19 and vimentin, the expression levels of HOXA10, LIF, CXCR4 proteins and CXCR4 mRNA in the BMSCs+EA+AMD3100 group were down-regulated (P<0.01). Results of H.E. staining showed thin endometrium with absence of epithelial cells, and sparse glands and blood vessels, with smaller glandular cavity in the model group, which was relative milder in BMSCs and BMSCs+EA groups. CONCLUSION: EA can promote the transfer of transplanted BMSCs to the damaged site through SDF-1/CXCR4 signaling related stem cell homing, thereby promoting thin endometrial regeneration, repairing endometrial injury, and improving endometrial tolerance in rats with thin endometrium.

Hyperbaric oxygen therapy: a possible choice for patients with resistant thin endometrium during frozen embryo transfer treatments.

BACKGROUND: Thin endometrium is considered suboptimal for embryo implantation, leading to compromised pregnancy rates without effective therapies. While some studies have reported promoted endometrial growth after a period of hyperbaric oxygen therapy (HBOT) in patients with intrauterine adhesion, there have been no reports in patients with resistant thin endometrium. The purpose of this study was to investigate the impact of HBOT on endometrium growth and pregnancy outcomes in patients with resistant thin endometrium during frozen embryo transfer (FET) treatments. METHODS: This prospective pre-post cohort study was conducted at a university-affiliated assisted reproductive medical center between October 2021 and December 2022. Patients who had experienced at least one canceled transfer cycle due to a thin endometrium(< 7 mm) on the endometrium transformation day, despite the use of standard therapies as well as adjuvant therapies, were enrolled in the study. Patients were assigned voluntarily to either the HBOT group or the concurrent control group. The HBOT group received daily HBOT for at least 10 days during the proliferative phase, in addition to the routine endometrium preparation methods and the concurrent control group underwent cycles without HBOT. Propensity score matching (PSM) was used to ensure comparability between the groups. Both self-control and case-control comparisons were conducted. The primary outcome measured was endometrial thickness (ET) on the day of endometrium transformation. Secondary outcomes included intrauterine pregnancy rate (IPR), embryo implantation rate (IR), miscarriage rate, and others. RESULTS: Patients in the HBOT group demonstrated a significantly thicker endometrial thickness on the day of endometrium transformation after undergoing therapy (5.76 ± 1.66 vs. 6.57 ± 1.23, P = 0.002). This improvement was accompanied by a decreased rate of cycle cancellations. Baseline parameters and endometrial thickness were comparable between the HBOT group and the concurrent control group during the cycle. The IPR was similar in patients who received cleavage-stage embryos (0.0% vs. 6.7%, P = 1.00), but significantly higher in patients in the HBOT group who received blastocysts (53.8% vs. 18.2%, P = 0.017). CONCLUSIONS: A period of HBOT prior to endometrium transformation contributes to increased endometrial thickness and facilitates blastocyst implantation in patients with resistant thin endometrium during FET treatments. TRIAL REGISTRATION: The trial was registered on the Chinese Clinical Trial Registry (registration no. ChiCTR2300072831, retrospectively registered).

Effect of Chinese patent medicine Kunling Pill on endometrial receptivity: A clinical trial, network pharmacology, and animal-based study.

Although pregnancy success rates are raised with assisted reproductive technology, it still cannot meet clinical demands. Kunling Pill (KLP), a traditional Chinese medicine, is widely used in various gynecological disorders, particularly in improving fertility and pregnancy rates. However, the underlying mechanism of how KLP affects pregnancy outcomes remains unclear. This study aimed to explore the effects and mechanisms of KLP on endometrial receptivity. Firstly, a retrospective trial was conducted to validate the efficacy of KLP on repeated implantation failure (RIF) patients. The result indicated a significant increase in the proportion of live birth in KLP group (30.56%) compared to the control group (16.89%). Secondly, network pharmacology methods predicted the active components and network targets of KLP. Endometrial receptivity is closely associated with the activation of inflammatory factors, predicting the function of KLP on the immune system. The estrogen and apoptotic signaling pathways were also highlighted in the gene ontology enrichment analysis. Thirdly, a decreased endometrial receptivity model was established by controlled ovarian hyperstimulation (COH) in female C57BL/6 mice, divided into the COH and KLP groups. Normal female mice are as control group. In vivo, KLP administration could increase endometrial thickness and the number of endometrial glands and pinopodes. In the endometrium, KLP supplementation upregulated the expressions of estrogen receptor α, progesterone receptor, endothelial nitric oxide synthase, and integrin αVß3 in the murine uterus and reduced serum levels of estrogen and progesterone. KLP regulated the uterine immune cells and inhibited cell apoptosis in the ovary via Bcl-2/Bax/caspase-3 pathway. In conclusion, KLP administration raised the live birth rate in RIF patients to optimize medication regimens, mainly because KLP ameliorated impaired endometrial receptivity.

A prospective study on the endometrium properties and the pregnancy rate of infertile women with thin endometrium receiving Fructus ligustri lucidi used in traditional Chinese medicine.

OBJECTIVE: To evaluate whether Zi Gui Nv Zhen capsules (ZGNZC) can increase the fertility rate of Chinese women with infertility due to thin endometrium. METHODS: Prospective, randomized, open-labeled 3-monthly study; 104 patients (aged 20-40 years) receiving either ZGNZC (experimental group, n = 55) or not (control group, n = 49). Main outcomes: thickness/type of the endometrium during ovulation and pregnancy rate. Between-group analysis (A) compares the experimental vs. control group, and within-group analysis (B) compares data at baseline and after study in the experimental group. RESULTS: (A) Between-group-analysis: Patients with ZGNZC had a higher endometrium thickness (0.8 ± 0.27 vs. 0.68 ± 0.10; p < .05) and higher type A endometrium rates (34.2% vs. 13.2%; p < .05) than the control group. Pregnancy rates were higher in the experimental than in the control group (43.6% vs. 22.4%; p < .05). (B) Within-group-analysis: ZGNZC increased endometrium thickness (0.58 ± 0.13 vs. 0.87 ± 0.24 vs. 0.83 ± 0.26 vs. 0.80 ± 0.27), and type A endometrium rates (10.9% vs. 60.0% vs. 49.0% vs. 34.2%) (all p < .05). Univariate analysis of pregnancy with other study parameters showed positive and significant correlations between pregnancy and administration of ZGNZC (p < .05). All hepato-renal biomarkers remained within the norm. There were no adverse events. CONCLUSIONS: In infertile women with thin endometrium who wish to conceive, two months' application of ZGNZC can improve endometrial properties and proliferation, which is necessary for a healthy pregnancy, and increase the clinical pregnancy rate in our prospective randomized observational study.

Cuyun Recipe ameliorates pregnancy loss by regulating macrophage polarization and hypercoagulable state during the peri-implantation period in an ovarian hyperstimulation mouse model.

BACKGROUND: The Chinese herbal prescription Cuyun Recipe (CYR) has been widely used to treat clinical infertility and has shown good efficacy. Animal experiments have shown that CYR can promote implantation in mice, however, the exact mechanism underlying the implantation has not been elucidated. PURPOSE: To investigate the effect and mechanism of CYR on regulating macrophage polarization and hypercoagulability during the peri-implantation period in mice with ovarian hyperstimulation. METHODS: An ovarian hyperstimulation mouse model was developed, followed by treatment with CYR. Mice were sacrificed on day (D)4.5, D6, or D8 of gestation. The number of implantation sites, the pathological changes of the uterus and ovaries were assessed. The polarization of monocytes/macrophages in the spleen and endometrium, the expression and localization of cytokines were further detected. Furthermore, analyses of hypercoagulable state of the blood were also performed. RESULTS: Treatment with CYR increased the average number of implantation sites, promoted angiogenesis in endometrial, and regulated monocytes/macrophages and the cytokine levels. Moreover, CYR downregulated the overexpression of D-dimer and fgl2 after ovarian hyperstimulation. CONCLUSION: CYR facilitates embryo implantation by alleviating ovarian hyperstimulation, promoting endometrial decidualization and angiogenesis, regulating macrophage polarization, and reversing the hypercoagulable state of the blood.

Endometriosis Treatment: Role of Natural Polyphenols as Anti-Inflammatory Agents.

Endometriosis is an estrogen-dependent common chronic inflammatory disease defined by the presence of extrauterine endometrial tissue that promotes pelvic pain and fertility impairment. Its etiology is complex and multifactorial, and several not completely understood theories have been proposed to describe its pathogenesis. Indeed, this disease affects women's quality of life and their reproductive system. Conventional therapies for endometriosis treatment primarily focus on surgical resection, lowering systemic levels of estrogen, and treatment with non-steroidal anti-inflammatory drugs to counteract the inflammatory response. However, although these strategies have shown to be effective, they also show considerable side effects. Therefore, there is a growing interest in the use of herbal medicine for the treatment of endometriosis; however, to date, only very limited literature is present on this topic. Polyphenols display important anti-endometriotic properties; in particular, they are potent phytoestrogens that in parallel modulates estrogen activity and exerts anti-inflammatory activity. The aim of this review is to provide an overview on anti-inflammatory activity of polyphenols in the treatment of endometriosis.