Endometrial Implants Research Articles

Structural and functional features of the endometrium in chronic endometritis and implantation failure in IVF cycles

BACKGROUND: Implantation failure of the endometrium in women with chronic endometritis remains a subject of discussion, despite certain progress in the diagnosis and treatment of this pathology. It has been proven that chronic endometritis is a predictor of endometrial dysfunction, which contributes to the disruption of the morphogenesis of the endometrium necessary for its implantation capacity. Implantation disorders associated with chronic endometritis are related to pathology of the receptor profile and vascular bed, as well as dysregulation of proliferative and apoptotic mechanisms. AIM: The aim of this study was to evaluate the expression of estrogen and progesterone receptors, endothelial marker and apoptosis-inducing factor in the endometrium of patients with chronic endometritis and ineffective embryo transfer in IVF protocols. MATERIALS AND METHODS: This study enrolled 50 patients with ineffective embryo transfers in IVF protocols and 25 women in the control group with positive reproductive outcomes. All patients underwent a comprehensive morphological study of the endometrium including histological examination, immunohistochemical study with assessment of the receptor profile [estrogen (ERα) and progesterone (PR) receptor expression], chronic endometritis (CD8+, CD4+, CD20+, CD138+), apoptosis [antibodies to apoptosis-inducing factor (anti-AIF [E20])], and angiogenesis (CD34+), with microscopic, morphometric and statistical methods used as well. RESULTS: Endometrial implantation failure is associated with a history of urogenital infection, benign cervical diseases, high incidence of endometrial pathology and reproductive loss. It is characterized by the presence of follicle-like mononuclear infiltrates in the endometrium, fibroplastic changes in the stromal component, vascular sclerosis, abnormal increase in the pro-inflammatory cell count, multifocal decrease in estrogen (ERα) and progesterone (PR) receptor expression, as well as activation of pathological neoangiogenesis and apoptosis in the stroma. CONCLUSIONS: Chronic endometritis leads to disruption of the structural and functional characteristics of the endometrium, stimulates pathological neoangiogenesis and dysregulates proliferative and apoptotic mechanisms, which forms a vicious circle of endometrial implantation failure and inefficiency of embryo transfer in IVF protocols.

Papillomavirus infection and male infertility: A systematic review and meta-analysis.

Increasing attention is being paid to the role of human papillomavirus (HPV) in men and specifically reproduction. Growing evidence suggests an association between HPV infection with many adverse effects including the impairment of semen parameters, the increase of blastocyst apoptosis, the reduction of endometrial implantation of trophoblastic cells, as well as the increase rate of miscarriages and spontaneous preterm birth. We systematically searched PubMed/MEDLINE, Scopus, Embase, Web of Science, CINHAL, PsycINFO, and ERIC from inception to 2nd of July 2024, for studies that investigated the association between HPV infection with sperm parameters and fertility outcomes. The meta-analysis was conducted on mean data and standard deviations. We included 25 studies with a total of 6942 patients. Sperm morphology was lower in HPV positive groups versus HPV negative control groups (SMD = -0.52 95% CI -0.84; -0.21; p = 0.001). Sperm motility was also significantly lower in HPV positive groups when compared to HPV negative controls (SMD = -0.82 95% CI -1.07; -0.57; p = <0.001). Sperm volume, concentration, and pH were not significantly different between the two groups. The other 15 studies included in the systematic review for which it was not possible to conduct a meta-analysis showed strong associations between HPV infection and impairment of sperm parameters, reduced couple fertility and increased risk of pregnancy loss. The current evidence highlights the link between HPV infection and sperm parameters, male fertility and reproductive outcomes, which has the potential to lead to a decreased couple fertility, increased risk of pregnancy loss, re-infection and increased treatment costs.

Endometriosis: A Comprehensive Exploration of Inflammatory Mechanisms and Fertility Implications.

Endometriosis is a prevalent gynecological disorder characterized by the ectopic growth of endometrial-like tissue outside the uterus. This condition poses significant challenges due to its chronic nature, debilitating symptoms such as pelvic pain and infertility, and substantial impact on quality of life. Central to the pathogenesis of endometriosis are inflammatory mechanisms that perpetuate tissue proliferation, adhesion formation, and immune dysregulation within the pelvic cavity. Inflammation plays a pivotal role in the development and progression of endometriosis, influencing the severity of symptoms and complications associated with the disease. Dysregulated immune responses contribute to the persistence of ectopic endometrial implants, exacerbating pelvic pain and other symptoms experienced by affected individuals. Moreover, the inflammatory milieu created by endometriotic lesions disrupts normal ovarian function, impairs follicular development, and compromises reproductive outcomes, thereby posing challenges to fertility. This review comprehensively explores the inflammatory mechanisms underlying endometriosis and their implications for fertility. Synthesizing current research and clinical insights elucidates the intricate interplay between inflammation, disease progression, and reproductive health outcomes. Understanding these complex interactions is essential for developing targeted diagnostic strategies and optimizing therapeutic approaches tailored to alleviate symptoms and improve fertility outcomes in individuals with endometriosis. Ultimately, this review aims to enhance the understanding of endometriosis pathophysiology, inform clinical practice, and stimulate further research to advance personalized care and management strategies for this challenging condition.

Immunological testing for female infertility: a modern view on the problem

Introduction. According to various estimates, in Russia 10 to 20 % of people of reproductive age are infertile. Changes in the immune system play a key role in the etiology and treatment of various infertility forms. The active introduction of immunological testing into clinical practice can potentially improve the results of infertility diagnostics and treatment.Aim: to analyze the current literature data on immunological testing in female infertility, as well as to assess its potential role in infertility diagnostics and treatment.Materials and Methods. There was conducted a search for publications in the electronic databases PubMed and eLibrary by using the following keywords and their combinations: "infertility", "immunology", "immune system", "immunological testing", "diagnostics", "treatment". The articles were evaluated in accordance with the PRISMA recommendations. Ultimately, 88 publications were included in the review.Results. Testing for antiphospholipid antibodies (APA) may be useful for women undergoing assisted reproductive technology (ART) therapy, as these antibodies increase the risk of pregnancy complications and thrombotic risks associated with ovarian stimulation, but studies assessing AFA effect on in vitro fertilization (IVF) outcomes have ambiguous results. The presence of antithyroid antibodies (ATA) may be associated with infertility, so their assessment is indeed important to determine treatment tactics. It has been suggested that antinuclear antibodies (ANA) may affect reproductive function by disrupting trophoblast cell development and interfering with RNA transcription, which may lead to lowered reproductive success. Studies have shown that patients with a positive ANA data have a lower incidence of pregnancy and a higher rate of miscarriages after IVF procedure. The human herpes virus type 6 (HHV-6) affects female fertility and is often the cause of spontaneous termination of pregnancy. B-cell lymphoma protein 6 (BCL-6) can serve as an important prognostic biomarker to identify individuals with endometriosis and related reproductive disorders, including idiopathic infertility. Evaluation of endometrial decidualization can be a useful tool to assess readiness for endometrial implantation and provide opportunities for targeted therapeutic interventions. The issue of testing for NK-cells in patients undergoing infertility screening remains controversial due to the difficulties of standardizing testing recommendations.Conclusion. To date, there is a limited number of reliable data on the role of various immunological tests in infertility diagnostics and treatment. With the exception of testing for AFA in patients with RPL, as well as for thyroid-stimulating hormone (TSH) and anti-thyroperoxidase antibodies (TPOAb) in patients undergoing therapy using various types of assisted reproductive technologies, the remaining immunological tests provide scant data to justify their routine use in clinical practice. The major limitations of existing studies are coupled to a small patient sample, as well as heterogeneity of inclusion criteria, patient groups and research methods.

Potential therapeutic properties of broccoli extract and soy isoflavones on improvement endometriosis and involved oxidative parameters.

In Endometriosis is a gynecological disorder characterized by the growth of endometrial tissue outside the uterine cavity that is associated with chronic pelvic pain and subfertility. The purpose of the study was to investigate the effect of broccoli extract (BE) alone and in combination with soy isoflavones (SI) on endometrial implants in female rat. In this study, endometriosis was induced surgically in 40 mature female rats. The rats were divided into 5 groups that were treated by oral gavage for 6weeks with 0.5 mL of saline 0.9 %/day (control group), BE (3,000 mg/kg/day), SI (50 mg/kg/day), BE/soy isoflavones (BE 3000 mg/kg/day+soy isoflavones 50 mg/kg/day) and diphereline as a standard medication (3 mg/kg) intramuscularly. At the end of treatments, the volume and histopathology of the endometrial implants were compared among the 5 groups. The serum levels of oxidative parameters including superoxide dismutase (SOD), malondialdehyde (MDA) and tumor necrosis factor alpha (TNF-α) were also compared between the groups. The volume of the implants significantly decreased in diphereline group (p=0.002). The histopathological grade of endometrial implants in BE/SI and diphereline group were significantly decreased compared to the control group (p=0.001). The serum levels of SOD in BE group were enhanced significantly in comparison to the control group (p=0.034). BE in combination with SI decreased the growth and histopathologic grades of transplanted endometrial implants. These herbal compounds may have the potential therapeutic effect to be used as an alternative medication for the treatment of endometriosis.

P-799 Augmenting endometrial thickness and implantation rates: investigating the efficacy of autologous blood cell derivative - endosera in women struggling with thin endometrium and infertility

Abstract Study question Could autologous platelet derived growth factor based strategies promote endometrial growth in women with refractory thin endometrium resulting in sustained implantation and live births? Summary answer Autologous platelet derived growth factor based strategies remarkably promote endometrial thickness and receptivity in women with refractory thin endometrium and pregnancy outcomes What is known already Repeated implantation failure (RIF) attributed to thin endometrium poses significant challenge in reproductive medicine. In cases where conventional treatments prove ineffective, couples often contemplate stressful option of surrogacy. Regenerative strategies, such as platelet-rich plasma (PRP), exhibit promise by capitalizing on anti-inflammatory and pro-regenerative attributes of platelet-derived growth factors. Recognizing the imperative for anti-inflammatory microenvironment conducive to successful implantation, Endosera, a cell-free growth factors concentrate tailored for implantation has been formulated. This study seeks to evaluate the effectiveness of Endosera in augmenting endometrial thickness and implantation rates, offering potential remedies for RIF and thin endometrium. Study design, size, duration In this prospective self-controlled study 62 women (24-47 years) were enrolled spanning May 2021 to December 2023. Women experienced &amp;gt;2 failed IVF cycles and multiple cycle cancellations due to refractory thin endometrium with poor endometrial receptivity defined as endometrial thickness of &amp;lt; 7mm despite standard medical therapy, abnormal endometrial pattern and suboptimal endometrialvascularity, negative hysteroscopic screening for endometrial pathology, and negative bacteriologicscreening and despite multiple failed conventional therapies were selected to undergo Endosera treatment. Participants/materials, setting, methods After obtaining informed consent, patients underwent three intrauterine infusions of Endosera on menstrual cycle Day 7, Day 12, and 2 days before frozen embryo transfer (FET). Alongside standard hormone replacement therapy (HRT), 0.8 ml of Endosera was introduced into the uterine cavity. Pregnancy confirmation relied on a positive serum β-HCG test two weeks post-ET, while clinical pregnancy was established by detecting fetal heartbeat through transvaginal ultrasound four weeks post-ET. Main results and the role of chance In this study involving 62 patients facing multiple cancelled IVF cycles due to refractory thin endometrium, the administration of three doses of Endosera demonstrated significant positive outcomes. Average endometrial thickness notably increased from 6.2 ± 1.2 mm to 8.4 ± 1.2 mm (P &amp;lt; 0.0001) after the 2nd Endosera dose, reaching an optimal thickness of ≥ 8mm. Among the participants, 43 achieved positive pregnancy result (69.4%), with 26 (41.9%) delivering healthy babies and 7 (11.3%) uneventful ongoing pregnancies. Poor outcomes included for 2 (3.2%) patients cycle was cancelled due to no endometrial improvement 5(8.1%) miscarriages before 10 weeks, while 12 (19.4%) women didn’t conceive and for 4.8%(3) patients results awaited post ET. No adverse effects were reported, indicating safety of Endosera.Cumulative positive pregnancy outcomes recorded to be 53.2% (33). The formulation, standardized to contain 6-9 times higher growth factor levels than peripheral blood, emphasizes implantation-friendly and anti-inflammatory cytokines. This optimized composition aligns with cyclical requirements of the endometrial phases (proliferation, secretion, and implantation), creating an angiogenic and anti-inflammatory environment crucial for successful implantation. Endosera’s safety, reproducibility, and efficacy in mimicking natural tissue repair and regeneration processes position it as promising solution for refractory thin endometrium-related challenges. Limitations, reasons for caution This study, though demonstrating improved outcomes in refractory thin endometrium cases with Endosera treatment, acknowledges limitations such as a small sample size and absence of a randomized control group. To establish Endosera as a routine adjuvant in IVF, larger study with stringent inclusion criteria for refractory thin endometrium is recommended. Wider implications of the findings Endosera demonstrated notable success with significant improvement in implantation, pregnancy, and live birth rates for refractory thin endometrium patients prompts rigorous randomized controlled studies. These will include diverse populations, validating results and offering patients hope to reduce financial and psychological stress and challenges associated with cycle cancellations and surrogacy. Trial registration number Not Applicable

P-329 Evaluating Implansera, autologous platelet-derived growth factors, to improve clinical pregnancy and live birth rates in frozen embryo transfer for women with recurrent implantation failure

Abstract Study question Can Implansera, Autologous Platelet-Derived Growth Factors, serve as a promising adjuvant therapy in assisted reproductive techniques to enhance endometrial thickness and implantation rates? Summary answer Use of Implansera significantly increased clinical pregnancy and Live Birth Rate In Patients With repeated implantation failure during FET cycles. What is known already Insufficient endometrial thickness and receptivity contribute to Recurrent Implantation Failure (RIF), prompting consideration of surrogacy. Despite global rise in Frozen Embryo Transfer (FET), quest for an optimal implantation protocol persists. Recent research highlights platelet-rich plasma (PRP) as promising solution for thin endometrium and RIF cases, attributing positive outcomes to PRP’s high growth factor concentration. Recognizing the need for precise platelet and growth factor concentrations, Implansera—a specialized platelet-derived growth factor concentrate—was developed. This innovation addresses the challenge of unresponsive endometrium, offering tailored solution to maximize therapeutic efficacy and improve pregnancy outcomes for patients facing RIF issues. Study design, size, duration Conducted from May 2021 to December 2023, this prospective interventional self-controlled study focused on 21 women (aged 30-48) experiencing recurrent IVF failure due to stubborn thin endometrium. Selected after negative hysteroscopic and bacteriologic screenings, and unsuccessful pregnancies despite multiple immune therapy regimens, such as intralipid and granulocyte colony-stimulating factor infusions, steroids, and endometrial scratching. The subjects underwent Implansera treatment, aiming to address refractory thin endometrium and potentially enhance their chances of successful conception. Participants/materials, setting, methods After obtaining informed consent women with at least two previous unexplained RIF, who were candidates for frozen-thawed embryo transfer were treated with intrauterine infusion of 0.8 ml of Implansera was infused into uterine cavity in addition to standard HRT protocols 2 days before ET. Clinical pregnancy was determined by positive serum β-HCG, 2weeks after ET and presence of fetal heart beat in trans-vaginal ultrasound 5weeks after ET. Main results and the role of chance Among 21 women, 10 achieved pregnancy (47.6%), while 11 did not conceive (52.4%). Clinical outcomes varied: 1 woman (4.8%) experienced chemical pregnancy, 3 (14.3%) miscarried before 6-12 weeks, 3 (14.3%) delivered healthy full-term babies, and 3 (14.3%) are in the 24th week of uneventful gestation accounts for 28.6% cumulative successful pregnancy outcomes. Implansera treatment yielded no reported adverse effects.Implansera, prepared through a patented proprietary process, standardized to contain 6-9 times higher growth factors than peripheral blood, particularly implantation-friendly and anti-inflammatory cytokines, addresses the crucial angiogenic and anti-inflammatory requirements for successful implantation. Tailored for the cyclical demands of endometrial proliferation, secretion, and implantation phases, it efficiently restores impaired uterine environments.Beyond efficacy, Implansera’s safety, reproducibility, and effectiveness in emulating natural tissue repair and regeneration processes are noteworthy. Studies underscore the significance of creating an environment conducive to successful implantation, and Implansera’s composition aligns with these requirements. Its capacity to mimic natural processes positions it as a promising solution, offering a comprehensive approach to enhancing endometrial receptivity, thus optimizing conditions for successful pregnancies. Limitations, reasons for caution This self-controlled prospective study, albeit with small sample size and lacking randomized control group, aligns with published data in demonstrating Implansera’s benefits. Proposing larger studies, particularly with unexplained RIF, to establish Implansera as routine IVF adjuvant. Further investigations aim to identify active molecules influencing implantation and suggest targeted recommendations. Wider implications of the findings Implansera improved implantation, pregnancy, and live birth rates (LBR) in unexplained RIF patients which indicates clearly endometrial thickness and receptivity improvement and motivated us to plan randomized controlled studies to confirm the results and provide opportunity for women with unexplained RIF to conceive and deliver in lesser number of cycles. Trial registration number Not applicable

P-305 Enhancing endometrial thickness and implantation rates: a study on the effectiveness of autologous blood cell derivative - endosera in infertile women with thin endometrium

Abstract Study question Could personalized regenerative medicine based strategies promote endometrial growth and receptivity in women with refractory thin endometrium resulting in successful implantation and live births? Summary answer Personalized regenerative medicine based strategies remarkably promote endometrial growth and receptivity in women with refractory thin endometrium resulting in successful implantation and live births. What is known already Repeated implantation failure (RIF) due to thin endometrium is a formidable challenge in reproductive medicine, often steering couples toward the stressful option of surrogacy. Regenerative approaches like platelet-rich plasma (PRP) have shown promise, leveraging the anti-inflammatory and pro-regenerative properties of platelet-derived growth factors. Recognizing the need for an anti-inflammatory milieu for successful implantation, Endosera, a cell-free implantation-friendly growth factors concentrate, has been developed. This study at Bloom IVF Centre-Mumbai, India, aims to assess Endosera’s efficacy in enhancing endometrial thickness and implantation rates, providing potential solutions for RIF and thin endometrium. Study design, size, duration In this prospective self-controlled study 72 women (27-49 years) were enrolled spanning December 2022 to December 2023. Women experienced two or more failed IVF cycles and multiple cycle cancellations due to refractory thin endometrium, negative hysteroscopic screening for endometrial pathology, and negative bacteriologic screening and despite multiple failed conventional therapies were selected to undergo Endosera treatment. Participants/materials, setting, methods After obtaining informed consent, patients underwent three intrauterine infusions of Endosera on menstrual cycle Day 7, Day 12, and 2 days before frozen embryo transfer (FET). Alongside standard hormone replacement therapy (HRT), Endosera was introduced into the uterine cavity. Pregnancy confirmation relied on a positive serum β-HCG test two weeks post-ET, while clinical pregnancy was established by detecting fetal heartbeat through transvaginal ultrasound four weeks post-ET. Main results and the role of chance In this study involving 72 patients facing recurrent implantation failure due to refractory thin endometrium, the administration of three doses of Endosera demonstrated significant positive outcomes. The average endometrial thickness notably increased from 6.4 ± 1.1 mm to 8.7 ± 1.0 mm (P &amp;lt; 0.0001) after the 2nd Endosera dose, reaching an optimal thickness of ≥ 7mm. Among the participants, 34 achieved a positive pregnancy result (47.2%), with 6 (8.3%) delivering healthy babies and 23 (31.9%) experiencing ongoing clinical pregnancies. Adverse outcomes included 2 (2.8%) ectopic pregnancies and 3 (4.2%) miscarriages before 10 weeks, while 36 (50%) women returned negative HCG results. No adverse effects were reported, indicating the safety of Endosera. The formulation, standardized to contain 6-9 times higher growth factor levels than peripheral blood, emphasizes implantation-friendly and anti-inflammatory cytokines. This optimized composition aligns with the cyclical requirements of the endometrial phases (proliferation, secretion, and implantation), creating an angiogenic and anti-inflammatory environment crucial for successful implantation. Endosera’s safety, reproducibility, and efficacy in mimicking natural tissue repair and regeneration processes position it as a promising solution for refractory thin endometrium-related challenges. Limitations, reasons for caution This study, though demonstrating improved outcomes in refractory thin endometrium cases with Endosera treatment, acknowledges limitations such as small sample size and absence of a randomized control group. To establish Endosera as a routine adjuvant in IVF, a larger study with stringent inclusion criteria for refractory thin endometrium is recommended. Wider implications of the findings Endosera demonstrated notable success with significant improvement in implantation, pregnancy, and live birth rates for refractory thin endometrium patients prompts rigorous randomized controlled studies. These will include diverse populations, validating results and offering patients hope to reduce financial and psychological stress and challenges associated with cycle cancellations and surrogacy. Trial registration number Not applicable

P-014 Extracellular vesicles (EVs) from female reproductive tracts positively affect sperm capacitation and progressive motility

Abstract Study question Do EVs from human follicular fluid (FF-EVs), cervicovaginal fluid (CVF-EVs), and endometrial stromal cells spent medium (ESCs-EVs) induce sperm capacitation and enhance sperm motility? Summary answer In humans, EVs from different female reproductive tracts actively contribute to sperm maturation by inducing capacitation and enhancing sperm progressive motility. What is known already In the human body, EVs represent crucial players in cell-to-cell communication. EVs presence has been detected also in the female and male reproductive systems, specifically in seminal, follicular, oviductal, endometrial and vaginal fluid, as well as in embryo secretions. Moreover, EVs have been shown to be of pivotal importance in the fertilization process, as they support the maturation of spermatozoa and oocytes, as well as embryos in their development and endometrial implantation. The majority of these studies, though, have been conducted in animal models, underscoring the necessity to replicate analogous investigations also in humans. Study design, size, duration Follicular fluid (FF, n = 8), cervicovaginal fluid (CVF, n = 20) and endometrial tissue (n = 8) samples were collected from women undergoing assisted reproductive technology (ART) cycles at San Raffaele Hospital Fertility Centre (Milan). CVF were collected 2 and 7 days after the LH peak (CVF LH + 2 and CVF LH + 7, respectively). ESCs-EVs were isolated from both decidualized (dESCs-EVs) and not decidualized (eESCs-EVs) endometrial cells. Semen samples (n = 15) were collected from normozoospermic men. Participants/materials, setting, methods EVs were isolated using differential centrifugations and characterized using Transmission Electron Microscopy (TEM), Nanoparticle Tracking Analysis (NTA), and western blotting (WB). EVs capture by spermatozoa was assessed using flow cytometry. EVs impact on sperm progressive motility was evaluated following WHO guidelines. EVs potential to induce sperm capacitation was examined through sperm acrosome reaction (AR) induction using a Ca2+ ionophore, followed by Coomassie blue staining. All experiments were performed using EVs at 50 and 100 μg/mL. Main results and the role of chance Isolated EVs displayed comparable size, with mean diameters ranging from 147,9±11,6 nm to 168,1±6,2 nm (mean ± SEM) and average concentrations from 9,53E+10 ± 8,11E+09 to 2,70E+11 ± 3,49E+10 particles/ml (mean ± SEM). TEM analysis confirmed their round-shaped morphology and structural integrity, while WB confirmed their positivity for EVs markers (Alix, TSG101, CD63, CD9). Spermatozoa efficiency in capturing these fluorescently-labeled EVs, measured as the percentage of fluorescence-positive spermatozoa, reached up to 3,6±1,1% for FF-EVs, 12,1±3,7% for CVF-EVs and 9,4±1,9% for ESCs-EVs (mean ± SEM). Sperm cells co-incubated with any EVs group maintained unaffected vitality. Following a 4-hour incubation with EVs, a rise in the proportion of progressively motile spermatozoa was observed, with percentages of progressively motile spermatozoa ranging from a minimum of 45,4±3,8% for CVF-EVs LH + 7 to a maximum of 52,7±2,7% for dESCs-EVs (mean ± SEM). This increase resulted to be statistically significant (p &amp;lt; 0.05 and p &amp;lt; 0.01, respectively) compared to the untreated (UT) condition, which exhibited 26,2±0,7% of progressively motile sperm cells. Spermatozoa that completed the AR, indicating capacitation, after 4-hour incubation with EVs resulted to be up to 25,8±3,2% for FF-EVs, 35,0±6,6% for CVF-EVs and 26,9±3,0% for ESCs-EVs, compared to an average of 13% in the UT control. Limitations, reasons for caution This study constitutes a pilot investigation characterized by a limited sample size, necessitating larger cohorts to validate results. Wider implications of the findings Studied EVs may exert beneficial effects on capacitation and progressive motility also on asthenozoospermic (AS) men’s spermatozoa. Therefore, they could be used in ART cycles to prime spermatozoa from AS men and increase the percentage of sperm cells displaying progressive motility, potentially favouring fertilization via classical IVF instead of ICSI. Trial registration number not applicable

P-397 A retrospective analysis shows a high effect in reproductive outcomes as age increases associated to endometrial factor in IVF advanced maternal age (AMA) patients

Abstract Study question Is there an increasing risk of a negative reproductive outcomes with every passing year associated with endometrial factor in AMA patients? Summary answer The risk of implantation failure and pregnancy loss associated with endometrial factor increase in 2.0% and 1.2% respectively with each passing year in AMA patients. What is known already Nowadays, there is a delay in maternity due to lifestyle, leading to a progressive increase in the age of women at the first childbirth increasing infertility. However, no universal definition of AMA has been established yet. Typically, studies are performed between different groups of age, establishing arbitrary thresholds. Besides, studies about AMA infertility mainly focus on the decline in oocyte quality, but the effects of age on the endometrium are often overlooked. Therefore, there is a lack of studies centered on endometrial factor and considering age year-by-year to be able to strictly establish when fertility decline starts and its magnitude. Study design, size, duration This retrospective study included a total of 33,735 single embryo transfers from 21,506 IVF AMA patients (&amp;gt;35 y.o, BMI 19-25 kg/m2) performed with high quality embryos (embryos from donated oocytes (donor &amp;lt;35 y.o) or euploid embryos from their own oocyte cohort) transferred in day 5 in an endometrial Hormonal Replacement Therapy (HRT) cycle. Evaluated transfers were performed between 2001 and 2022 in 19 clinics in Spain. This study was approved by an IRB (2203-FIVI-030-PD). Participants/materials, setting, methods A logistic regression analysis was performed, considering the patient’s age (42.93±3.45 y.o.) as a continuous and dependent variable and the reproductive outcome as an independent variable. The risk of obtaining a negative pregnancy outcome was estimated as risk ratios at a 95% confidence interval. A p-value&amp;lt;0.05 was considered as statistically significant. Reproductive rates (live birth and pregnancy loss) were calculated and compared each passing year in the studied range of ages (35-51 y.o). Main results and the role of chance The rates of live birth, Implantation failure and pregnancy loss associated to endometrial factor were affected as age increased in IVF AMA patients (&amp;gt;35 y.o). The implantation failure risk was increased in each passing year a 2% (Risk Ratio (RR) = 1.020 (1.003, 1.021) 95% confidence interval (95%CI), p-value=6.0E-09) and pregnancy loss risk was increased in each passing year a 1.2% (RR = 1.012, (1.003, 1.021) 95%CI, p-value=0.0096). Live birth rate decreased progressively from 42% observed in 35 y.o. to 32% in 50 y.o. Nevertheless, the pregnancy loss rate increased progressively from 29% in 35 y.o. to 37% in 50 y.o. Moreover, results shown a more drastic decrease of implantation failure rate after 42 years old and a pregnancy loss rate after 45 years old. Limitations, reasons for caution Only transfers from Spanish population under a HRT cycle were included, decreasing the results variability. However, the results could not be generalized to other hormonal cycles. Moreover, biochemical and clinical pregnancy loss could not be distinguished in this study considering pregnancy loss as a term that includes both of them. Wider implications of the findings To our knowledge, this is the first study in which the endometrial factor in AMA patients was analyzed year by year, helping to define in a more reliable way the ages at which the sharpest changes occur to improve clinical practices on endometrial implantation failure in AMA patients. Trial registration number Not applicable

P-331 Unlocking metformin’s potential: exploring its impact on inflammation and oxidative pathways in an endometriosis mouse model

Abstract Study question Could endometriosis-associated pro-oxidative and pro-inflammatory condition be mitigated by metformin? Summary answer Metformin treatment tends to reduce fibrosis in ectopic tissues, while up-regulating anti-oxidative response enzymes. What is known already Endometriosis is a pro-inflammatory and pro-oxidative disease, which causes pain and infertility. Hormonal treatments used to control the disease must be stopped when a woman attempts to conceive, meaning new therapies are necessary. We had shown that endometriosis reduces fertility in a mouse model, which metformin treatment led to its reversal. Metformin, in addition to reducing blood glucose, reduces inflammation and systemic oxidation. It has also been shown to mitigate the growth of endometrial implants in the animal model. The objective of this work is to evaluate the intervention of metformin in the inflammatory and oxidative response pathways associated with endometriosis. Study design, size, duration Female mice were randomly divided into groups: 1-Endometriosis (n = 24); 2-Sham (n = 12); 3-Endometriosis with metformin (n = 21); 4-Sham with metformin (n = 16). Endometriosis was surgically induced by heterologous transplantation of endometrium from a donor into recipients of the same strain. Metformin (0,5mg/mL) was administered orally for 3 months. Half of the mice in each group were mated for the fertility study. Tissues analysed are described with “Eu” for eutopic and “Ec” for ectopic tissues, respectively. Participants/materials, setting, methods The implants were confirmed and monitored by ultrasound and serum glucose, estradiol and lipoprotein values were quantified by enzymatic-colorimetric methods. Fibrosis in eutopic and ectopic tissues was quantified after computer-assisted analysis in Masson’s Trichrome images. Dual-labelling immunofluorescence was performed for the proteins F4/80 and Galectin-3 (gal-3) and Western Blot (WB) was performed to quantify their expression and also NFkB, HO-1 and GPX-1, both in eutopic and ectopic tissues. Main results and the role of chance No differences were found in serum analysis between groups, apart from a decrease in total cholesterol and an increase in estradiol in pregnant compared to non-mated mice. Masson’s Trichrome stained sections revealed increased stromal fibrosis in both eutopic and ectopic tissues of non-mated Endometriosis groups compared to Sham groups(EuEnd vs. EuSham p = 0.018; EcEnd vs. EuSham p = 0.022; EuEnd vs. EuShamMet p = 0.0008; EcEnd vs EuShamMet p = 0,001). Metformin treatment showed a decreasing trend in fibrosis both in non-mated and pregnant, particularly in ectopic tissue, although statistical significance was not reached. Both eutopic and ectopic endometrium presented positive cells for f4/80 and gal-3, with some co-localization, in all groups. Metformin treatment led to the increase of the inflammation markers in the tissues of mice with ENDO: F4/80 was increased in eutopic tissue(p = 0.0002), while gal-3 (p = 0.004) and NF-kB (p = 0.03) augmented in ectopic tissue of non-mated. Pregnancy condition, on the other hand had a tendency for the decrease of these proteins. Metformin increased the expression of antioxidant defense enzymes HO-1 (p = 0,005) and GPX-1 (p = 0,0001) in the ectopic endometrium of non-mated mice. Metformin has shown to act in the up-regulation of anti-oxidative defenses that intervene in the control of endometriosis progression, but this effect was not evident during pregnancy. Limitations, reasons for caution Caution is necessary when extrapolating animal model data to humans due to species-specific differences in endometriosis patterns. Further studies, assessing pro-oxidative species, total antioxidant capacity, and potential anti-inflammatory roles of studied molecules, are essential to clarify the unexpected results in inflammatory pathways. Wider implications of the findings These findings highlight metformin, in a subtherapeutic dose, as a novel and safe approach to alleviate oxidative stress in endometriosis, offering a promising pharmacological solution for improving endometriosis-associated infertility, once we also had shown the reversal of infertility associated with endometriosis through metformin treatment. Trial registration number not applicable

Immune Checkpoints in Endometriosis-A New Insight in the Pathogenesis.

Endometriosis (EMS) is an oestrogen-dependent, chronic disease affecting women of a reproductive age. One of the important factors involved in the development of this disease is the complex disorders associated with the functioning of the immune system. Recent evidence has shown that EMS development is associated with changes in systemic and local immunity, including functional disturbances of effector and antigen-presenting cells. One of the reasons for immune imbalance can be the improper expression of immune checkpoints (ICPs). ICPs and their ligands are responsible for maintaining self-tolerance and the modulation of the initiation, duration, and magnitude of the immune response of effector cells in normal tissues to avoid tissue damage. Considering the complex nature of co-stimulatory or co-inhibitory ICPs and the signalling between effector cells and APCs, we hypothesise that changes in cells' activity caused by ICPs may lead to serious immune system disturbances in patients with endometriosis. Moreover, both upregulation and downregulation in the expression of ICPs may be implicated in this process, including the reduced activity of effector cells against endometrial implants and disturbances in the antigen-presenting process. In this narrative review, we discuss, for the first time, key findings from the emerging literature, describing the associations between ICPs and their possible implication in the pathogenesis of endometriosis.

Malignancy in Abdominal Wall Endometriosis: Is There a Way to Avoid It? A Systematic Review.

Background: Malignant-associated abdominal wall endometriosis (AWE) is a rare pathology, likely to occur in 1% of scar endometriosis. The objectives of this study were to update the evidence on tumor degeneration arising from AWE to notify about the clinical characteristics, the different treatments offered to patients and their outcomes. Methods: A comprehensive systematic review of the literature was conducted. PubMed, Embase and Cochrane Library databases were used. Prospero (ID number: CRD42024505274). Results: Out of the 152 studies identified, 63 were included, which involved 73 patients. The main signs and symptoms were a palpable abdominal mass (85.2%) and cyclic pelvic pain (60.6%). The size of the mass varied between 3 and 25 cm. Mean time interval from the first operation to onset of malignant transformation was 20 years. Most common cancerous histological types were clear cell and endometrioid subtypes. Most widely accepted treatment is the surgical resection of local lesions with wide margins combined with adjuvant chemotherapy. The prognosis for endometriosis-associated malignancy in abdominal wall scars is poor, with a five-year survival rate of around 40%. High rates of relapse have been reported. Conclusions: Endometrial implants in the abdominal wall should be considered as preventable complications of gynecological surgeries. Special attention should be paid to women with a history of cesarean section or uterine surgery.

Neu5Gc regulates decidual macrophages leading to abnormal embryo implantation.

Repeated implantation failure (RIF) is one of the most prominent problems in the field of assisted reproduction. Neu5Gc on the surface of decidual macrophages (dMΦ) leads to different activation patterns of dMΦ, which affects embryo implantation and development. Cmah-/- (Neu5Gc-deficient) mice induced to produce anti-Neu5Gc antibodies in vivo were given a special diet rich in Neu5Gc and their fertility was monitored. The long-term diet rich in Neu5Gc induced the decrease of endometrial receptivity of female mice. The pregnancy rate of female mice fed the normal diet was 63.6% (n = 11) and the average number of embryos was 9.571 ± 1.272, while the pregnancy rate of female mice fed the diet rich in Neu5Gc was 36.4% (n = 11) and the average number of embryos in pregnant mice was 5.750 ± 3.304. The intake of Neu5Gc and the production of anti-Neu5Gc antibody led to M1 polarization of endometrial dMΦ and abnormal embryo implantation.

In-depth proteome characterization of endometrium and extraembryonic membranes during implantation in pig

BackgroundProteome characterization of the porcine endometrium and extraembryonic membranes is important to understand mother-embryo cross-communication. In this study, the proteome of the endometrium and chorioallantoic membrane was characterized in pregnant sows (PS) during early gestation (d 18 and 24 of gestation) and in the endometrium of non-pregnant sows (NPS) during the same days using LC-MS/MS analysis. The UniProtKB database and ClueGO were used to obtain functional Gene Ontology annotations and biological and functional networks, respectively.ResultsOur analysis yielded 3,254 and 3,457 proteins identified in the endometrium of PS and NPS, respectively; of these, 1,753 being common while 1,501 and 1,704 were exclusive to PS and NPS, respectively. In addition, we identified 3,968 proteins in the extraembryonic membranes of PS. Further analyses of function revealed some proteins had relevance for the immune system process and biological adhesion in endometrium while the embryonic chorion displayed abundance of proteins related to cell adhesion and cytoskeletal organization, suggesting they dominated the moment of endometrial remodeling, implantation and adhesion of the lining epithelia. Data are available via ProteomeXchange with identifier PXD042565.ConclusionThis is the first in-depth proteomic characterization of the endometrium and extraembryonic membranes during weeks 3 to 4 of gestation; data that contribute to the molecular understanding of the dynamic environment during this critical period, associated with the majority of pregnancy losses.

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.

The effect of pentoxifylline and different types of exercise training on coagulation factors in a rat endometriosis model

ObjectivesThis study evaluated the effects of high-intensity interval training (HIIT), moderate-intensity continuous training (MICT), and pentoxifylline (PTX) on coagulation factors, including the amount and percentage of lymphocytes, PLC, PLR, aPTT, PT, PT.I.N. R in a model of rats with endometriosis. MethodsEndometriosis was surgically induced in female SpragueDawley rats. The rats with confirmed endometrial implants were divided into control, MICT, pentoxifylline (D), HIIT+D, and MICT+D, HIIT groups. D and exercise interventions were performed for eight weeks. Then, the macroscopic size of endometriosis lesions was measured, and inflammatory factors (count and percentage of lymphocytes) and coagulation factors, including PLC, PLR, aPTT, PT, PT.I.N. R, and PLR in blood samples were evaluated. ResultsD significantly decreased the volume of lesions and significantly increased PT and PT.I.N. R in blood. HIIT decreased the volume of lesions and significantly increased PT. MICT did not cause significant effects on the target variables. MICT+D decreased the volume of lesions. HIIT+D significantly decreased the volume of lesions and PLC and increased aPTT as well as the count and percentage of lymphocytes, PT, and PT.I.N. R, and decreased PLR. ConclusionsAll interventions(except for MICT) especially HIIT+D and D by priority, induced a significant effect on reducing some indices of inflammation and coagulation.

Integration of 16 S rRNA gene sequencing, metabonomics and metagenome analysis to investigate the mechanism of Sparganium stoloniferum-Curcuma phaeocaulis in treating of endometriosis in rats

BackgroundEndometriosis is a gynecological disease that causes severe chronic pelvic pain and infertility in women. The therapeutic efficacy of the traditional herbal combination of Sparganium stoloniferum-Curcuma phaeocaulis (Sangleng-Ezhu, SL-EZ) in the treatment of endometriosis has been established. However, the precise mechanism by which this treatment exerts its effects remains elusive. MethodsTo gain further insights, UPLC-MS/MS was employed to identify the primary chemical constituents of SL-EZ in serum. Additionally, network pharmacology was utilized to analyze the active ingredients and their corresponding targets. Furthermore, the impact of SL-EZ on ectopic endometrial growth in endometrial implants was assessed using a rat model. The therapeutic mechanism of SL-EZ in rats with endometriosis was further investigated through the application of 16 S rRNA gene sequencing, metagenomic sequencing, and metabolomics. ResultsThe primary compounds in serum were zederone, p-coumaric acid, dehydrocostus lactone, curdione, curcumol. The growth of ectopic lesions in a rat model was effectively inhibited by SL-EZ. In comparison to the control group, the endometriotic rats exhibited a decrease in α-diversity of the gut microbiota, an increase in the Firmicutes/Bacteroidetes ratio, and a reduction in the abundance of Ruminococcaceae. Following SL-EZ intervention, the potential probiotic strains Lactobacillus gasseri and Lactobacillus johnsonii were able to restore the intestinal microenvironment at the species level. The altered metabolites were significantly correlated with Verrucomicrobia, Proteobacteria, and Bacteroidetes. The metabolomic analysis demonstrated significant alterations in intestinal metabolites. And SL-EZ intervention also exerted regulatory effects on various metabolic pathways in gut microbiota, including aminoacyl-tRNA biosynthesis, monobactam biosynthesis, cyanoamino acid metabolism, glycine, serine and threonine metabolism, plant secondary metabolite biosynthesis, and amino acid biosynthesis. ConclusionThe identification of novel treatment formulations for endometriosis was achieved through the utilization of network pharmacology and gut microbiota analyses. Our findings revealed simultaneous alterations in the microbiota within the rat model of endometriosis. The therapeutic efficacy of SL-EZ in treating endometriosis is attributed to its ability to restore the gut microbiota and regulate metabolism. This investigation offers valuable insights into the therapeutic mechanisms of traditional Chinese medicine (TCM) for endometriosis.

Transcriptome analysis of adenomyosis eutopic endometrium reveals molecular mechanisms involved in adenomyosis-related implantation failure and pregnancy disorders

BackgroundWomen with adenomyosis are characterized by having defective decidualization, impaired endometrial receptivity and/or embryo-maternal communication, and implantation failure. However, the molecular mechanisms underlying adenomyosis-related infertility remain unknown, mainly because of the restricted accessibility and the difficult preservation of endometrial tissue in vitro. We have recently shown that adenomyosis patient-derived endometrial organoids, maintain disease-specific features while differentiated into mid-secretory and gestational endometrial phase, overcoming these research barriers and providing a robust platform to study adenomyosis pathogenesis and the associated molecular dysregulation related to implantation and pregnancy disorders. For this reason, we aim to characterize the dysregulated mechanisms in the mid-secretory and gestational endometrium of patients with adenomyosis by RNA-sequencing.MethodsEndometrial organoids were derived from endometrial biopsies collected in the proliferative phase of women with adenomyosis (ADENO) or healthy oocyte donors (CONTROL) (n = 15/group) and differentiated into mid-secretory (-SECorg) and gestational (-GESTorg) phases in vitro. Following RNA-sequencing, the significantly differentially expressed genes (DEGs) (FDR < 0.05) were identified and selected for subsequent functional enrichment analysis and QIAGEN Ingenuity Pathway Analysis (IPA). Statistical differences in gene expression were evaluated with the Student’s t-test or Wilcoxon test.ResultsWe identified 1,430 DEGs in ADENO-SECorg and 1,999 DEGs in ADENO-GESTorg. In ADENO-SECorg, upregulated genes included OLFM1, FXYD5, and RUNX2, which are involved in impaired endometrial receptivity and implantation failure, while downregulated genes included RRM2, SOSTDC1, and CHAC2 implicated in recurrent implantation failure. In ADENO-GESTorg, upregulated CXCL14 and CYP24A1 and downregulated PGR were related to pregnancy loss. IPA predicted a significant inhibition of ID1 signaling, histamine degradation, and activation of HMGB1 and Senescence pathways, which are related to implantation failure. Alternatively, IPA predicted an inhibition of D-myo-inositol biosynthesis and VEGF signaling, and upregulation of Rho pathway, which are related to pregnancy loss and preeclampsia.ConclusionsIdentifying dysregulated molecular mechanisms in mid-secretory and gestational endometrium of adenomyosis women contributes to the understanding of adenomyosis-related implantation failure and/or pregnancy disorders revealing potential therapeutic targets. Following experimental validation of our transcriptomic and in silico findings, our differentiated adenomyosis patient-derived organoids have the potential to provide a reliable platform for drug discovery, development, and personalized drug screening for affected patients.

Comparative analysis of superovulated versus uterine-embryo synchronized recipients for embryo transfer in cynomolgus monkeys (Macaca fascicularis).

Assisted reproductive technologies (ARTs), such as intracytoplasmic sperm injection and embryo transfer, are essential for generating genetically edited monkeys. Despite their importance, ARTs face challenges in recipient selection in terms of time and the number of animals required. The potential of superovulated monkeys, commonly used as oocyte donors, to serve as surrogate mothers, remains underexplored. The study aimed to compare the efficacy of superovulated and uterine-embryo synchronized recipients of embryo transfer in cynomolgus monkeys (Macaca fascicularis). This study involved 23 cynomolgus monkeys divided into two groups-12 superovulated recipients and 11 synchronized recipients. The evaluation criteria included measuring endometrial thickness on the day of embryo transfer and calculating pregnancy and implantation rates to compare outcomes between groups. The study found no statistically significant differences in endometrial thickness (superovulated: 4.48 ± 1.36 mm, synchronized: 5.15 ± 1.58 mm), pregnancy rates (superovulated: 30.8%, synchronized: 41.7%), and implantation rates (superovulated: 14.3%, synchronized: 21.9%) between the groups (p > 0.05). The observations indicate that superovulated recipients are as effective as synchronized recipients for embryo transfer in cynomolgus monkeys. This suggests that superovulated recipients can serve as viable options, offering an efficient and practical approach to facilitate the generation of gene-edited models in this species.