Genital tract dysbiosis in infertile women with a history of repeated implantation failure and pilot study for reproductive outcomes following oral enteric coating lactoferrin supplementation.

PURPOSE: We prospectively investigated if oral enteric coating lactoferrin supplementation improves the reproductive outcomes in infertile women with a history of repeated implantation failure (RIF) and non-Lactobacillus-dominant (Lactobacillus rate < 90%) microbiota (NLDM) in vaginal secretions (VS)/endometrial fluid (EF). METHODS: Paired VS/EF samples were obtained from RIF women and control infertile women (non-RIF group) for microbiome analysis. Chronic endometritis (CE) was diagnosed histopathologically and hysteroscopically. In a pilot study, oral enteric coating lactoferrin (700 mg/day, at least 28 consecutive days) was administered to eligible patients with NLDM in VS/EF. Their reproductive outcomes in the subsequent vitrified-warmed embryo transfer cycles were followed up. RESULTS: While CE was more prevalent (OR 2.41, 95% CI 1.02-5.63, p = 0.042) in the RIF group (29.1%, n = 117) than in the non-RIF group (14.5%, n = 55), The NLDM rate was similar between the two groups (44.4 vs 52.7%). Lactoferrin supplementation improved NLDM in 43.2% of RIF women (n = 37). Within the RIF group, the live birth rate in the subsequent cycles was higher (OR 10.67, 95% CI 1.03 - 110.0, p = 0.046) in women with improved microbiota (57.1%, n = 14) than in those with unimproved microbiota (11.1%, n = 9). CONCLUSION: Unlike CE, NLDM was not unique to RIF but was common in infertile women. Although the therapeutic effect of the oral lactoferrin supplementation on NLDM was limited in a pilot study, the reproductive outcomes were better in RIF women who overcame NLDM than in those who failed. Randomized controlled trials are required to confirm the results. TRIAL REGISTRATION NUMBER AND DATE FOR PROSPECTIVELY REGISTERED TRIALS: UMIN-CTR 000036990, June 7, 2019.

Multi-drug-resistant chronic endometritis in infertile women with repeated implantation failure: trend over the decade and pilot study for third-line oral antibiotic treatment.

PURPOSE: To evaluate the yearly prevalence and annual transition of multi-drug-resistant-chronic endometritis (MDR-CE) in infertile women with a history of repeated implantation failure (RIF) and to establish the third-line antibiotic treatment regimen against MDR-CE. METHODS: This retrospective/prospective cohort and pilot study included 3473 RIF women between April 2010 and September 2021. The endometrial stromal plasmacyte density index (ESPDI) was calculated in 3449 CD138-immunostained endometrial sections to evaluate CE. The microbiota in the vaginal secretions and endometrial fluid was compared between 17 patients with MDR-CE and 16 patients with antibiotics-sensitive CE. In a pilot study, oral moxifloxacin (400 mg/day, 10 days, n = 24) or azithromycin (500 mg/day, 3 days, n = 24) was administered to eligible patients with MDR-CE. RESULTS: From April 2010 to March 2020, CE was detected in 31.4% of RIF women and MDR was detected in 7.8% of CE. While the prevalence of CE was stable for a decade, MDR in CE increased steadily (OR 8.27, 95% CI 2.58-26.43, p trend < 0.001). The bacterial species/communities unique to MDR-CE were not found. The histopathologic cure rate of MDR-CE was similar between the moxifloxacin and azithromycin groups (79.2% vs 75.0%, OR 1.27, 95% CI 0.32-4.89, p value 0.73), as well as reproductive outcomes in subsequent embryo transfer cycles. CONCLUSION: In RIF women, MDR in CE increased over the decade. As a third-line treatment for MDR-CE, azithromycin may have a clinical advantage due to its shorter time administration periods. CLINICAL TRIAL NUMBER: ClinicalTrials.gov Identifier: UMIN-CTR 000029449/000031909.

Differential Vaginal Microbiota Profiling in Lactic-Acid-Producing Bacteria between Infertile Women with and without Chronic Endometritis.

PURPOSE: Chronic endometritis (CE) is an infectious and inflammatory disorder associated with infertility of unknown etiology, repeated implantation failure, and recurrent pregnancy loss. In the current clinical practice, intrauterine interventions such as endometrial biopsy/histopathologic examinations and/or hysteroscopy are required for the diagnosis of CE. In this study, we analyzed the microbiota in vaginal secretions (VS) as a potential prediction tool for CE in infertile women. METHODS: Using next-generation sequencing analysis, we compared the VS and endometrial fluid (EF) microbiota in infertile women with (n = 20) or without CE (n = 103). RESULTS: The detection rate of Streptococcus and Enterococcus as well as the bacterial abundance of Atopobium and Bifidobacterium in the VS microbiota was significantly lower in the CE group than in the non-CE group. Meanwhile, the detection rate and bacterial abundance of Lactobacillus in the EF and VS microbiota were at similar levels between the two groups. CONCLUSION: These findings suggest that VS microbiota in infertile women with CE is characterized by the reduction in Bifidobacterium and lactic-acid-producing bacteria other than Lactobacillus. Our results hold promise for the prediction of CE, not by somewhat interventional intrauterine procedures, but by less invasive VS sampling. TRIAL REGISTRATION NUMBER: UMIN000029449 (registration date 6 October 2017).

Characterization of Microbiota in Endometrial Fluid and Vaginal Secretions in Infertile Women with Repeated Implantation Failure.

Studies suggest that persisting intrauterine bacterial infectious conditions such as chronic endometritis potentially impair the embryo implantation process. The microbial environment in the female reproductive tract, however, remains largely undetermined in infertile patients with a history of repeated implantation failure (RIF). Using next-generation sequencing, we aimed to characterize the microbiota in the endometrial fluid (EF) and vaginal secretions (VS) in women with RIF. Twenty-eight infertile women with a history of RIF and eighteen infertile women undergoing the first in vitro fertilization-embryo transfer attempt (the control group) were enrolled in the study. On days 6-8 in the luteal phase of the natural, oocyte-pickup, or hormone replacement cycle, the paired EF and VS samples were obtained separately. Extracted genomic DNA was pyrosequenced for the V4 region of 16S ribosomal RNA using a next-generation sequencer. The EF microbiota had higher α-diversity and broader bacterial species than the VS microbiota both in the RIF and control groups. The analysis of the UniFrac distance matrices between EF and VS also revealed significantly different clustering. Additionally, the EF microbiota, but not the VS microbiota, showed significant variation in community composition between the RIF group and the control group. Burkholderia species were not detected in the EF microbiota of any samples in the control group but were detectable in a quarter of the RIF group. To our best knowledge, this is the first study investigating the microbiota in the paired EF and VS samples in infertile women with RIF.

Endometritis: new time, new concepts.

Endometritis is subdivided into two categories. Acute endometritis is symptomatic and characterized by microabscess formation and neutrophil invasion in the endometrial superficial epithelium, gland lumina, and uterine cavity. Chronic endometritis is rather silent and recognized as unusual plasmacyte infiltration in the endometrial stromal areas. Over the last decade, studies have disclosed the potential association between poor reproductive outcomes and endometritis, particularly chronic endometritis. The aim of this review is to address the current literature surrounding chronic endometritis and highlight recent advances in the research of this long-neglected gynecologic disease.

Minimum values for midluteal plasma progesterone and estradiol concentrations in patients who achieved pregnancy with timed intercourse or intrauterine insemination without a human menopausal gonadotropin.

OBJECTIVE: The aim of the study was to assess the lower limits of midluteal plasma progesterone and estradiol concentrations in patients who achieved pregnancy with timed intercourse or intrauterine insemination without a human menopausal gonadotropin stimulation. RESULTS: We included 297 pregnant cycles of 297 women and assessed midluteal plasma progesterone and estradiol concentrations and pregnancy outcomes, retrospectively. These cycles were compared with the non-pregnant cycles (406 cycles) of the same women who became pregnant. Mean midluteal plasma P4 and E2 concentrations were significantly (P < 0.01) higher in pregnant cycles (14.5 and 188.5 pg/mL) than in non-pregnant cycles (10.7 and 162.6 pg/mL). The 5 percentiles of progesterone and estradiol in pregnant cycles were 5.6 and 70.2 pg/mL, respectively. The lowest progesterone and estradiol levels in pregnant cycles were 2.3 and 23.4 pg/mL, respectively. In non-pregnant cycles, many women had low P4 levels that were less than 5.6 ng/mL. Subgroup analyses showed slight differences among the four groups, which may have been due to the ovarian function of each group. Miscarriage was not related to progesterone and estradiol concentrations. These values may be useful for the evaluation of necessary values for pregnancy with timed intercourse or intrauterine insemination.

Live birth rate following oral antibiotic treatment for chronic endometritis in infertile women with repeated implantation failure.

PROBLEM: The aim of this prospective study was to investigate the prevalence of chronic endometritis (CE) in infertile women with a history of repeated implantation failure (RIF) and to determine whether oral antibiotic treatment improves their live birth rate in the following embryo transfer (ET) cycles. METHOD OF STUDY: Endometrial biopsy samples obtained from infertile women with RIF were subjected to immunohistochemistrical/histopathologic diagnosis of CE. Following antibiotic administration to the RIF/CE group, their histopathologic cure rate, microbial detection rate, and reproductive outcome in the subsequent ET cycles were prospectively studied. RESULTS: 33.7% of infertile women with RIF were diagnosed with CE. Following the first-line doxycycline treatment, the histopathologic cure rate in the subsequent endometrial biopsy was 92.3%. Following the second-line metronidazole/ciprofloxacin treatment, the overall cure rate was 99.1%. The live birth rate in the first ET cycle (P=.031, RR 1.48, 95% CI 1.03-2.12) and cumulative three ET cycles (P=.037, RR 1.39, 95% CI 1.02-1.90) following antibiotic treatment in the cured RIF/CE group (32.8% and 38.8%, respectively) was significantly higher than in the RIF/non-CE group (22.1% and 27.9%, respectively). CONCLUSION: Chronic endometritis was found in one-third of infertile women with RIF. The oral antibiotic treatment against CE might be a promising therapeutic option for infertile women with RIF.

Salvage hormonal therapy after failed microdissection testicular sperm extraction: A multi-institutional prospective study.

OBJECTIVE: To validate the efficacy of salvage hormonal therapy in men with non-obstructive azoospermia at their second microdissection testicular sperm extraction. METHODS: This was a multi-institutional study registered at the Japanese University Hospital Medical Information Network clinical trial center. After 1 month of human chorionic gonadotropin therapy (5000 IU, three times a week), patients were treated with recombinant human follicle-stimulating hormone (150 IU, three times a week) and human chorionic gonadotropin for the next 3 months. Three testicular samples were obtained randomly from both testes, and sent for pathological diagnosis at the first and second microdissection testicular sperm extraction. RESULTS: A total of 21 men, excluding those with chromosomal abnormalities, azoospermia factor a or b deletions, extremely small testes (<2 mL), or prior hormonal therapy, were eligible to participate based on our inclusion criteria. At the first microdissection testicular sperm extraction, 13 and six patients had Sertoli cells only and an early maturation arrest, respectively. With the second microdissection testicular sperm extraction, sperm were successfully obtained from two patients (10%). Patient age, testicular volume and hormone profiles were not associated with the results of the second microdissection testicular sperm extraction. However, the testicular histology of the two successful patients were late maturation arrest and hypospermatogenesis. CONCLUSIONS: Effectiveness of human chorionic gonadotropin-based salvage hormonal therapy preceding a second microdissection testicular sperm extraction seems to be limited. Non-obstructive azoospermia men who have differentiated cells in their testes are likely to respond to hormonal stimulation.

Clinical background affecting pregnancy outcome following local endometrial injury in infertile patients with repeated implantation failure.

Local endometrial injury (LEI) has been performed as a promising medical intervention to improve the pregnancy outcome in infertile women suffering from repeated implantation failure (RIF) in in vitro fertilization-embryo transfer cycles. The effect of LEI, however, remains controversial. The aim of this retrospective study was to identify the subgroups of patients with RIF who benefit from LEI. We compared the clinical parameters between the patients who had had a clinical pregnancy in the subsequent embryo transfer cycle following the LEI cycle (LEI-CP group, n = 94) and those who had resulted in negative pregnancy test (LEI-NP group, n = 114). The female age, basal follicle stimulating hormone concentration, number of past oocyte pickup cycles, and embryos/blastocysts transferred in the past three RIF cycles were significantly (p < 0.047) lower in the LEI-CP group than the LEI-NP group. The prevalence of polycystic ovarian syndrome was significantly (p = 0.0059) higher in the LEI-CP group than in the LEI-NP group. These findings suggest that LEI is most effective to improve the pregnancy outcome in patients undergoing RIF with uncompromised ovarian reserve, particularly in those with polycystic ovarian syndrome.

Noninvasive cellular quantity measurement in Bone Marrow Stromal Cells/ Beta-tricalcium phosphate.

This paper describes noninvasive cellular quantity measurement in Bone Marrow Stromal Cells/ ß-tricalcium phosphate. We attempt to identify cellular quantity with an ultrasonic system. The ultrasonic waves are reflected at boundaries where there is a difference in acoustic impedances of the materials on each side of the boundary. Therefore, we focus on the reflected signal. From the obtained ultrasonic data, we extract two features; amplitude and frequency. Amplitude is obtained from the raw ultrasonic wave, and frequency is calculated from frequency spectrum obtained by applying cross-spectrum method. Therefore, we suggest the superiority of frequency to analyze Bone Marrow Stromal Cells. This study shows the ability of intervention to produce the desired beneficial effect.

A neonatal brain MR image template of 1 week newborn.

PURPOSE: Magnetic resonance imaging (MRI) is often used to detect and treat neonatal cerebral disorders. However, neonatal MR image interpretation is limited by intra- and inter-observer variability. To reduce such variability, a template-based computer-aided diagnosis system is being developed, and several methods for creating templates were evaluated. METHOD: Spatial normalization for each individual's MR images is used to accommodate the individual variation in brain shape. Because the conventional normalization uses as adult brain template, it can be difficult to analyze the neonatal brain, as there are large difference between the adult brain and the neonatal brain. This article investigates three approaches for defining a neonatal template for 1-week-old newborns for diagnosing neonatal cerebral disorders. The first approach uses an individual neonatal head as the template. The second approach applies skull stripping to the first approach, and the third approach produces a template by averaging brain MR images of 7 neonates. To validate the approaches, the normalization accuracy was evaluated using mutual information and anatomical landmarks. RESULTS: The experimental results of 7 neonates (revised age 5.6 ± 17.6 days) showed that normalization accuracy was significantly higher with the third approach than with the conventional adult template and the other two approaches (P < 0.01). CONCLUSION: Three approaches to neonatal brain template matching for spinal normalization of MRI scans were applied, demonstrating that a population average gave the best results.

Normal live birth after testicular sperm extraction and intracytoplasmic sperm injection in variant primary ciliary dyskinesia with completely immotile sperm and structurally abnormal sperm tails.

OBJECTIVE: To report on the investigation and fertility management of variant primary ciliary dyskinesia (PCD). DESIGN: Case report. SETTING: University-affiliated assisted reproductive technologies practice. PATIENT(S): A 40 year-old man presenting with 12 months' primary infertility, complete sperm immotility, severe morphologic defects, and moderate sinopulmonary disease. INTERVENTION(S): Electron microscopy (EM) of sperm, nasal cilial function studies, open testis biopsy, and sperm extraction for intracytoplasmic sperm injection (ICSI). MAIN OUTCOME MEASURE(S): Outcome of ICSI treatment using immotile testicular sperm. RESULT(S): EM revealed abnormal connecting pieces, shortened midpieces with attenuated mitochondrial sheaths, poorly developed annulus, abnormal outer dense fibers, and axonemes missing the two central mircotubules. Nasal ciliary beat frequency was subnormal and dyssynchronous. Immotile testicular sperm were selected for ICSI based on physical characteristics and fertilized 12 of 18 eggs. A single day-5 blastocyst achieved a normal pregnancy and delivery of a healthy 3,840-g girl at 38 weeks' gestation. CONCLUSION(S): Nonclassic PCD may present with structurally abnormal completely immotile sperm, with seemingly little prospect of fertility, and moderate respiratory dysfunction supporting the presence of an underlying ciliopathy. Despite testicular sperm also being immotile and showing profound structural defects that would seem to preclude fertilization, more morphologically normal sperm are capable of establishing a normal pregnancy.

Surgical recovery of sperm in non-obstructive azoospermia.

The development of intracytoplasmic sperm injection (ICSI) opened a new era in the field of assisted reproduction and revolutionized the assisted reproductive technology protocols for couples with male factor infertility. Fertilisation and pregnancies can be achieved with spermatozoa recovered not only from the ejaculate but also from the seminiferous tubules. The most common methods for retrieving testicular sperm in non-obstructive azoospermia (NOA) are testicular sperm aspiration (TESA: needle/fine needle aspiration) and open testicular biopsy (testicular sperm extraction: TESE). The optimal technique for sperm extraction should be minimally invasive and avoid destruction of testicular function, without compromising the chance to retrieve adequate numbers of spermatozoa to perform ICSI. Microdissection TESE (micro-TESE), performed with an operative microscope, is widely considered to be the best method for sperm retrieval in NOA, as larger and opaque tubules, presumably with active spermatogenesis, can be directly identified, resulting in higher spermatozoa retrieval rates with minimal tissue loss and low postoperative complications. Micro-TESE, in combination with ICSI, is applicable in all cases of NOA, including Klinefelter syndrome (KS). The outcomes of surgical sperm retrieval, primarily in NOA patients with elevated serum follicle-stimulating hormone (FSH) (NOA including KS patients), are reviewed along with the phenotypic features. The predictive factors for surgical sperm retrieval and outcomes of treatment were analysed. Finally, the short- and long-term complications in micro-TESE in both 46XY males with NOA and KS patients are considered.

Molecular changes induced by bisphenol-A in rat Sertoli cell culture.

Bisphenol-A (BPA) is an industrial chemical and is known to act as an endocrine disrupter. This study was designed to evaluate how BPA regulates Sertoli cell (SC) signal molecules. Purified rat SCs were cultured and treated with BPA (200 µmol/l) at various time points. Western blot analysis was used to determine the activation of extracellular signal-related kinases 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK), p38 mitogen activated protein kinase (MAPK), nuclear factor kappa B (NF-κB), cyclooxygenase-1,2 (COX-1, 2), estrogen receptor-α (ER-α), and androgen receptor (AR). The levels of transferrin (TF), prostaglandin E(2) (PGE(2)), and prostaglandin F(2α) (PGF(2α)) in culture medium were quantified by ELISA. Interleukin (IL)-1ß and IL-6 mRNAs were measured by quantitative real-time PCR (QRT-PCR). Compared with the control, BPA activated the phosphorylation of ERK1/2 (p-ERK1/2) through 30 min to 6 h. TF was down-regulated at 6 and 24 h. Furthermore, IL-1ß was up-regulated at 30 min and IL-6 was up-regulated at 1 and 24 h. ERK activity inhibitor (PD98059, 10 µmol/l) inhibited these molecular changes. These results reveal the possibility that BPA may have adverse effects on spermatogenesis via ERK1/2.

Learning curves of microdissection testicular sperm extraction for nonobstructive azoospermia.

OBJECTIVES: Microdissection testicular sperm extraction (micro-TESE) has been performed at some experienced male infertility centers. Micro-TESE in patients with nonobstructive azoospermia (NOA) has been shown to be not only feasible and safe but also a technically demanding operation. To evaluate whether the improvement of surgical outcomes of micro-TESE, we hereby report the learning curves of micro-TESE for NOA patients. DESIGN: Retrospective clinical analysis. SETTING: Male infertility center. PATIENT(S): Since 2006, micro-TESE was performed in 150 patients with NOA by a single surgeon. INTERVENTION(S): After the tunica albuginea was opened, direct examination of the testicular parenchyma was performed at x25 magnification. MAIN OUTCOME MEASURE(S): Sperm retrieval rate and clinical factors in the first 50 patients (group A), the middle 50 patients (group B), and the last 50 patients (group C) were examined. RESULT(S): There were no differences in clinical factors among the three groups. Total operation times were shorter in group B and C than in group A (P<0.05). The sperm retrieval rate in group B (44%) and C (48%) was significantly higher than in group A (32%; P<0.05). CONCLUSION(S): As cases increase, surgical outcomes and sperm retrieval rate have improved. We think that this report showed substantial learning curves for a microscopic surgery such as micro-TESE.

Serum hormones in patients with nonobstructive azoospermia after microdissection testicular sperm extraction.

PURPOSE: Microdissection testicular sperm extraction combined with intracytoplasmic sperm injection is currently used to treat infertility in cases of nonobstructive azoospermia. Even in patients with nonmosaic Klinefelter's syndrome, who usually present with small testes and hypogonadism, the procedure has been done successfully. We assessed serum hormones after microdissection testicular sperm extraction and compared postoperative testicular damage between 46XY males with nonobstructive azoospermia and those with Klinefelter's syndrome. MATERIALS AND METHODS: We retrospectively reviewed the records of 140 men with azoospermia, including 100 46XY males with nonobstructive azoospermia and 40 with nonmosaic Klinefelter's syndrome, who underwent microdissection testicular sperm extraction. Serum follicle-stimulating hormone, luteinizing hormone and testosterone were evaluated before, and 1, 3, 6, 9, 12 and 18 months after surgery. RESULTS: In 46XY males with nonobstructive azoospermia serum follicle-stimulating hormone during 18 months of followup, and luteinizing hormone 1 and 3 months postoperatively were significantly increased vs baseline. No significant differences were observed in testosterone at any postoperative time point vs baseline. In men with Klinefelter's syndrome who underwent sperm extraction mean testosterone significantly decreased an average of 30% to 35% vs baseline when assessed 1, 3, 6, 9 and 12 months postoperatively. It returned to 75% of the preoperative level after 18 months. In Klinefelter's syndrome cases no significant differences were observed in follicle-stimulating hormone and luteinizing hormone at each postoperative time point. CONCLUSIONS: Hormonal followup after microdissection testicular sperm extraction is recommended, particularly in patients with Klinefelter's syndrome, to prevent the deleterious consequences of hypogonadism.

The efficacy of adult orchidopexy as a treatment of male infertility: our experience of 20 cases.

The objective of this study was to determine whether adult orchidopexy and testicular sperm extraction are effective or not as a treatment of infertility. In this study, we could not show the efficacy of orchidopexy as a treatment of infertility in adulthood, and testicular sperm extraction can be a useful treatment for adult patients with unilateral undescended testis.

Cyclin and protamine as prognostic molecular marker for testicular sperm extraction in patients with azoospermia.

The CCNA1, CCNB1, CCNB2, PRM1, and PRM2 messenger RNA transcript ratios were significantly decreased in patients with spermatogenic disorders. These messenger RNA transcript ratios in patients with successful sperm retrieval were higher than in patients with failed sperm retrieval.

Rapidly progressing, late-onset obstructive azoospermia linked to herniorrhaphy with mesh.

OBJECTIVE: To report a case of a man presenting with rapidly progressing inguinal vasal obstruction 5 years after bilateral herniorrhaphy with polypropylene mesh. DESIGN: Case report with a review of the scientific literature. SETTING: Male infertility clinic, Kobe University Hospital (Kobe, Japan). PATIENT(S): A 30-year-old patient who had undergone bilateral inguinal herniorrhaphy using polypropylene mesh 5 years previously. INTERVENTION(S): The patient underwent bilateral vasography and left testicular sperm extraction (TESE). MAIN OUTCOME MEASURE(S): Long-term effects of the adult inguinal herniorrhaphy with polypropylene mesh. RESULT(S): Rapid progression of vas deferens obstruction occurred within several months. We performed left TESE for intracytoplasmic sperm injection (ICSI) and retrieved many motile sperm. CONCLUSION(S): Before azoospermia, men who undergo inguinal herniorrhaphy using polypropylene mesh need to rapidly cryopreserve their sperm for future fertility; TESE-ICSI is also a selectable treatment.