Therapeutic efficacy of gentle endometrial curettage on antibiotic-resistant chronic endometritis in infertile women.

Purpose: To identify the efficacy of endometrial curettage on antibiotic-resistant chronic endometritis (CE) in infertile women. Methods: Of 1580 women with CE, 87 with antibiotic-resistant CE after two to five cycles of antibiotic treatment were recruited between 2019 and 2021. The women who underwent endometrial curettage without applying any force and, in the subsequent menstrual cycle, endometrial sampling for CD138 immunostaining without antibiotic use. Pregnancy outcomes after in vitro fertilization treatment were analyzed in women who did not desire endometrial curettage and in those with cured and persistent CE after endometrial curettage. Results: In 64 women who underwent endometrial curettage, the number of CD138-positive cells decreased from 28.0 ± 35.3 to 7.7 ± 14.0 (p < 0.0001), and CE in 41 women (64.1%) was cured (<5 CD138-positive cells). The pathological findings detected 3.1% of endometrial hyperplasia and 1.6% of endometrial cancer. The ongoing pregnancy rates in women aged ≤42 without endometrial curettage were significantly lower than those of women with cured and persistent CE (26.7%, 67.6%, and 57.1%, respectively, p = 0.03). Conclusions: Gentle endometrial curettage for antibiotic-resistant CE significantly decreased the number of CD138-positive cells, resulting in improved pregnancy outcomes regardless of remaining CE. Endometrial curettage is also important as a screening for endometrial malignancy.

Prevalence of and risk factors for chronic endometritis in patients with intrauterine disorders after hysteroscopic surgery.

OBJECTIVE: To identify the prevalence of and risk factors for chronic endometritis (CE) in patients with intrauterine disorders and the therapeutic efficacy of hysteroscopic surgery in the treatment of CE without antibiotic therapy. DESIGN: Prospective cohort study. SETTING: Hospital specializing in reproductive medicine. PATIENT(S): The study population consisted of 350 women with infertility, of whom 337 were recruited, who underwent hysteroscopic surgery between November 2018 and June 2021. Eighty-nine consecutive patients without intrauterine disorders were also recruited as controls. INTERVENTION(S): Endometrial samples were collected during the surgery for CD138 immunostaining for the diagnosis of CE. In women diagnosed with CE, endometrial biopsy was performed without antibiotic use in the subsequent menstrual cycle. MAIN OUTCOME MEASURE(S): Prevalence of and risk factors for CE in intrauterine disorders and therapeutic effects of hysteroscopic surgery on CE. RESULT(S): The prevalence of CE with ≥5 CD138-positive cells in women with no intrauterine disorder and with endometrial polyps, myomas, intrauterine adhesions (IUAs), and septate uterus was 15.7%, 85.7%, 69.0%, 78.9%, and 46.2%, respectively. A multivariate analysis revealed that CE was diagnosed significantly more often in the endometrial polyp (odds ratio, 27.69; 95% confidence interval, 15.01-51.08) and IUA groups (odds ratio, 8.85; 95% confidence interval, 3.26-24.05). The rate of recovery from CE with surgery in women with endometrial polyps, myomas, IUA, and septate uterus was 89.7%, 100%, 92.8%, and 83.3%, respectively. CONCLUSION(S): Endometrial polyp and IUA were risk factors for CE. Most CE cases with intrauterine disorders were cured with hysteroscopic surgery without antibiotic therapy, regardless of the type of intrauterine abnormalities.

Analysis of the therapeutic effects of hysteroscopic polypectomy with and without doxycycline treatment on chronic endometritis with endometrial polyps.

PROBLEM: We aimed to compare the therapeutic effects of hysteroscopic polypectomy with and without doxycycline treatment on chronic endometritis (CE) with endometrial polyps. METHOD OF STUDY: DESIGN: A cross-sectional study was conducted on 267 infertile patients, of whom 243 were recruited, who underwent hysteroscopic polypectomy between March 2019 and March 2020. During surgery, the endometrial specimens for the immunohistochemistry analysis of the plasma cell marker CD138 and for the intrauterine bacterial culture were obtained to diagnose CE, and the prevalence of CE was analyzed. Of the 222 women who were diagnosed with CE after polypectomy, we treated 62 women with doxycycline (antibiotic group) and did not provide antibiotics in 160 women (non-antibiotic group). RESULTS: Most of the infertile patients with endometrial polyps had CE (92.6%). The recovery rate from CE by hysteroscopic polypectomy was significantly higher in the non-antibiotic group than in the antibiotic group (88.8% and 58.1%, respectively, p < 0.0001). The duration of recovery from CE in the non-antibiotic group was shorter than that in the antibiotic group (42.6 ± 41.0 and 56.5 ± 32.3 days, respectively, p < 0.0001). The clinical pregnancy rate within 6 months in non-antibiotic group was higher than that in the antibiotic group (63.2% and 43.8%, respectively, p = 0.034). CONCLUSION: Endometrial polyps are significantly associated with CE. Most CE patients with endometrial polyps had been cured by polypectomy without doxycycline. Inappropriate antibiotic therapy may delay recovery from CE and decrease the efficacy of polypectomy on CE and pregnancy rates.

Analysis of the risk factors and treatment for repeated implantation failure: OPtimization of Thyroid function, IMmunity, and Uterine Milieu (OPTIMUM) treatment strategy.

PROBLEM: What are the pregnancy outcomes after the OPtimization of Thyroid function, Immunity, and Uterine Milieu (OPTIMUM) treatment strategy in patients with repeated implantation failure (RIF)? METHOD OF STUDY: Infertile women with a history of RIF after more than three embryo transfer (ET) cycles underwent implantation testing, including a hysteroscopy, endometrial biopsy for CD138 immunostaining and bacterial culture, and serum 25-hydroxyvitamin D3 , interferon-γ-producing helper T (Th1) cell, IL-4-producing helper T (Th2) cell, thyroid-stimulating hormone, thyroid peroxidase antibody, and thrombophilia screening between April 2017 and August 2018. We treated chronic endometritis with antibiotics, aberrant high Th1/Th2 cell ratios with vitamin D and/or tacrolimus intake, overt/subclinical hypothyroidism with levothyroxine, and thrombophilia with low-dose aspirin. Of the 116 RIF women, 88 women with 133 ET cycles were recruited from a questionnaire-based survey regarding pregnancy outcomes. Fifty-nine consecutive RIF patients without the OPTIMUM treatment strategy were also recruited as a control. RESULTS: The 116 women with RIF after the OPTIMUM treatment strategy were 38.3 ± 3.8 years old and had an implantation failure history over 5 (3-19) ET cycles. Implantation testing identified impaired intrauterine circumstances in 75 women (64.7%), an aberrant elevated Th1/Th2 cell ratio in 56 women (48.3%), and thyroid abnormalities in 33 women (28.4%). Cumulative ongoing pregnancy rates including spontaneous pregnancy in the patients aged < 40 and ≥ 40 years were 72.7% and 45.5% within two ET cycles, respectively. The pregnancy outcomes in the OPTIMUM group were significantly higher than those in the control. CONCLUSIONS: The OPTIMUM treatment strategy improved pregnancy outcomes in patients with RIF.

The actions of resveratrol in decidualizing endometrium: acceleration or inhibition?†.

Resveratrol, a natural polyphenolic compound, is widely studied for its anti-inflammatory and antisenescent properties. Recently, two studies reported seemingly conflicting findings on the actions of resveratrol on decidualization of human endometrial stromal cells (HESCs). One study by Ochiai et al. demonstrated that resveratrol inhibits decidual transformation of primary cultured HESCs. The other study by Mestre Citrinovitz et al., showed that resveratrol enhances decidualization of HESCs in culture. At a glance, the reason for these opposing observations seems puzzling. However, recent studies demonstrated that decidualization is a multistep process, which starts with an acute proinflammatory stress response that lasts for several days and is followed by the emergence of stress-resistant decidual cells as well as senescent decidual cells. The balance between these decidual subpopulations may determine if the cycling endometrium can successfully transition into the decidua of pregnancy upon embryo implantation. Here, we explore the importance of timing of drugs aimed at modulating the decidual response. We posit that resveratrol treatment during the initial proinflammatory decidual phase, i.e., coinciding with the implantation window in vivo, inhibits decidual transformation of the endometrium. However, when given after the initial phase, resveratrol may promote decidualization by inhibiting decidual senescence. Further, if restricted to the proliferative phase, resveratrol may promote ovarian function without adversely impacting on embryo implantation or decidualization. Thus, failure to align drug interventions with the correct phase of the menstrual cycle may negate beneficial clinical effects and results in adverse reproductive outcomes.

Impact of chronic endometritis on endometrial receptivity analysis results and pregnancy outcomes.

BACKGROUND: The aim of this study is to evaluate the relationship between chronic endometritis (CE) and a personalized window of implantation (WOI), identified by results of endometrial receptivity analysis (ERA), and pregnancy outcomes following embryo transfer (ET) based on the ERA outcomes. METHODS: The single-center, cross-sectional study was designed. The study population consisted of 101 infertile women who underwent endometrial sampling between June 2018 and February 2020. We recruited 88 patients who underwent ERA testing and immunohistochemistry of the plasma cell marker CD138 to diagnose CE within 3 months of testing. Subjects were divided into three groups as follows: 33 without CE (non-CE group); 19 with untreated CE at ERA testing (CE group); and 36 successfully treated for CE before ERA testing (cured-CE group). CE diagnosis was defined as ≥5 CD138-positive plasma cells per 10 random stromal areas at ×400 magnification. RESULTS: In non-CE, CE, and cured-CE groups, the numbers of CD138-positive cells were 0.7 ± 1.0, 28.5 ± 30.4, and 1.3 ± 1.3, respectively (p < .001). The rates of "receptive" endometrium in non-CE and cured-CE groups were 57.6% (19 women) and 50.0% (18 women), respectively; however, in the CE group, this rate was significantly lower than the other two groups (p = .009) at only 15.8% (3 women). After CE were treated prior or posterior to the ERA test in cured-CE or CE groups, the clinical pregnancy rates at the first ET in non-CE, CE, and cured-CE groups were 77.8% (21/27 cycles), 22.2% (4/18 cycles), and 51.7% (15/29 cycles), respectively (p < 0.001). CONCLUSION: CE had detrimental effects on the individual WOI, leading to embryo-endometrial asynchrony; therefore, diagnosis and treatment of CE should be done before ERA testing.

Preconception resveratrol intake against infertility: Friend or foe?

BACKGROUND: Resveratrol is an antiaging, antioxidant, anti-inflammatory, and insulin-sensitizing natural polyphenolic compound. Growing evidence indicates that resveratrol has potential therapeutic effects in infertile women with diminished ovarian function, polycystic ovary syndrome (PCOS), or endometriosis. However, only one clinical trial in women undergoing in vitro fertilization (IVF) cycles using resveratrol has ever been reported. This review focuses on the potential therapeutic effects of resveratrol on pregnancy and on its advantages and disadvantages in pregnancy outcomes during infertility treatment. METHODS: We performed a literature review to describe the known impacts of resveratrol on the ovary and endometrium. RESULTS: Resveratrol upregulates sirtuin (SIRT)1 expression in ovaries, which is associated with protection against oxidative stress. It leads to the activation of telomerase activity and mitochondrial function, improving ovarian function. In the endometrium, resveratrol downregulates the CRABP2-RAR pathway leading to suppressing decidual and senescent changes of endometrial cells, which is essential for embryo implantation and placentation. Moreover, resveratrol may also induce deacetylation of important decidual-related genes. CONCLUSIONS: Resveratrol has potential therapeutic effects for improving ovarian function; however, it also has anti-deciduogenic actions in uterine endometrium. In addition, its teratogenicity has not yet been ruled out; thus, resveratrol should be avoided during the luteal phase and pregnancy.

Influence of resveratrol supplementation on IVF-embryo transfer cycle outcomes.

RESEARCH QUESTION: Does resveratrol, a polyphenolic compound, affect IVF-embryo transfer outcomes? DESIGN: This single-centre, cross-sectional retrospective study was designed to compare the outcomes of embryo transfer cycles in women receiving resveratrol supplementation (200 mg/day) continuously (RES group) with a control group (non-RES group). Of 8686 embryo transfer cycles, 1409 cycles with poor prognostic factors were excluded, including cycles in women aged ≥43 years and those with poor-quality embryos. The RES group (204 cycles, 102 women) was compared with the non-RES group (7073 cycles, 2958 women). RESULTS: After matching patients by age at the time of oocyte retrieval, grade and developmental stage of embryos, number of embryos transferred, and fresh or vitrified-warmed embryo transfer, multivariate logistic regression analysis showed that resveratrol supplementation is strongly associated with a decrease in clinical pregnancy rate [odds ratio (OR) 0.539, 95% confidence interval (CI) 0.341-0.853] and an increased risk of miscarriage (OR 2.602, 95% CI 1.070-6.325). CONCLUSIONS: Resveratrol supplementation during embryo transfer cycles appears to be detrimental for pregnancy outcomes. An analysis of the supplementation protocol and randomized controlled studies are needed.

Resveratrol inhibits decidualization by accelerating downregulation of the CRABP2-RAR pathway in differentiating human endometrial stromal cells.

Pregnancy critically depends on the transformation of the human endometrium into a decidual matrix that controls embryo implantation and placenta formation, a process driven foremost by differentiation and polarization of endometrial stromal cells into mature and senescent decidual cells. Perturbations in the decidual process underpin a spectrum of prevalent reproductive disorders, including implantation failure and early pregnancy loss, emphasizing the need for new therapeutic interventions. Resveratrol is a naturally occurring polyphenol, widely used for its antioxidant and anti-inflammatory properties. Using primary human endometrial stromal cell (HESC) cultures, we demonstrate that resveratrol has anti-deciduogenic properties, repressing not only the induction of the decidual marker genes PRL and IGFBP1 but also abrogating decidual senescence. Knockdown of Sirtuin 1, a histone deacetylase activated by resveratrol, restored the expression of IGFBP1 but not the induction of PRL or senescence markers in decidualizing HESCs, suggesting involvement of other pathways. We demonstrate that resveratrol interferes with the reprogramming of the retinoic acid signaling pathway in decidualizing HESCs by accelerating down-regulation of cellular retinoic acid-binding protein 2 (CRABP2) and retinoic acid receptor (RAR). Notably, knockdown of CRABP2 or RAR in HESCs was sufficient to recapitulate the anti-deciduogenic effects of resveratrol. Thus, while resveratrol has been advanced as a potential fertility drug, our results indicate it may have detrimental effects on embryo implantation by interfering with decidual remodeling of the endometrium.

Combination Treatment of Preoperative Embryo Cryopreservation and Endoscopic Surgery (Surgery-ART Hybrid Therapy) in Infertile Women with Diminished Ovarian Reserve and Uterine Myomas or Ovarian Endometriomas.

STUDY OBJECTIVE: To analyze the clinical outcomes of and predictive factors for the therapeutic effect of combination treatment of preoperative embryo cryopreservation and endoscopic surgery (surgery-assisted reproductive technology [ART] hybrid therapy) in infertile women with diminished ovarian reserve (DOR) with uterine fibroids and/or ovarian endometriomas. DESIGN: Retrospective cohort study. SETTING: Data from all patients who underwent surgery-ART hybrid therapy at Juntendo University Hospital and Sugiyama Clinic between 2014 and 2016 were analyzed retrospectively. We compared women who experienced live birth (success group) and implantation failure or miscarriage (failure group) after surgery-ART hybrid therapy and evaluate the predictive factors for live birth. PATIENTS: A total of 39 infertile women underwent surgery-ART hybrid therapy with 86 embryo transfer cycles. INTERVENTIONS: All women underwent ART treatment for embryo cryopreservation preoperatively, reproductive surgery, and warmed embryo transfer after the postoperative contraceptive interval (surgery-ART hybrid therapy) for women with DOR (anti-Müllerian hormone <1.0 ng/mL) and/or advanced reproductive age (>40 years) with uterine myomas and/or ovarian endometriomas who required surgery. RESULTS: Among 39 women underwent surgery-ART hybrid therapy, 1 woman acquired no embryo after oocyte retrieval trials and abandoned efforts to conceive, 14 experienced childbirth (success group) and 24 (63.2%) experienced implantation failure or miscarriage (failure group) after surgery-ART hybrid therapy. The median patient age was 40 years (interquartile range [IQR], 38-41 years) in the success group and 41.5 years (IQR, 41-42 years) in the failure group (p = .032). The respective serum anti-Müllerian hormone levels were 2.5 ng/mL (range, 0.1-8.6 ng/mL) and 1.3 ng/mL (range, 0.1-4.2 ng/mL) (p = .396), and the respective numbers of preoperative frozen were 5.0 (range, 4.0-6.0) and 2.0 (range, 1.0-3.0) (p < .001). There were no significant differences in surgical findings of myomas and endometriosis between the 2 groups. Compared with the 24 women who experienced hybrid therapy failure, the 14 who underwent successful surgery-ART hybrid therapy were significantly younger and had a greater number of cryopreserved embryos. CONCLUSION: Successful surgery-ART hybrid therapy requires a sufficient preoperative age-specific number of frozen embryos, establishment of ART treatment with stable pregnancy outcomes and skillful reproductive surgery, and a strong desire of the patient and doctor for pregnancy.

Reciprocal changes of H3K27ac and H3K27me3 at the promoter regions of the critical genes for endometrial decidualization.

AIM: Decidualization is essential for embryo implantation and placental development. We aimed to obtain transcriptome and epigenome profiles for primary endometrial stromal cells (ESCs) and in vitro decidualized cells. MATERIALS & METHODS: ESCs isolated from human endometrial tissues remained untreated (D0), or decidualized for 4 days (D4) and 8 days (D8) in the presence of 8-bromo-cAMP and progesterone. RESULTS: Among the epigenetic modifications examined (DNA methylation, H3K27ac, H3K9me3 and H3K27me3), the H3K27ac patterns changed most dramatically, with a moderate correlation with gene expression changes, upon decidualization. Subsets of up- and down-regulated genes upon decidualization were associated with reciprocal changes of H3K27ac and H3K27me3 modifications at their promoter region, and were enriched with genes essential for decidualization such as WNT4, ZBTB16, PROK1 and GREB1. CONCLUSION: Our dataset is useful to further elucidate the molecular mechanisms underlying decidualization.

Vitamin D Regulates Maternal T-Helper Cytokine Production in Infertile Women.

Vitamin D (VD) deficiency is associated with reproductive failure. However, the relationship between VD and maternal immunity remains unclear. We investigated the clinical efficacy of VD in maternal T-helper (Th) cytokines in 276 infertile women and examined for Th1 and Th2 cells based on the deficient, insufficient, and sufficient serum 25-hydroxyvitamin D3 (25[OH]VD) levels (<12, 12⁻30, and >30 ng/mL, respectively). Most infertile women had a low-level of VD (87.3%). Immunological tests of pre-/post-VD supplementation were performed in patients who were deficient and insufficient in VD. Of 23 patients, 11 (47.8%) exhibited sufficient VD levels after supplementation. Th1/Th2 cell ratio in patients with insufficient VD was significantly decreased after supplementation (p = 0.004). After supplementation, serum 25(OH)VD levels of the patients: 11 in the sufficient group showed significant decreases in Th1 cell level and Th1/Th2 cell ratio (p = 0.032 and 0.010, respectively), whereas no significant differences in Th1/Th2 cell ratio were recognized in the insufficient group. Furthermore, mid-luteal endometrial biopsies (n = 18) were processed for primary cultures and measured interferon [IFN]-γ and interleukin [IL]-4 in condition media. Decidualizing cultures with 1,25-dihydroxvitamin D3 (1,25[OH]2VD) decreased IFN-γ. Sufficient VD supplementation in women with insufficient VD may optimize maternal T-helper cytokines during pregnancy via rebalancing the Th1/Th2 cell ratio.

Reprogramming of the retinoic acid pathway in decidualizing human endometrial stromal cells.

Upon breaching of the endometrial surface epithelium, the implanting embryo embeds in the decidualizing stroma. Retinoic acid (RA), a metabolite of vitamin A, is an important morphogen during embryonic and fetal development, although the role of the RA pathway in the surrounding decidual cells is not understood. Here we show that decidual transformation of human endometrial stromal cells (HESCs) results in profound reprogramming of the RA signaling and metabolism pathways. Differentiating HESCs downregulate the intracellular carrier proteins CRABP2 and FABP5, responsible for transfer and binding of RA to the nuclear receptors RAR and PPARß/δ, respectively. Furthermore, the expression of RAR, the receptor that mediates the pro-apoptotic effects of RA, was also inhibited. By contrast, PPARß/δ, which transduces the differentiation responses of RA, was upregulated. Decidualization was also associated with increased expression of retinol-binding protein 4 (RBP4) and various enzymes involved in the metabolism of RA and its precursor, retinaldehyde (Rald), including CYP26A1, DHRS3, and RDH12. Exposure of differentiating HESCs to RA or Rald reversed the inhibition of the CRABP2-RAR pathway, perturbed the expression of decidual marker genes and triggered cell death. Taken together, the data demonstrate that decidualizing HESCs silence RA signaling by downregulating key cytoplasmic binding proteins and by increasing retinoid metabolism. However, excessive RA exposure is toxic for decidual cells and triggers a response that may lead to pregnancy failure.

The establishment of appropriate methods for egg-activation by human PLCZ1 RNA injection into human oocyte.

Phospholipase C-zeta (PLCZ1), a strong candidate of egg-activating sperm factor, can induce Ca2+ oscillations and cause egg activation. For the application of PLCZ1 to clinical use, we examined the pattern of Ca2+ responses and developmental rate by comparing PLCZ1 RNA injection methods with the other current methods, such as cytosolic aspiration, electrical stimulation and ionomycin treatment in human oocytes. We found that the pattern of Ca2+ oscillations after PLCZ1 RNA injection exhibited similar characteristics to that after ICSI treatment. We also determined the optimal concentration of human PLCZ1 RNA to activate the human oocytes. Our findings suggest that human PLCZ1 RNA is a better therapeutic agent to rescue human oocytes from failed activation, leading to normal and efficient development.

Minimum ovarian stimulation involving combined clomiphene citrate and estradiol treatment for in vitro fertilization of Bologna-criteria poor ovarian responders.

AIM: The aim of this study was to assess the efficacy of a minimal ovarian stimulation involving combined clomiphene citrate (CC) and estradiol (E2) administration for poor responders with diminished ovarian reserve (DOR). MATERIAL AND METHODS: In this case-control study, we recruited 41 consecutive hypergonadotropic poor responders (69 cycles) who met Bologna-criteria and had experienced cancellation of oocyte retrieval. In 10 (20 cycles), 11 (21 cycles) and 20 patients (28 cycles) between 2012 and 2014, follicular development was induced using an E2 cycle, CC cycle and CC + E2 cycle, respectively. After confirmation of high follicle-stimulating hormone levels (15-40 mIU/ml) at menstrual day 3, DOR patients were treated with oral E2 of 1.0 mg/day, CC of 100 mg/day, or both CC and E2 continuously, until ovulation induction. Two days later, we transvaginally aspirated the follicles, performed in vitro fertilization, and cryopreserved the cleavage embryos. One warmed embryo was transferred into the uterus during the hormone replacement cycles. RESULTS: For the E2, CC, and CC + E2 cycles, the median patient age was 41 years in all groups, and the serum anti-Müllerian hormone levels were 0.2 ± 0.3, 0.4 ± 0.4, and 0.2 ± 0.3 ng/mL, respectively (P = 0.258); follicular development failure rates were 50.0%, 19.0%, and 3.6%, respectively (P < 0.001); numbers of retrieved oocytes (/cycle) were 0.5 ± 0.6, 0.8 ± 0.7, and 1.2 ± 1.1, respectively (P = 0.033); and clinical pregnancy rates (/cycle) were 5.0%, 4.8%, and 10.7%, respectively (P = 0.725). CONCLUSION: CC + E2 administration for the patients with DOR was effective with a lower cancellation rate of oocyte retrieval and a higher number of retrieved oocytes.

Fourteen babies born after round spermatid injection into human oocytes.

During the human in vitro fertilization procedure in the assisted reproductive technology, intracytoplasmic sperm injection is routinely used to inject a spermatozoon or a less mature elongating spermatid into the oocyte. In some infertile men, round spermatids (haploid male germ cells that have completed meiosis) are the most mature cells visible during testicular biopsy. The microsurgical injection of a round spermatid into an oocyte as a substitute is commonly referred to as round spermatid injection (ROSI). Currently, human ROSI is considered a very inefficient procedure and of no clinical value. Herein, we report the birth and development of 14 children born to 12 women following ROSI of 734 oocytes previously activated by an electric current. The round spermatids came from men who had been diagnosed as not having spermatozoa or elongated spermatids by andrologists at other hospitals after a first Micro-TESE. A key to our success was our ability to identify round spermatids accurately before oocyte injection. As of today, all children born after ROSI in our clinic are without any unusual physical, mental, or epigenetic problems. Thus, for men whose germ cells are unable to develop beyond the round spermatid stage, ROSI can, as a last resort, enable them to have their own genetic offspring.

Menstrual cycle could affect Ki67 expression in estrogen receptor-positive breast cancer patients.

BACKGROUND: Ki67 is a potent prognostic marker for determining systemic treatment of patients with hormone receptor-positive breast cancer. However, evaluation of Ki67 expression can be difficult, due mostly to its heterogeneity. The Ki67 expression level, which indicates that a cell is undergoing division (cell cycle), rises when proliferation activity increases. Thus, Ki67 expression might be affected by hormonal stimuli. We hypothesised that Ki67 expression level might change during the menstrual cycle. We examined pairs of biopsy and surgical specimens from individual patients to evaluate this hypothesis. METHODS: First, the effects of estradiol on Ki67 expression in breast cancer cell lines were examined employing immunocytochemistry and Western blotting. Next, differences in Ki67 expression between biopsy and surgical specimens from 131 patients with estrogen receptor-positive tumours were retrospectively examined. RESULTS: In vitro experiments showed Ki67 expression in estrogen receptor-positive cancer cells to be dependent on estradiol stimulation. Ki67 expression was higher in biopsy samples collected in the luteal phase than in those from other phases. When biopsy and surgical samples were obtained at different times during the menstrual cycle in the same individual, there were differences in Ki67 expression between these samples. Those collected in the luteal phase showed higher Ki67 expression than samples obtained during other phases (p<0.01). CONCLUSIONS: Ki67 expression varied in the same patients according to menstrual cycle phase. Our results suggest that Ki67 expression in estrogen receptor-positive breast cancer should be carefully assessed bearing in mind the patient's menstrual cycle, since the interpretation of expression could affect treatment decisions.

Elevated periimplantation uterine natural killer cell density in human endometrium is associated with impaired corticosteroid signaling in decidualizing stromal cells.

BACKGROUND: Decidualizing human endometrial stromal cells (HESCs) profoundly up-regulate 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1), the enzyme that converts inert cortisone to active cortisol. We postulated that the induction of a cortisol gradient upon decidualization of the periimplantation endometrium may impact on the uterine natural killer (uNK) cell population and on local expression of corticosteroid-dependent target genes. METHODS: Midluteal endometrial biopsies (n = 55) were processed for uNK cell (CD56) analysis and primary HESC cultures. The cultures remained either untreated or were decidualized for 4 or 8 days. A tissue microarray was constructed from endometria with normal (n = 18) and elevated uNK cell (n = 18) scores. An abnormal uNK cell test was defined as greater than 5% CD56(+) cells in the subluminal stroma. RESULTS: Increased uNK cell density was associated with lower endometrial expression of 11ßHSD1 and mineralocorticoid receptor (MR) but not glucocorticoid receptor in vivo. Elevated uNK cell density also corresponded to impaired induction of key decidual markers (11ßHSD1, prolactin, and insulin-like growth factor binding protein-1) and MR-dependent enzymes (dehydrogenase/reductase member 3 and retinol saturase) in differentiating HESC cultures. Increased uNK cell density in vivo was not associated with increased in vitro expression of either IL-15 or IL-11, two cytokines implicated in uNK cell regulation. CONCLUSIONS: Elevated levels of uNK cells in the stroma underlying the surface epithelium are associated with inadequate cortisol biosynthesis by resident decidualizing cells and suboptimal induction of key MR-dependent enzymes involved in lipid biogenesis and the retinoid transport pathway. Our observations suggest that uNK cell testing identifies those women at risk of reproductive failure due to relative uterine cortisol deficiency.

Induction of 11ß-HSD 1 and activation of distinct mineralocorticoid receptor- and glucocorticoid receptor-dependent gene networks in decidualizing human endometrial stromal cells.

The actions of glucocorticoids at the feto-maternal interface are not well understood. Here, we show that decidualization of human endometrial stromal cells (HESCs) in response to progesterone and cAMP signaling is associated with a strong induction of 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1) expression and enzyme activity. Decidualization also triggered a gradual decrease in glucocorticoid receptor (GR) expression and reciprocal increase in mineralocorticoid receptor (MR) levels. Gene expression profiling of differentiating HESCs after small interfering RNA (siRNA)-mediated knockdown of either GR or MR identified 239 and 167 significantly regulated genes, respectively. Interestingly, GR-repressed genes were enriched for Krüppel-associated box domain containing zinc-finger proteins, transcriptional repressors involved in heterochromatin formation. In agreement, GR knockdown was sufficient to enhance trimethylated H3K9 levels in decidualizing cells. Conversely, we identified several MR-dependent genes implicated in lipid droplet biogenesis and retinoid metabolism. For example, the induction in differentiating HESCs of DHRS3, encoding a highly conserved enzyme that catalyzes the oxidation/reduction of retinoids and steroids, was enhanced by aldosterone, attenuated in response to MR knockdown, and abolished upon treatment with the MR antagonist RU26752. Furthermore, we demonstrate that decidualization is associated with dynamic changes in the abundance and distribution of cytoplasmic lipid droplets, the formation of which was blocked by RU26752. In summary, progesterone drives local cortisol biosynthesis by decidual cells through induction of 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1), leading to transcriptional regulation of distinct GR and MR gene networks involved in epigenetic programming and lipid and retinoid metabolism, respectively.

Histological assessment of impact of ovarian endometrioma and laparoscopic cystectomy on ovarian reserve.

AIM: The rate of oocyte decline follows a biphasic pattern, characterized by acceleration between 32 and 38 years old. Ovarian reserve is also affected by external factors, including ovarian disease and iatrogenic damage. The aim of this study was to histologically evaluate the impact of ovarian endometriomas, laparoscopic cystectomy, and age on follicle reserve in healthy ovarian tissues and in surgically resected cyst walls. MATERIAL AND METHODS: Sixty-one patients were found to have ovarian endometriomas and 42 patients non-endometriotic cysts. A small amount of normal ovarian tissue was obtained during ovarian cystectomy. The follicles in normal ovarian tissue and resected cyst walls were histologically evaluated. RESULTS: The density of follicles in ovarian tissues correlated with the age of the patients in both groups. In women aged <35 years, the relative density of follicles in healthy ovarian tissues was consistently lower in the endometriotic cyst group compared to the non-endometriotic cyst group, with the relative ratio at age 20, 30 and 35 years calculated to be 35.4%, 46.8% and 62.7%, respectively. There was no significant difference between the groups in patients over the age of 35. The resection rate of normal ovarian tissue in cystectomy specimen of the endometriosis group was significantly higher than in the non-endometriotic cyst group (P < 0.001). CONCLUSIONS: Our data suggest that ovarian endometriomas have a detrimental impact on follicle reserve in younger patients. Further, laparoscopic cystectomy for endometriomas may accelerate the rate of oocyte loss associated with aging.