Infertility treatment strategy involving combined freeze-all embryos and single vitrified-warmed embryo transfer during hormonal replacement cycle for in vitro fertilization of women with hypogonadotropic hypogonadism.

AIM: Hypogonadotropic hypogonadism (HH) is a condition caused by the deficient secretion of pituitary gonadotropins, leading to diminished ovarian function. Several studies of in vitro fertilization (IVF) in women with HH revealed acceptable clinical pregnancy outcomes but high multiple pregnancy rates after multiple fresh embryo transfer (ET). The purpose of this study was to analyze the outcomes of combined freeze-all embryos and single vitrified-warmed ET in women with HH. METHODS: Of 91 infertile women with HH (basal luteinizing hormone and follicle-stimulating hormone levels <2.0 mIU/mL), we excluded patients aged ≥40 years (n = 2) and women who preferred fresh ET (n = 10). Seventy-nine women underwent 117 oocyte retrieval cycles and 135 vitrified-warmed ET during hormone replacement (HR) cycles from 2008 to 2014 at the Kato Ladies Clinic and Juntendo University Hospital. RESULTS: In 26 single cleavage ET cycles, the rates of clinical pregnancy and live birth were 34.6% (9/26 ET) and 26.9% (7/26 ET), respectively. Regarding the outcomes after single vitrified-warmed blastocyst transfer, clinical pregnancy and live birth rates were 65.1% (71/109 ET) and 50.5% (55/109 ET), respectively. Multiple conceptions and ovarian hyperstimulation syndrome did not occur in any of the women with HH. CONCLUSION: Our results demonstrated that IVF followed by single vitrified-warmed ET in adjusted endocrine milieu during the HR cycle is an effective fertility treatment for women with HH and decreases the incidence of complications, including multiple conceptions.

An improved method for isolation of epithelial and stromal cells from the human endometrium.

We aimed to improve the efficiency of isolating endometrial epithelial and stromal cells (EMECs and EMSCs) from the human endometrium. We revealed by immunohistochemical staining that the large tissue fragments remaining after collagenase treatment, which are usually discarded after the first filtration in the conventional protocol, consisted of glandular epithelial and stromal cells. Therefore, we established protease treatment and cell suspension conditions to dissociate single cells from the tissue fragments and isolated epithelial (EPCAM-positive) and stromal (CD13-positive) cells by fluorescence-activated cell sorting. Four independent experiments showed that, on average, 1.2 × 10(6) of EMECs and 2.8 × 10(6) EMSCs were isolated from one hysterectomy specimen. We confirmed that the isolated cells presented transcriptomic features highly similar to those of epithelial and stromal cells obtained by the conventional method. Our improved protocol facilitates future studies to better understand the molecular mechanisms underlying the dynamic changes of the endometrium during the menstrual cycle.

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.

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.