Reproduction & Fertility最新文献

Preovulatory follicle growth and the luteal transition requires intense angiogenesis. This enables progesterone production to increase sufficiently to support a pregnancy. Inadequate follicular or luteal vascularisation can lead to reduced ovarian function and thus compromise fertility. Insulin-like growth factor 1 (IGF1) and IGF2 regulate multiple ovarian processes and are key links between an animal's reproductive and metabolic status. This study investigated the role that the IGF system plays in regulating luteinising follicular endothelial cell (EC) networks and progesterone production in vitro. Bovine luteinising follicular angiogenesis cultures were treated with 1) LR3-IGF1 (10 or 100ng/ml) under basal and angiogenic-stimulated conditions or 2) IGF1 receptor inhibitor (picropodophyllin (PPP); 1µM) in the presence or absence of LR3-IGF1, IGF2, or combined LR3-IGF1+IGF2 (10ng/ml). EC networks were quantified by von Willebrand factor immunohistochemistry. Progesterone production was analysed by ELISA and cell proliferation was determined by MTT assay. LR3-IGF1 had limited effects on EC growth parameters, whilst PPP (p<0.001) markedly reduced EC growth parameters (by 60-70%). Cell proliferation was slightly increased (by 3-5%) by LR3-IGF1 (p<0.001). LR3-IGF1 had variable effects on progesterone production, whilst PPP reduced progesterone concentration (p<0.001) with or without LR3-IGF1 or IGF2 alone or in combination. IGF1 was detected in cell conditioned media and was increased by LH (50ng/ml) (p<0.001). In conclusion, exogenous IGF1 and IGF2 had minimal effects on luteinising follicular angiogenesis and progesterone production, but the inhibitory effect of the IGFR1 inhibitor (PPP) suggests that IGF1 receptor signalling is critical for the development of EC networks and progesterone production in luteinising follicular cells.

A prospective longitudinal cohort study aimed to longitudinally examine the kinetics of Anti-müllerian hormone (AMH) during the first two trimesters of pregnancy. Pregnant women with stored 1st trimester serum samples were recruited at 24-28 weeks gestation during their gestational diabetes testing, where they provided an additional serum sample. The samples were analysed for AMH, oestradiol and progesterone concentrations. A decrease in serum AMH was observed in 40 out of 45 (88.9%) (95% CI 75.9% to 96.3%) of the participants in this study. The median serum AMH concentration was 10.9 pmol/L in the 1st trimester and 6.5 pmol/L during the 2nd trimester, with a significantly different distribution of the values between the 1st and the 2nd trimester AMH samples (p<0.001). The median percentage of AMH difference of -39.8%. This study demonstrated a significant decrease in serum AMH levels from the 1st to the 2nd trimester of pregnancy. The absolute decrease in AMH levels seems to be positively associated with 1st trimester AMH levels, whereas the percentage of AMH difference is not. Further studies are required to elucidate the potential physiological mechanisms of this finding.

Androgens are essential in normal ovarian function and follicle health but hyperandrogenism, as seen in polycystic ovary syndrome, is associated with disordered follicle development. There are few data on the effect of long-term exposure to high levels of testosterone as found in transgender men receiving gender-affirming endocrine therapy. In this study, we investigate the effect of testosterone on the development, morphological health and DNA damage and repair capacity of human ovarian follicles in vivo and their survival in vitro. Whole ovaries were obtained from transgender men (mean age: 27.6 ± 1.7 years; range 20-34 years, n = 8) at oophorectomy taking pre-operative testosterone therapy. This was compared to cortical biopsies from age-matched healthy women obtained at caesarean section (mean age: 31.8±1.5 years; range= 25-35 years, n=8). Cortical tissues were dissected into fragments and either immediately fixed for histological analysis or cultured for 6 days and subsequently fixed. Follicle classification and morphological health were evaluated from histological sections stained with H&E and expression of γH2AX as a marker of DNA damage by IHC. In uncultured tissue, testosterone exposure was associated with reduced follicle growth activation, poor follicle health and increased DNA damage. After 6 days of culture, there was enhanced follicle activation compared to control with further deterioration in morphological health and increased DNA damage. These data indicate that high circulating concentrations of testosterone have effects on the primordial and small-growing follicles of the ovary. These results may have implications for transgender men receiving gender-affirming therapy prior to considering pregnancy or fertility preservation measures.

To evaluate the proportion of chromosomal abnormalities in recurrent pregnancy loss (RPL) assisted by array comparative genomic hybridization (aCGH) bright out with higher detection rate, more accuracy, and less sample failure as compared with conventional cytogenetic analysis. In this study, product of conception samples with abnormal USG findings of the fetus and clinical history of RPL were first processed for karyotyping and Fluorescence In Situ Hybridization analysis. Normal results given by Karyotype and FISH samples with major anomalies detected by Ultrasound with RPL were divided into six groups and aCGH was performed to detect the gain or loss and copy number variations (CNVs) of a particular gene present in chromosomal segments. Among a total of 300 POC samples, 100 abnormal samples were identified either by karyotype (n=70) or by FISH (n=30). From the remaining 200 samples, 5 showed the presence of maternal cell contamination excluded. aCGH analysis revealed (n=195) that 74 (38%) samples with copy number variations (CNVs) and two samples with variants of unknown clinical significance (VOUS) were clinically associated with the clinical findings and 121(62%) samples showed no change in CNVs. The most frequent CNVs were loss of chromosome regions at 2q33.1, 7q11.21, 15q11.1, 16p11.2, Xp22.33, and Yp11.32. CNVs at arr[GRCh37]7p22.3,p21.2(830852-15124702)×1,7q34q36.3(141464180_158909738)×3, 14.2Mbp deletion of 7p22.3p21.2 (SUN1 gene) and 17.4Mbp duplication of 7q34q36.3 (KCNH2, CNTNAP2, and SHH genes) in one sample, CNVs at arr[GRCh37]8p22.2q22.3 (86326349_105509986)×1, 2.48Mbp deletion of 8p22.2q22.3 (GRHL1 gene) were found in another sample.

The hypothesis that CSF2 plays a role in the preimplantation development of the bovine embryo was tested by evaluating consequences of inactivation of CSF2RA (the functional receptor in the embryo) for development of embryos in utero. CRISPR/Cas9 was used to alter sequences on exon 5 and intron 5 of CSF2RA, Control embryos were injected with Cas9 mRNA only. Embryos > 16 cells at day 5 after insemination were transferred to synchronized recipient females in groups of 7 to 24. Embryos were flushed from the uterus two days later. The proportion of recovered embryos that developed to the blastocyst stage was lower for knockout embryos (39%) than for control embryos (63%). RNA sequencing of individual morulae and blastocysts indicated a total of 27 (morula) or 15 (blastocyst) differentially-expressed genes (false discovery rate <0.05). Gene set enrichment analysis indicated that the knockout affected genes playing roles in several functions including cell signaling and glycosylation. It was concluded that signaling through CSF2RA is not obligatory for development of the bovine preimplantation embryo to the blastocyst stage but that CSF2 signaling does enhance the likelihood that the embryo can become a blastocyst and result in specific changes in gene expression.

Kisspeptin, a hypothalamic neuropeptide encoded by the KISS1 gene, has a pivotal role in promoting GnRH secretion in mammals. Kisspeptin and its receptor (KISS1R) are also expressed in certain peripheral tissues including gonads, suggesting intra-gonadal roles. Such actions at the level of the bovine ovary have not been explored previously. The current aims were to determine whether KISS1 and its receptor (KISS1R) are expressed in the bovine ovary and whether kisspeptin or a kisspeptin antagonist can modulate ovarian steroid production by cultured ovarian cells. Granulosa (GC) and theca interna (TC) were collected from antral follicles (3-18 mm) categorized into five class sizes. Early, mid and regressing corpora lutea (CL) were also collected for RT-qPCR analysis of KISS1 and KISS1R expression. Bovine TC and GC cultured under both non-luteinizing (serum-free) and luteinizing (serum-supplemented) conditions were treated for 4 days with kisspeptin-10 (10-10-10-6M) or kisspeptin antagonist (p234; 10-10-10-6M), alone and in combination with either FSH (GC), LH (TC) or forskolin (luteinized GC/TC). Steroid secretion (GC: oestradiol, progesterone; TC: androstenedione, progesterone; luteinized GC/TC: progesterone) was measured by ELISA and viable cell number determined by neutral red uptake assay. KISS1 and KISS1R transcripts were detected in TC, GC and CL with significant differences between follicle categories and CL stages. However, neither kisspeptin-10 nor kisspeptin antagonist affected steroid secretion or viable cell number in any of the four ovarian cell culture models. As such, the hypothesis that kisspeptin has a direct intra-ovarian role to modulate follicular or luteal steroidogenesis, or cell proliferation/survival, is not supported.

In the present work, we emphasize the studies about integrins and their receptors in pig placental interface at different times of gestation. Uterine placental interface (n=24) of 17-, 30-, 60- and 70-days gestation (dg) and non-pregnant uterus (n=4) of crossbred sows were used. The presence of αvβ3 and α5β1 integrins, and their ligands fibronectin (FN), and osteopontin (OPN) were detected by immunohistochemistry, and the immunolabelled area percentage (IAP) and the optical density (OD) were determined. The integrins and its ligands analyzed have presented peaks of expression in early and mid-gestation, both in IAP and the OD area decreasing at 70 dg. These temporal changes showed us that the molecules studied in this work participate in embryo/feto-maternal attachment, variably. Besides, we found a significant correlation both in the intensity and in the extension of immunostaining for trophoblastic FN and endometrial αvβ3, and trophoblastic OPN and endometrial α5β1, throughout the entire pig pregnancy. At late gestation, take place a notable placental remodelation with subsequent removal or renewal of folds at the uterine-placental interface that results in the loss of focal adhesions. The decrease of the expression of some integrins and their ligands in late gestation, particularly at 70 dg, would demonstrate that there would be other adhesion molecules and other ligands that could be participating in the establishment of the maternal-fetal interface.

Background: The distribution of the blood vessel network at any point in time in any body tissue, may provide valuable information with regards to the tissue condition and its angiogenesis functionality. The blood vessel three-dimensional network of the endometrium goes through a process of change over a relatively short period of 4 weeks on average. It is well accepted that this angiogenesis is closely related to the success or failure of the implantation of the embryo Objective and rationale: Our study aims to present a method to follow the three-dimensional evolution of the superficial blood vessel distribution in the endometrium throughout the uterine cycle.

Method: This method utilizes differences in the observed broadband colors of the blood vessels in order to assess their depth coordinate below the endometrial tissue surface. We implemented the method using microscopic images of fresh, ex-vivo, endometrial samples of different cycle days to obtain the statistical evolution track of the superficial blood vessel population in both human and animal (swine) samples.

Outcomes: In human samples we observed a systematic and consistent trend in the BV diameter distribution at different tissue depths. We demonstrate that the magnitude of this trend evolves throughout the course of the female cycle.

Wider implications: This method has the potential to further our understanding of the mechanisms of angiogenesis in tissues other than the endometrium. We propose that this method may also contribute to more precise endometrial dating and may assist in more accurate determination of embryo transfer timing within IVF treatments.

Treatment of sub-fertile women aged ≥ 40 years old (AMA) is challenging. Co-treatment with growth hormone (GH) is suggested to improve reproductive outcomes in poor responders. However, few studies, and with conflicting results, focused on women with AMA. A systematic review and meta-analysis of randomized controlled trials (RCTs) and comparative retrospective trials (CRTs) of GH cotreatment in AMA women undergoing in vitro fertilization or intracytoplasmic injection treatment using their autologous oocytes was performed. The search included studies published in English up to the end of 2021. The primary outcome was the clinical pregnancy rate per embryo transfer. Secondary outcomes were the number of mature and retrieved oocytes and the rate of live birth. 406 studies were found. The final analysis included three RCTs and four CRTs with 481 patients who used GH and 400 patients who did not. Clinical pregnancy and live birth rates were significantly higher in the GH cotreatment group compared to the placebo as well as the group without GH co-treatment, (OR 2.2; 95% CI 1.34 - 3.61 and OR 4.12; 95% CI 1.82 - 9.32, respectively). Intriguingly, the subgroup analysis showed that poor-responder patients did not benefit from co-treatment with GH. There were no statistically significant differences in the number of mature or retrieved oocytes. GH cotreatment in a subgroup of women with AMA improves clinical pregnancy and live birth per fresh embryo transfer. However, this conclusion must be taken with caution and further research is needed. The review is registered in PROSPERO database (CRD42021252618). www.crd.york.ac.uk/prospero/.