Molecular human reproduction最新文献

Infertility impacts up to 17.5% of reproductive-aged couples worldwide. To aid in conception, many couples turn to ART, such as IVF. IVF can introduce both physical and environmental stressors that may alter DNA methylation regulation, an important and dynamic process during early fetal development. This meta-analysis aims to assess the differences in the placental DNA methylome between spontaneous and IVF pregnancies. Potential datasets were identified by searching the NCBI Gene Expression Omnibus (GEO) using keywords related to IVF in human participant studies published before November 2023. In our combined fetal sex population (N = 575) from three eligible GEO datasets, 127 autosomal cytosine guanine dinucleotides (CpGs) were significant (False Discovery Rate (FDR) <0.05) between IVF (n = 96) and spontaneous (n = 479) placentae, with 47 CpGs considered differentially methylated (FDR < 0.05 and |Δβ| > 0.05). Stratification by fetal sex revealed no significant autosomal CpGs in fetal female placentae (N = 281); however, in the fetal male placentae (N = 294), we identified nine autosomal CpGs that reached statistical significance between IVF (n = 56) and spontaneous (n = 238) placentae, with three CpGs considered differentially methylated. Fetal male placentae had lower proportions of trophoblasts (P < 0.0001) and stromal cells (P = 0.007) and higher proportions of syncytiotrophoblasts (P = 0.0001) compared to fetal female placentae, regardless of conception type. IVF placentae had higher proportions of stromal cells (P = 0.01) and lower proportions of syncytiotrophoblasts (P = 0.01) compared to spontaneous placentae, regardless of sex. Controlling for cell-type proportions in linear models reduced test statistic inflation and identified new significant CpGs that may previously have been masked by cell-type heterogeneity. The results of this meta-analysis are critical to further understand the impact of IVF on tissue epigenetics, which may help with understanding the connections between IVF and negative pregnancy outcomes. Additionally, our study suggests that sex-specific differences in placental DNA methylation and cell composition should be considered as factors for future placental DNA methylation analyses.

Premature ovarian insufficiency (POI) is a major cause of female infertility, for which effective therapies remain limited. S14G-Humanin (HNG), a potent analogue of Humanin, exhibits strong antioxidant and anti-apoptotic properties and has demonstrated cytoprotective effects in various tissues, including the ovary. In this study, a cyclophosphamide (CP)-induced POI mouse model was established to evaluate both the ovarian damage induced by chemotherapy and the protective effects of HNG. HNG administration significantly increased the number of primordial follicles (P = 0.044) and growing follicles (all P > 0.05), as well as corpora lutea (P = 0.09). Moreover, HNG markedly improved oocyte quality (P = 0.009), significantly lowering the proportion of abnormal ovulated oocytes (P = 0.002). Fertility outcomes were also enhanced: CP treatment significantly reduced litter size compared to controls (4.6 ± 1.1 vs 8.0 ± 1.0; P < 0.001), whereas HNG treatment significantly mitigated this reduction (6.2 ± 0.8 vs 4.6 ± 1.1; P = 0.029). Mechanistically, HNG alleviated oxidative stress and apoptosis in ovarian tissues (all P < 0.05), reduced ROS levels (P = 0.034), and restored mitochondrial membrane potential (P = 0.004) in a human granulosa cell line. Furthermore, HNG significantly upregulated PGC-1α expression and enhanced AMPK phosphorylation in both in vivo and in vitro models (both P < 0.05). Collectively, these findings demonstrate that HNG confers significant protection against chemotherapy-induced ovarian damage and highlight its potential as a novel therapeutic agent for chemotherapy-induced ovarian damage.

While advanced maternal age is associated with significant changes in oocyte gene expression, these are not global changes but limited to a fraction of the transcriptome. However, there is little consensus on the specific genes affected, and on the transcriptomic signatures of age-related declines in oocyte quality. To characterize the effects of age on the human MII oocyte transcriptome, here we take a two-part approach. We first generated single-oocyte Smart-seq2 datasets from 10 younger (21-29 years) and 10 older (37-43 years) donors, identifying genes differentially expressed between the two groups, then cross-referenced our results with those of 12 studies (9 human, 3 mouse) performing equivalent analyses using a variety of single-cell transcriptomic or microarray platforms. Technical differences notwithstanding, we found considerable discordance between the datasets, suggesting that age-related signatures of differential gene expression are not easily reproducible. Independent corroboration of age-associated changes in expression was limited to few genes, with the vast majority only supported by one of the 13 datasets, including our own. Nevertheless, we identified 40 genes whose expression significantly altered with age in multiple studies, highlighting common processes underlying ageing, including dysregulated proteostasis. As human Smart-seq2 oocyte libraries are challenging to procure and rare in public archives, we next implemented a meta-analytic method for their re-use, combining our 20 oocytes with 130 pre-existing libraries sourced from 12 different studies and representing a continuous age range of 18-43 years. We identified 25 genes whose expression level significantly correlated with age and corroborated 14 of these genes with RT-PCR, including the proteasomal subunits PSMA1 and PSMA2, both of which were downregulated in older oocytes. Overall, our findings are consistent with both pronounced inter-oocyte heterogeneity in transcription and with oocyte ageing being a multifactorial process to which bona fide transcriptomic changes may only play a restricted role, while proteomic changes play more pronounced roles.

In vitro follicle culture (IFC) is an emerging fertility preservation alternative for women and children with cancer. Because two-dimensional (2D) IFC results in oocytes of suboptimal quality in mice and cannot support follicle growth in humans, the search for an optimal three-dimensional (3D) method that preserves the follicular structure is ongoing, and both matrix-free and hydrogel encapsulation systems are being explored. Our aim was to compare several 3D mouse IFC systems, including matrix-free and hydrogel encapsulation approaches. Secondary follicles were cultured for 12 days in a matrix-free non-attachment (NA) system, a Poly-Ethylene-Glycol (PEG) hydrogel, an extracellular-matrix-derived soft hydrogel (ES), and a 2D attachment (AT) control. We assessed follicle growth, survival, hormone secretion, theca cell localization, oocyte meiotic competence and diameter, gene expression in oocytes and cumulus cells, as well as oocyte fertilization potential. Metaphase II oocyte rates were significantly higher in the NA (75 ± 12.4%, n = 79) and AT systems (77 ± 12.6%, n = 109) compared to the ES (33.4 ± 9.5%, n = 40, P < 0.01), while low antral follicle rates from the PEG system led to its exclusion from the comparison. Similarly, following IVF, 2-cell rates were significantly higher in the NA (47.7 ± 17.6%, n = 147, P < 0.01) and AT (40.2 ± 9.7%, n = 132, P < 0.05) systems compared to the ES (23.5 ± 9.3%, n = 63). Furthermore, cumulus cells from the NA condition displayed a more in vivo-like gene expression profile than other conditions. No differences were detected in follicle survival, oocyte diameter, blastocyst rate, or quality between conditions. Lastly, we observed major differences in theca cell localization and hormone secretion levels that require further investigation. Our findings demonstrate the efficiency of the NA system over complex encapsulation methodologies, as it enhanced oocyte meiotic and developmental competence compared to the ES. However, as the study is limited by the lack of human data and the use of Fetal Bovine Serum (FBS) in the culture medium, further research is required to translate our findings to humans.

Spermatogonial stem cells (SSCs) serve as the foundational units for all adult male germ cells, playing a critical role in male fertility. These cells are characterized by their unique ability to balance self-renewal and differentiation, ensuring the maintenance of tissue homeostasis while simultaneously producing mature spermatozoa. Recent studies have highlighted the pivotal role of fibroblast growth factor (FGF) signaling in regulating SSC self-renewal, with various FGFs and their corresponding FGF receptors (FGFRs) being implicated in the intricate processes governing SSC homeostasis. This review aims to provide a comprehensive overview of the characteristic expression patterns of FGFs and FGFRs within germ cells and their surrounding somatic cells. In addition, we will conduct in-depth research on the multifaceted functions and regulatory mechanisms of FGF signaling in SSCs and elucidate its potential clinical significance. Understanding these mechanisms not only enhances our knowledge of male reproductive biology but also opens avenues for innovative clinical applications aimed at safeguarding fertility in vulnerable populations.

The aetiology of endometriosis remains poorly understood. In vitro model systems provide the opportunity to identify the mechanisms driving disease pathogenesis using human cells. Three-dimensional models, particularly organoid systems, have revolutionized how we study epithelial biology and are powerful tools for modelling endometriosis. As an emerging model system, it is important to define protocols and identify the remaining challenges surrounding endometrial organoid culture to increase reproducibility and scientific rigour in endometriosis research. The World Endometriosis Research Foundation (WERF) established an international working group comprised of experts using in vitro approaches for the study of endometriosis. This working group harmonized protocols and documentation of existing and emerging organoid systems to maximize comparison and replication across the field and guide specific research hypotheses testing. This evaluation of organoid protocols, limitations, challenges, and alternative approaches assessed both published and grey literature papers across several disciplines pertinent to endometriosis research. Recommendations for protocol and documentation harmonization are presented, and we created the first-ever decision tree diagram to guide and facilitate the selection of existing models best suited for specific areas of endometriosis research. Rigorous and systematic assessment of emerging organoid systems, recognizing the inferential strengths and limitations of these approaches, is vital for endometriosis research. This comprehensive review of the benefits, limitations, and utilization of organoid models, as well as the consequent integration of protocols and documentation, will contribute to the scientific knowledge base by maximizing the reproducibility, comparability, and interpretation of research studies in endometriosis. Additionally, these newly developed protocols and documentation should serve as a resource for, and facilitate collaboration between, endometriosis investigators using organoids in their research methods.

Maternal diet-induced obesity (DIO) may affect adult offspring oocyte quality due to mitochondrial dysfunction. Here, we investigated whether offspring of DIO mothers exhibit mitochondrial abnormalities in their primordial follicle oocytes (PFOs) already at birth and if (further) alterations can be detected at weaning. Female Swiss mice were fed a control or obesogenic diet for 7 weeks before mating and throughout pregnancy and lactation. Offspring ovaries were collected at birth and at weaning. Offspring PFOs were examined by transmission electron microscopy of ovarian sections. Key markers of cell stress (HSP70), mitochondrial biogenesis (PGC-1α), mtDNA replication (TFAM), fusion (MFN2, OPA1), and fission (DRP1) were examined using immunofluorescence and confocal microscopy. Maternal DIO did not alter HSP70 or PGC-1α expression in the PFOs at birth, suggesting that cellular homeostasis and mitochondrial biogenesis were unaffected. TFAM expression was reduced at both time points. DRP1 and cytoplasmic OPA1 expression were reduced at birth, but without ultrastructural changes in mitochondrial shape and density, suggesting that these alterations are regulatory. No inborn mitochondrial structural abnormalities could be detected. In contrast, at weaning, offspring born to and nursed by DIO mothers exhibited a high number of lipid droplets (LDs) in their ovaries, some of which were detectable in the PFOs, while no LDs were detected in the PFOs of the controls. Maternal DIO increased PGC-1α expression, suggesting postnatal effects on PFO mitochondrial biogenesis. MFN2 and OPA1 expression also increased, together with increased mitochondrial elongation and a reduced mitochondrial density. Mitochondrial abnormalities, such as vacuolation, loose inner membranes, the number of detected autophagosomes, and signs of lipophagy, were also significantly increased by maternal DIO at weaning. In conclusion, the oocyte mitochondrial structural abnormalities previously reported in adult offspring from DIO mothers were not detected in the PFOs at birth. Significant changes in primordial follicles linked to maternal DIO were detected only at weaning.

Endometriosis, defined as the growth of endometrial-like tissues outside the uterus, is a common disease among women. Numerous in vivo rodent models of endometriosis have been developed to explore multiple aspects of this poorly understood disease. Heterologous models utilize human endometrial tissues engrafted into immunocompromized mice, while homologous models engraft rodent endometrium into immunocompetent mice or rats. Heterologous models of endometriosis more closely replicate the human disease; however, the murine humoral immune response must be suppressed to prevent rejection of the xenograft tissue. Although the innate immune system remains intact, suppression of the humoral response leads to a markedly different local and systemic immune environments compared to humans. Despite this limitation, experiments using heterologous models have contributed significantly to our understanding of endometriosis establishment and progression, the pre-clinical effectiveness of various therapeutic strategies, and genetically modifiable host factors that contribute to disease. Unfortunately, a lack of harmonization of the models used by different laboratories has impeded the reproducibility and comparability of results between groups. Therefore, the World Endometriosis Research Foundation (WERF) formed an international working group of experts in heterologous models of endometriosis to develop guidelines and protocols that could contribute to unifying experimental approaches across laboratories. Nine critical variables were identified: (i) mouse strain; (ii) human tissue type; (iii) hormonal status of the human tissue donor; (iv) human tissue preparation; (v) method and location of tissue placement; (vi) hormonal status of the recipient animal; (vii) whether or not mice were engrafted with human immune cells; (viii) endpoint assessments; and (ix) number and type of replicates. Herein, we outline important considerations for each major variable and make recommendations for unification of approaches. Widespread adoption of harmonized protocols and implementation of standardized documentation and reporting should further improve the reproducibility and translation of experimental findings both within and between laboratories.

In vivo models of endometriosis enable the discovery and preclinical testing of new therapies. Several rodent models of endometriosis exist, but a lack of harmonization impedes reproducibility and comparability of results among investigators. Homologous models are advantageous as they allow the contribution of the immune system/inflammation to be studied. We reviewed published homologous rodent models of endometriosis to develop standard operating procedures ('EPHect-EM-Homologous-SOPs') to guide and facilitate the choice and implementation of these models and harmonize documentation to enhance interpretation and comparability of results. The World Endometriosis Research Foundation (WERF) established an international working group of experts in models of endometriosis and formed a working sub-group to discuss homologous rodent models of endometriosis. A systematic literature review and detailed analysis of protocols was performed. The identified models have advantages and limitations regarding physiological relevance and utility. To harmonize key variables for endometriosis rodent models, the working group focused on species and animal strains, placement of ectopic tissue, uterine tissue volume, method of induction, hormonal status, and uterine tissue 'type'. A decision tree and recommendations on model use were developed for mice and rats to serve as guides for the use of harmonized EPHect-EM-Homologous-SOPs, experimental design, reporting standards, and research of question-dependent key variables. No 'ideal' homologous model of endometriosis was identified. The choice of model for specific research should be guided according to a best-fit strategy. Harmonization of SOPs, documentation, and reporting standards will improve replicability and translational applicability of studies and better highlight where de novo model creation is needed.