Search Results (24)

Premature ovarian insufficiency (POI) is a clinical condition characterized by loss of ovarian function indicated by amenorrhea or irregular menstrual cycles for at least 4 months and elevated gonadotrophins (FSH > 25 IU/L, measured on one occasion) and low estrogen serum levels in women under the age of 40. Premature ovarian insufficiency can be non-iatrogenic or spontaneous (idiopathic or due to genetic, autoimmune, or metabolic reasons, or infections) and iatrogenic (a consequence of oophorectomy, chemotherapy, radiotherapy, or uterine artery embolization). Women with POI are faced not only with estrogen deficiency but also with infertility and psychological implications. Hormonal replacement therapy is effective in treating the symptoms of premature ovarian insufficiency as well as in lowering the health risk of long-term consequences of premature ovarian insufficiency. Currently, oocyte donation is the standard treatment for patients with POI desiring pregnancy. Recently developed methods for the regeneration of ovarian tissue, such as stem cell therapy, platelet-reach plasma therapy and in vitro activation of ovarian tissue, are still under research and further adequate multicentric clinical studies are needed to develop standardized effective and safe protocols for the infertility treatment of patients with premature ovarian insufficiency. Full article

Ovarian function relies on a network of well-coordinated molecular mechanisms that regulate follicular development, oocyte maturation, ovulation, and corpus luteum function. When these processes are disrupted, infertility can result. Extracellular matrix (ECM) remodeling represents a central regulatory component in these processes and is essential for follicle rupture and oocyte release. This mechanism involves metalloproteinases (MMPs), mainly MMP-2 and MMP-9, which degrade the ECM and allow the necessary structural changes. Other ECM-modulating proteases, such as ADAM and ADAMTS families, also contribute to this process. Their activity is tightly regulated by tissue inhibitors of metalloproteinases (TIMPs), ensuring that tissue remodeling occurs in a controlled manner. Disruption of the balance between MMPs and TIMPs increases the risk of infertility-related conditions such as polycystic ovary syndrome (PCOS), endometriosis, luteinizing hormone (LH) deficiency syndrome, and ovarian aging. In addition to the ECM, other factors, including intracellular signaling pathways, oxidative stress (OS), and mitochondrial function, contribute to ovarian physiology and directly affect oocyte quality and viability. This narrative review focuses on the molecular mechanisms governing ovarian function, with particular emphasis on the remodeling of the ECM by MMPs during ovulation, and examines how their disorders contribute to infertility. A deeper understanding of these mechanisms may lead to the identification of new therapeutic targets and the improvement of assisted reproduction outcomes. Full article

Fertilisation failure following intracytoplasmic sperm injection (ICSI) is a significant challenge in assisted reproductive technology (ART), particularly in the absence of an identifiable cause. Artificial oocyte activation (AOA), typically with calcium ionophores, has emerged as a potential solution in scenarios characterised by a deficiency of phospholipase C zeta (PLCζ). This narrative review consolidates the latest clinical and experimental data regarding the application of calcium ionophores for oocyte activation, the significance of PLCζ testing in instances of unexplained fertilisation failure, and the impact of AOA on the morphokinetics and developmental potential of embryos. AOA has demonstrated an enhancement in fertilisation, cleavage, and pregnancy outcomes in specific patient populations, including individuals with diminished ovarian reserve or those who have previously attempted conception unsuccessfully. Although AOA appears to have no impact on embryo morphokinetics, certain studies indicate slight alterations in early cleavage features. The available statistics indicate that there are no significant safety concerns about outcomes for babies. This finding underscores the significance of tailored ART methodologies that incorporate molecular diagnostics and targeted AOA therapies. It emphasises the necessity for additional prospective trials to enhance patient selection and long-term safety surveillance. Full article

Organ functions generally decline with age, but the ovary is a prototypical organ that undergoes functional loss over time. Autophagy plays a crucial role in maintaining organ homeostasis, and age-related upregulation of the autophagy inhibitor protein, Rubicon, has been linked to cellular and tissue dysfunction. This review describes how granulosa cell autophagy supports follicular growth and oocyte selection and maturation by regulating cellular energy metabolism and protein quality control. We then introduce the role of selective autophagy, including mitophagy or lipophagy, in steroidogenesis and cellular remodeling during luteinization. In aged ovaries, Rubicon accumulation suppresses autophagic flux, leading to diminished oxidative-stress resilience and enhanced DNA damage. Moreover, impaired autophagy drives the accumulation of ATP citrate lyase, which correlates with poor oocyte quality and reduced ovarian reserve. Following fertilization, oocytes further upregulate autophagy to provide the energy required for blastocyst transition. Conversely, in infertility-related disorders, such as premature ovarian insufficiency, endometriosis, and polycystic ovary syndrome, either deficient or excessive autophagy contributes to disease pathogenesis. Both autophagy inhibitors (e.g., Rubicon) and activators (e.g., Beclin1) could be emerging as promising biomarkers for assessing ovarian autophagy status. Therapeutically, Rubicon inhibition by trehalose in aged ovaries and autophagy suppression by agents such as hydroxychloroquine in polycystic ovary syndrome and endometriosis hold potential. Establishing robust methods to evaluate ovarian autophagy will be essential for translating these insights into targeted treatments. Full article

Abdominal fat necrosis is a dystrophic–necrotic process that is relatively common in dairy cows. It is determined by productive strain (excess fat in the diet), negative energy balance after calving, a lack of physical activity, vitamin E and selenium deficiency, etc. Lipomatous masses are predominantly located in the omentum and mesentery in cattle, potentially causing intestinal obstruction. We report on an outbreak of abdominal fat necrosis that affected 135 of 220 cows and heifers (61.36%); this involved massive fat accumulation in the uterine and salpingian ligaments and severe reproductive disorders (reducing fertility to 20% in cows and 10% in heifers) caused by a hyperenergetic diet (supplementation with saturated fats). A transrectal ultrasound examination of the genital apparatus—both in heifers and in cows in the puerperium—revealed a diffuse pathological hyperechogenicity of the cervical folds, suggesting lipid infiltration, proliferation of the endocervical folds and hyperechogenic lipogranulomas located paracervically or in the uterine ligaments. An ultrasound examination of the ovaries showed the presence of parasalpingial lipogranulomas on the mesovarium, with a uniformly pixelated greasy appearance, that altered the topography of the salpinx, leading to the impossibility of oocyte retrieval. At the histopathological examination, in addition to the necrosis of adipocytes and the subacute–chronic inflammation of the abdominal and retroperitoneal adipose tissue, lipid infiltration of the uterine walls was also observed in the uterine ligaments and lymph nodes. Additionally, lipid infiltration was observed in the wall of the uterine artery. All muscular-type branches of the ovarian artery exhibited subendothelial (subintimal) amyloid deposits, severely reducing their lumen and leading to ischaemia. Amyloidosis was secondary to the systemic inflammatory process triggered by lipid deposition and necrosis. Fertility returned to normal 45–60 days after the exclusion of fat supplements from the diet and their replacement with a vitamin–mineral supplement rich in antioxidants. Full article

In addition to the male genome, the fertilizing spermatozoon delivers to the oocyte several factors whose deficiency can cause embryo dysfunction. Sperm oocyte-activating factor, identified as phoshoplipase C zeta (PLCζ), drives oocyte exit from meiotic arrest through a signaling pathway initiated by periodic rises of free cytosolic Ca²⁺ concentration (calcium oscillations). Sperm centrioles, together with oocyte proteins, form centrosomes that are responsible for aster formation, pronuclear migration, and DNA polarization before nuclear syngamy and subsequent mitotic divisions. Sperm DNA fragmentation can be at the origin of aneuploidies, while epigenetic issues, mainly abnormal methylation of DNA-associated histones, cause asynchronies of zygotic gene activation among embryonic cells. Sperm long and short non-coding RNAs are important epigenetic regulators affecting critical developmental processes. Dysfunction of sperm PLCζ, centrioles, DNA, and RNA mostly converge to aneuploidy, developmental arrest, implantation failure, miscarriage, abortion, or offspring disease. With the exception of DNA fragmentation, the other sperm issues are more difficult to diagnose. Specific tests, including heterologous human intracytoplasmic sperm injection (ICSI) into animal oocytes, genetic testing for mutations in PLCZ1 (the gene coding for PLCζ in humans) and associated genes, and next-generation sequencing of sperm transcriptome, are currently available. Oral antioxidant treatment and in vitro selection of healthy spermatozoa can be used in cases of sperm DNA fragmentation, while ICSI with assisted oocyte activation is useful to overcome oocyte-activation defects. No clinically confirmed therapy is yet available for sperm RNA issues. Full article

Phospholipase C zeta (PLCζ) has emerged as a pivotal sperm-specific factor responsible for triggering oocyte activation, a process essential for successful fertilization and early embryogenesis. A narrative review was conducted to examine the molecular architecture and biochemical features of PLCζ, with particular emphasis on how its distinctive structural domains facilitate the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP₂) and the induction of calcium (Ca²⁺) oscillations in the oocyte. Notably, PLCζ exhibits unique sensitivity to basal Ca²⁺ levels and the capacity to sustain repetitive intracellular Ca²⁺ transients that drive meiotic progression and block polyspermy. Clinically, PLCζ deficiency—whether caused by genetic mutations, reduced expression, or improper localization—represents a unifying explanation for certain forms of male infertility, including total fertilization failure (TFF) following intracytoplasmic sperm injection (ICSI). Globozoospermia is a prime example; this condition is characterized by round-headed sperm devoid of acrosomes and exhibiting significantly reduced or absent PLCζ and often results in fertilization failure. Diagnostic methods such as immunofluorescence, Western blotting, and the mouse oocyte-activation test collectively support the identification and characterization of PLCζ-related defects, while genetic testing for mutations in the PLCZ1 gene has proven valuable for identifying hereditary causes of sperm-borne oocyte-activation deficiency (OAD). Therapeutic approaches range from assisted oocyte activation (AOA) with calcium ionophores to emerging interventions that introduce functional PLCζ protein or mRNA directly into the oocyte. These advancements demonstrate the rapid translation of foundational discoveries into clinically actionable interventions. Future investigations are poised to refine diagnostic assays, standardize measurement protocols, and explore the potential of gene therapy or CRISPR/Cas9-mediated correction for heritable PLCζ abnormalities. By addressing both the molecular basis and translational applications of PLCζ, recent findings underscore its indispensable role in fertility care and lay out a path toward further innovation in assisted reproductive technologies. Full article

Spermatozoon volume regulation is an essential determinant of male fertility competence in mammals and oviparous fishes. In mammals, aquaporin water channels (AQP3, -7 and -8) have been suggested to play a role in spermatozoon cell volume regulatory responses in the hypotonic female oviduct. In contrast, the ejaculated spermatozoa of marine teleosts, such as the gilthead seabream (Sparus aurata), experience a high hypertonic shock in seawater, initially resulting in an Aqp1aa-mediated water efflux, cell shrinkage and the activation of motility. Further regulatory recovery of cell volume in post-activated spermatozoa is mediated by Aqp4a in cooperation with the Trpv4 Ca²⁺ channel and other ion channels and transporters. Using a paralog-specific antibody, here, we show that seabream spermatozoa also express the aquaglyceroporin AQP3 ortholog Aqp3a, which is highly accumulated in the mid posterior region of the spermatozoon flagella, in a similar pattern to that described in mouse and human sperm. To investigate the role of Aqp3a in seabream sperm motility, we used a recently developed AQP3 antagonist (DFP00173), as well as the seabream Aqp3a-specific antibody (α-SaAqp3a), both of which specifically inhibit Aqp3a-mediated water conductance when the channel was heterologously expressed in Xenopus laevis oocytes. Inhibition with either DFP00173 or α-SaAqp3a did not affect sperm motility activation but did impair the spermatozoon motion kinetics at 30 s post activation in a dose-dependent manner. Interestingly, in close resemblance to the phenotypes of AQP3-deficient murine sperm, electron microscopy image analysis revealed that both Aqp3a inhibitors induce abnormal sperm tail morphologies, including swelling and angulation of the tail, with complete coiling of the flagella in some cases. These findings suggest a conserved role of Aqp3a as an osmosensor that regulates cell volume in fish spermatozoa under a high hypertonic stress, thereby controlling the efflux of water and/or solutes in the post-activated spermatozoon. Full article

The sperm-specific phospholipase C zeta (PLCζ) protein is widely considered as the predominant physiological stimulus for initiating the Ca²⁺ release responsible for oocyte activation during mammalian fertilization. The increasing number of genetic and clinical reports that directly link PLCζ defects and/or deficiencies with oocyte activation failure (OAF) necessitates the use of a powerful therapeutic intervention to overcome such cases of male factor infertility. Currently, in vitro fertilization (IVF) clinics treat OAF cases after intracytoplasmic sperm injection (ICSI) with Ca²⁺ ionophores. Despite their successful use, such chemical agents are unable to trigger the physiological pattern of Ca²⁺ oscillations. Moreover, the safety of these ionophores is not yet fully established. We have previously demonstrated that recombinant PLCζ protein can be successfully used to rescue failed oocyte activation, resulting in efficient blastocyst formation. Herein, we produced a maltose binding protein (MBP)-tagged recombinant human PLCζ protein capable of inducing Ca²⁺ oscillations in mouse oocytes similar to those observed at fertilization. Circular dichroism (CD) experiments revealed a stable, well-folded protein with a high helical content. Moreover, the recombinant protein could retain its enzymatic properties for at least up to 90 days after storage at −80 °C. Finally, a chick embryo model was employed and revealed that exposure of fertilized chicken eggs to MBP-PLCζ did not alter the embryonic viability when compared to the control, giving a first indication of its safety. Our data support the potential use of the MBP-PLCζ recombinant protein as an effective therapeutic tool but further studies are required prior to its use in a clinical setting. Full article

During mammalian fertilization, repetitive intracellular Ca²⁺ increases known as Ca²⁺ oscillations occur. These oscillations are considered crucial for successful fertilization and subsequent embryonic development. Numerous researchers have endeavored to elucidate the factors responsible for inducing Ca²⁺ oscillations across various mammalian species. Notably, sperm-specific phospholipase C zeta (PLCζ) emerged as a prominent candidate capable of initiating Ca²⁺ oscillations, particularly in mammals. Genetic mutation of PLCζ in humans results in the absence of Ca²⁺ oscillations in mouse oocytes. Recent studies further underscored PLCζ’s significance, revealing that sperm from PLCζ-deficient (Plcz1^−/−) mice fail to induce Ca²⁺ oscillations upon intracytoplasmic sperm injection (ICSI). Despite these findings, observations from in vitro fertilization (IVF) experiments using Plcz1^−/− sperm revealed some residual intracellular Ca²⁺ increases and successful oocyte activation, hinting at potential alternative mechanisms. In this review, we introduced the current hypothesis surrounding oocyte activation in mammals, informed by contemporary literature, and probed into the enigmatic mechanisms underlying mammalian fertilization-induced oocyte activation. Full article

Background: Glutathione S-transferase (GST) M1 belongs to a family of detoxification enzymes and deficiency in enzyme activity is due to a homozygous deletion of the GSTM1 gene. Several studies reveal a possible correlation between female infertility and GSTM1 polymorphisms. The aim of this study is to investigate the effect of the GSTM1-null polymorphism in female infertility as well as in IVF parameters. Methods: In the study group 125 women were classified as infertile according to WHO and 49 women with at least one successful pregnancy and no miscarriages, as control group. Genomic DNA from blood samples was isolated and PCR amplification was applied to determine the presence of GSTM1-null genotype. Results: Data analysis demonstrated a statistically significant higher presence of GSTM1-null variant in the infertile group compared to the control group. In a subgroup analysis of the infertile group, the estradiol levels, the number of fertilized oocytes as well as the number and the quality of the cumulus-oocyte complex, were statistically significant higher in women detected with the wildtype of GSTM1 gene compared to those who had the GSTM1 null genotype (deletion). Conclusions: Our study results propose a possible involvement of GMST1 in female infertility and may help elucidate possible interactions between the microenvironment of oocytes and the oxidative stress. Full article

Oocyte activation, a fundamental event during mammalian fertilisation, is initiated by concerted intracellular patterns of calcium (Ca²⁺) release, termed Ca²⁺ oscillations, predominantly driven by testis-specific phospholipase C zeta (PLCζ). Ca²⁺ exerts a pivotal role in not just regulating oocyte activation and driving fertilisation, but also in influencing the quality of embryogenesis. In humans, a failure of Ca²⁺ release, or defects in related mechanisms, have been reported to result in infertility. Furthermore, mutations in the PLCζ gene and abnormalities in sperm PLCζ protein and RNA, have been strongly associated with forms of male infertility where oocyte activation is deficient. Concurrently, specific patterns and profiles of PLCζ in human sperm have been linked to parameters of semen quality, suggesting the potential for PLCζ as a powerful target for both therapeutics and diagnostics of human fertility. However, further to PLCζ and given the strong role played by Ca²⁺ in fertilisation, targets down- and up-stream of this process may also present a significantly similar level of promise. Herein, we systematically summarise recent advancements and controversies in the field to update expanding clinical associations between Ca²⁺-release, PLCζ, oocyte activation and human fertility. We discuss how such associations may potentially underlie defective embryogenesis and recurrent implantation failure following fertility treatments, alongside potential diagnostic and therapeutic avenues presented by oocyte activation for the diagnosis and treatment of human infertility. Full article

Phospholipase C zeta1 (Plcz1) was known to be a physiological factor in sperm that activates oocytes to complete meiosis by triggering Ca²⁺ oscillations after fertilisation. However, the role of male Plcz1 in spermatogenesis and early embryo development in progeny has been controversial. Plcz1 knockout (Plcz1^−/−) mouse model (Plcz1^m3 and Plcz1^m5) was generated by using the CRISPR-Cas9 system. The fertility of Plcz1^−/− mice was evaluated by analysing the number of offsprings, sperm quality, pathological changes in the testis and epididymis. RNA-seq and RT-PCR were performed to screen differentially expressed genes and signalling pathways related to fertility in Plcz1^−/− mice. Further mechanism was explored by using Plcz1^−/− cells. Plcz1 knockout led to hypofertility in male mice. In particular, a significant time delay in development and polyspermy was found in eggs fertilized by both Plcz1^m3 and Plcz1^m5 sperm. Interestingly, a decline in sperm quality combined with pathological changes in epididymis was found in Plcz1^m3 mice but not in Plcz1^m5 mice. Notably, abnormal cytoskeleton appears in epididymis of Plcz1^m3 mice and Plcz1^−/− cells. Cytoskeleton damage of epididymis is involved in fertility decline of males upon Plcz1 deficiency in this model. Full article

The group X secreted phospholipase A2 (PLA2G10) is present at high levels in mouse sperm acrosome. The enzyme is secreted during capacitation and amplifies the acrosome reaction and its own secretion via an autocrine loop. PLA2G10 also improves the rate of fertilization. In in vitro fertilization (IVF) experiments, sperm from Pla2g10-deficient mice produces fewer two-cell embryos, and the absence of PLA2G10 is rescued by adding recombinant enzymes. Moreover, wild-type (WT) sperm treated with recombinant PLA2G10 produces more two-cell embryos. The effects of PLA2G10 on mouse fertility are inhibited by sPLA₂ inhibitors and rescued by products of the enzymatic reaction such as free fatty acids, suggesting a role of catalytic activity. However, PLA2G10 also binds to mouse PLA2R1, which may play a role in fertility. To determine the relative contribution of enzymatic activity and PLA2R1 binding in the profertility effect of PLA2G10, we tested H48Q-PLA2G10, a catalytically-inactive mutant of PLA2G10 with low enzymatic activity but high binding properties to PLA2R1. Its effect was tested in various mouse strains, including Pla2r1-deficient mice. H48Q-PLA2G10 did not trigger the acrosome reaction but was as potent as WT-PLA2G10 to improve IVF in inbred C57Bl/6 mice; however, this was not the case in OF1 outbred mice. Using gametes from these mouse strains, the effect of H48Q-PLA2G10 appeared dependent on both spermatozoa and oocytes. Moreover, sperm from C57Bl/6 Pla2r1-deficient mice were less fertile and lowered the profertility effects of H48Q-PLA2G10, which were completely suppressed when sperm and oocytes were collected from Pla2r1-deficient mice. Conversely, the effect of WT-PLA2G10 was not or less sensitive to the absence of PLA2R1, suggesting that the effect of PLA2G10 is polymodal and complex, acting both as an enzyme and a ligand of PLA2R1. This study shows that the action of PLA2G10 on gametes is complex and can simultaneously activate the catalytic pathway and the PLA2R1-dependent receptor pathway. This work also shows for the first time that PLA2G10 binding to gametes’ PLA2R1 participates in fertilization optimization. Full article

Glycosylphosphatidylinositol-anchored sperm hyaluronidases (HYAL) assist sperm penetration through the cumulus–oocyte complex (COC), but their role in mammalian fertilization remains unclear. Previously, we demonstrated that sperm from HYAL 5 and 7 double-knockout (dKO) mice produced significantly less offspring than sperm from wild-type mice due to defective COC dispersal. However, the HYAL6 gene remained active in the sperm from the dKO mice, indicating that they were not entirely infertile. This study explored the role of HYAL6 in fertilization by analyzing HYAL6-mutant mice. In this mouse model, HYAL5 and HYAL7 were present in the HYAL6-knockout sperm, and they could disperse hyaluronic acid. We found that HYAL6 was present on the surface of sperm. However, male mice lacking the HYAL6 gene had normal fertility, testicular integrity, and sperm characteristics. Furthermore, in vitro fertilization assays demonstrated that HYAL6-deficient epididymal sperm functioned normally. Therefore, HYAL6 is dispensable for fertilization. Full article