Effect of single versus grouped culture of human cumulus-oocyte complexes in PCOS women treated with biphasic in vitro maturation: A sibling oocyte pilot study.

Purpose: This study investigated the differences in the maturation rate of single versus grouped cumulus-oocyte complexes (COCs) culture methods for capacitation in vitro maturation (CAPA-IVM) in women with polycystic ovary syndrome (PCOS).

Methods: This study was performed at My Duc Phu Nhuan Hospital, Vietnam from October 1, 2020 to October 24, 2021. Women aged 18-37 years with a diagnosis of PCOS were recruited.

Mouse oocytes restore antral stage partial mechanical denudation in vitro.

In Brief: Partially denuded mouse cumulus-oocyte complexes restore likely functional transzonal projections in culture, under meiotic inhibition, with no detectable impact on oocyte competence. This proof-of-concept study constitutes positive premises for improving the developmental competence of human capacitation (CAPA)-in vitro maturation (IVM) oocytes with inadequate somatic cell connections.

Abstract: In vitro oocyte culture might be the sole option for fertility preservation in some patients.

Glucose and redox metabolism in meiotically blocked in vitro grown mouse antral follicles.

Purpose: Glucose and redox metabolism characterization in mouse antral follicles with meiotically blocked oocytes, after in vitro follicle culture (IFC) from the early secondary stage.

Methods: Following IFC (10 days), oocytes, corresponding cumulus (CC), and granulosa cells (GC) were collected from antral follicles: (i) on day 9-immature, germinal vesicle (GV) stage; (ii) on day 10, after hCG/EGF stimulation-mature, metaphase II (MII) stage and meiotically blocked (MB) immature GV stage. The metabolic profiles of all samples (GV, MII, and MB) were compared by measuring changes in metabolites involved in glycolysis, tricarboxylic acid (TCA) cycle, pentose phosphate pathway (PPP), and redox activity via enzymatic spectrophotometric assays in each cell type.

Effects of lactate, super-GDF9, and low oxygen tension during bi-phasic in vitro maturation on the bioenergetic profiles of mouse cumulus-oocyte complex†.

In vitro maturation (IVM) is an alternative assisted reproductive technology with reduced hormone-related side effects and treatment burden compared to conventional IVF. Capacitation (CAPA)-IVM is a bi-phasic IVM system with improved clinical outcomes compared to standard monophasic IVM. Yet, CAPA-IVM efficiency compared to conventional IVF is still suboptimal in terms of producing utilizable blastocysts.

IVF characteristics and the molecular luteal features of random start IVF cycles are not different from conventional cycles in cancer patients.

Study Question: Are the IVF parameters and the steroidogenic luteal characteristics of random-start IVF cycles different from conventional cycles in cancer patients?

Summary Answer: No; controlled ovarian stimulation cycles randomly started at late follicular phase (LFP) and luteal phase (LP) are totally comparable to those conventional IVF cycles started at early follicular phase (EFP) in terms of the expression of the enzymes involved in cholesterol utilization and steroid hormone biosynthesis pathways, gonadotropin receptor expression and, estradiol (E2) and progesterone (P4) production in addition to the similarities in ovarian response to gonadotropin stimulation, oocyte yield, fertilization rate and embryo development competency in cancer patients.

What Is Known Already: Random start ovarian stimulation protocols are commonly employed for oocyte and embryo freezing for fertility preservation in cancer patients with time constraints who do not have sufficient time to undergo ovarian stimulation initiated conventionally at EFP of the next cycle. No data is available regarding the molecular steroidogenic features of these cycles analyzed together with the clinical IVF characteristics in cancer patients.

Characterization of carbohydrate metabolism in in vivo- and in vitro-grown and matured mouse antral follicles†.

Establishing an ideal human follicle culture system for oncofertility patients relies mainly on animal models since donor tissue is scarce and often of suboptimal quality. The in vitro system developed in our laboratory supports the growth of prepubertal mouse secondary follicles up to mature oocytes. Given the importance of glucose in preparing the oocyte for proper maturation, a baseline characterization of follicle metabolism both in the culture system and in vivo was carried out.

Cholesterol uptake or trafficking, steroid biosynthesis, and gonadotropin responsiveness are defective in young poor responders.

Objective: To investigate whether poor ovarian response in young patients undergoing in vitro fertilization simply involves lesser follicle growth due to diminished ovarian reserve or whether there are intrinsic perturbations in the ovary.

Design: A translational research study.

Setting: University Hospital Translational Research Center.

Effect of cumulin and super-GDF9 in standard and biphasic mouse IVM.

Purpose: In vitro maturation (IVM) is a technology that generates mature oocytes following culture of immature cumulus-oocyte complexes (COC) in vitro. IVM is characterized by minimal patient stimulation, making it attractive for certain patient groups. Recently, a biphasic IVM system, capacitation (CAPA)-IVM, has shown improved clinical outcomes relative to standard IVM; however, it remains less efficient than IVF.

Positive effects of amphiregulin on human oocyte maturation and its molecular drivers in patients with polycystic ovary syndrome.

Study Question: Does use of medium containing amphiregulin improve meiotic maturation efficiency in oocytes of women with polycystic ovary syndrome (PCOS) undergoing in vitro maturation (IVM) preceded by a capacitation culture step capacitation IVM (CAPA-IVM)?

Summary Answer: Use of medium containing amphiregulin significantly increased the maturation rate from oocytes retrieved from follicles with diameters <6 or ≥6 mm pre-cultured in capacitation medium.

What Is Known Already: Amphiregulin concentration in follicular fluid is correlated with human oocyte developmental competence. Amphiregulin added to the meiotic trigger has been shown to improve outcomes of IVM in a range of mammalian species.

Reversing complete mechanical transzonal projections disruption during mouse in vitro follicle culture with unaltered oocyte competence†.

In vitro oocyte growth is widely studied as an alternative fertility preservation approach. Several animal models are used to generate extensive information on this complex process regulated by the constant and dynamic interaction between the oocyte and its somatic compartment throughout follicle growth and maturation. A two-dimensional attachment mouse secondary follicle culture system was used to assess the oocyte's capacity to overcome disconnection from its somatic companions at different developmental stages for final competence acquisition.

Glucose metabolism characterization during mouse in vitro maturation identifies alterations in cumulus cells†.

In vitro maturation (IVM) is an assisted reproduction technique with reduced hormone-related side-effects. Several attempts to implement IVM in routine practice have failed, primarily due to its relatively low efficiency compared with conventional in vitro fertilization (IVF). Recently, capacitation (CAPA)-IVM-a novel two-step IVM method-has improved the embryology outcomes through synchronizing the oocyte nuclear and cytoplasmic maturation.

Terminal differentiation of human granulosa cells as luteinization is reversed by activin-A through silencing of Jnk pathway.

Molecular mechanisms underlying luteinization (terminal differentiation of granulosa and theca cells after ovulation) and luteolysis (demise of corpus luteum) are poorly understood in human ovary. Here we report that activin-A, after binding to its cognate receptors induces a functional luteolytic state and reverses luteinization phenotype by downregulating the expression of the steroidogenic enzymes, LH receptor and VEGF and reducing estradiol (E) progesterone (P) production and upregulating FSH receptor and cyclin D1 expression in human primary luteinized granulosa cells. Further, this action of activin-A involves downregulation of JNK signaling pathway and is opposite to that of human chorionic gonadotropin (hCG), which acts as a luteotropic hormone and improves luteal function through the activation of JNK pathway in the same cell type.

hCG Improves Luteal Function and Promotes Progesterone Output through the Activation of JNK Pathway in the Luteal Granulosa Cells of the Stimulated IVF Cycles†.

Unlabelled: Human chorionic gonadotropin (hCG) is a luteotropic hormone that promotes the survival and steroidogenic activity of corpus luteum (CL) by acting through luteinizing hormone receptors (LHRs) expressed on luteinized theca and granulosa cells (GCs). Therefore, it is used to support luteal phase in in vitro fertilization (IVF) cycles to improve clinical pregnancy rates and prevent miscarriage. However, the molecular mechanism underlying this action of hCG is not well characterized.

Luteal granulosa cells from natural cycles are more capable of maintaining their viability, steroidogenic activity and LH receptor expression than those of stimulated IVF cycles.

Study Question: Are there any differences in the molecular characteristics of the luteal granulosa cells (GC) obtained from stimulated versus non-stimulated (natural) IVF cycles that may help explain the defective luteal phase in the former?

Summary Answer: Luteal GC of stimulated IVF cycles, particularly those of agonist-triggered antagonist cycles, are less viable ex vivo, express LH receptor and anti-apoptotic genes at lower levels, undergo apoptosis earlier and fail to maintain their estradiol (E2) and progesterone (P4) production in comparison to natural cycle GC.

What Is Known Already: Luteal function is defective in stimulated IVF cycles, which necessitates P4 and/or hCG administration (known as luteal phase support) in order to improve clinical pregnancy rates and prevent miscarriage. The luteal phase becomes shorter and menstruation begins earlier than a natural cycle if a pregnancy cannot be achieved, indicative of early demise of corpus luteum (premature luteolysis).

Endogenous c-Jun N-terminal kinase (JNK) activity marks the boundary between normal and malignant granulosa cells.

Granulosa cell tumor of the ovary (GCT) is a very rare tumor, accounting for only 2% of all ovarian tumors. It originates from sex cords in the ovary and can be divided into adult (95%) and juvenile (5%) types based on histologic findings. To date, no clear etiologic process has been identified other than a missense point mutation in the FOXL2 gene.

FSH Stimulation promotes progesterone synthesis and output from human granulosa cells without luteinization.

Study Question: Can granulosa cells produce progesterone (P) in response to FSH stimulation?

Summary Answer: FSH actively promotes P synthesis and output from granulosa cells without luteinization by up-regulating the expression and increasing enzymatic activity of 3β-hydroxysteriod dehydrogenoase (3β-HSD), which converts pregnenolone to P.

What Is Known Already: Serum P level may rise prematurely prior to ovulation trigger in stimulated IVF cycles and adversely affect implantation and clinical pregnancy rates by impairing endometrial receptivity.

Study Design, Size, Duration: A translational research study.

Cytotoxicity and mitogenicity assays with real-time and label-free monitoring of human granulosa cells with an impedance-based signal processing technology intergrating micro-electronics and cell biology.

A recently developed technology (xCelligence) integrating micro-electronics and cell biology allows real-time, uninterrupted and quantitative analysis of cell proliferation, viability and cytotoxicity by measuring the electrical impedance of the cell population in the wells without using any labeling agent. In this study we investigated if this system is a suitable model to analyze the effects of mitogenic (FSH) and cytotoxic (chemotherapy) agents with different toxicity profiles on human granulosa cells in comparison to conventional methods of assessing cell viability, DNA damage, apoptosis and steroidogenesis. The system generated the real-time growth curves of the cells, and determined their doubling times, mean cell indices and generated dose-response curves after exposure to cytotoxic and mitogenic stimuli.

The magnitude of gonadotoxicity of chemotherapy drugs on ovarian follicles and granulosa cells varies depending upon the category of the drugs and the type of granulosa cells.

Study Question: Do different chemotherapy drugs exert the same magnitude of cytotoxicity on dormant primordial follicles and the growing follicle fraction in the ovary in vivo and on mitotic non-luteinized and non-mitotic luteinized granulosa cells in vitro?

Summary Answer: Cyclophosphamide (alkylating agent) and cisplatin (alkylating like) impacted both primordial and pre-antral/antral follicles and both mitotic and non-mitotic granulosa cells, whereas the anti-metabolite cancer drug gemcitabine was detrimental only to pre-antral/antral follicles and mitotic non-luteinized granulosa cells.

What Is Known Already: It is already known that anti-metabolite cancer drugs are less detrimental to the ovary than alkylating and alkylating like agents, such as cyclophosphamide and cisplatin. This assumption is largely based on the results of clinical reports showing lower rates of amenorrhea in women receiving anti-metabolite agent-based regimens compared with those treated with the protocols containing an alkylating drug or a platinum compound.

GnRH agonist leuprolide acetate does not confer any protection against ovarian damage induced by chemotherapy and radiation in vitro.

Study Question: Is there any in vitro evidence for or against ovarian protection by co-administration of a GnRH agonist with chemotherapy in human?

Summary Answer: The co-administration of GnRH agonist leuprolide acetate with cytotoxic chemotherapy agents does not preserve ovarian reserve in vitro.

What Is Known Already: Randomized controlled trials of the co-administration of gonadotrophin-releasing hormone (GnRH) agonists with adjuvant chemotherapy to preserve ovarian function have shown contradictory results. This fact, together with the lack of a proven molecular mechanism of action for ovarian protection with GnRH agonist (GnRHa) places this approach as a fertility preservation strategy under scrutiny.