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A Zinc-Dependent Mechanism Regulates Meiotic Progression in Mammalian Oocytes.

A Zinc-Dependent Mechanism Regulates Meiotic Progression in Mammalian Oocytes.

Biol Reprod. 2012 Feb 1;

Authors: Bernhardt ML, Kong BY, Kim AM, O'Halloran TV, Woodruff TK

Abstract

Precise coordination of meiotic progression is a critical determinant of an egg's capacity to be fertilized successfully, and zinc has emerged as a key regulatory element in this process. An early manifestation of a regulatory role for this transition metal is the significant increase in total intracellular zinc: this accumulation is essential for meiotic progression beyond telophase I, and the establishment of meiotic arrest at metaphase II. The subsequent developmental event, fertilization, induces a rapid expulsion of labile zinc that is a hallmark event in meiotic resumption. Here, we show that the zinc fluxes work in part by altering the activity of the cytostatic factor (CSF), the cellular activity required for the establishment and maintenance of metaphase II arrest in the mature, unfertilized egg. We propose a model in which zinc exerts concentration-dependent regulation of meiosis through the CSF component EMI2, a zinc-binding protein. Together, the data support the conclusion that zinc itself, through its interaction with EMI2, is a central component of the CSF.

PMID: 22302686 [PubMed - as supplied by publisher]

Inhibin α-Subunit N-terminus Interacts With Activin Type IB receptor To Disrupt Activin Signaling.

Inhibin α-Subunit N-terminus Interacts With Activin Type IB receptor To Disrupt Activin Signaling.

J Biol Chem. 2012 Jan 20;

Authors: Zhu J, Lin SJ, Zou C, Makanji Y, Jardetzky TS, Woodruff TK

Abstract

Inhibin is a heterodimeric peptide hormone produced in the ovary that antagonizes activin signaling and FSH synthesis in the pituitary. The inhibin β-subunit interacts with the activin type II receptor (ActRII) to functionally antagonize activin. The inhibin α-subunit mature domain (N-terminus) arose relatively early during the evolution of the hormone, and inhibin function is decreased by an antibody directed against the α-subunit N-terminal extension region or by deletion of the N-terminal region. We hypothesized that the α-subunit N-terminal extension region interacts with the activin type I receptor (ALK4) to antagonize activin signaling in the pituitary. Human or chicken free α-subunit inhibited activin signaling in a pituitary gonadotrope derived cell line (LβT2) in a dose-dependent manner, whereas an N-terminal extension deletion mutant did not. An α-subunit N-terminal peptide, but not a control peptide, was able to inhibit activin A signaling and decrease activin-stimulated FSH synthesis. Biotinylated inhibin A, but not activin A, bound ALK4. Soluble ALK4-ECD bioneutralized human free α-subunit in LβT2 cells, but did not affect activin A function. Competitive binding ELISAs with N-terminal mutants and an N-terminal region peptide confirmed that this region is critical for direct interaction of the α-subunit with ALK4. These data expand our understanding of how endocrine inhibin achieves potent antagonism of local, constitutive activin action in the pituitary, through a combined mechanism of competitive binding of both ActRII and ALK4 by each subunit of the inhibin heterodimer, in conjunction with the co-receptor betaglycan, to block activin receptor-ligand binding, complex assembly, and downstream signaling.

PMID: 22267736 [PubMed - as supplied by publisher]

Improved Fertility Preservation Care for Male Patients With Cancer After Establishment of Formalized Oncofertility Program.

Improved Fertility Preservation Care for Male Patients With Cancer After Establishment of Formalized Oncofertility Program.

J Urol. 2012 Jan 18;

Authors: Sheth KR, Sharma V, Helfand BT, Cashy J, Smith K, Hedges JC, Köhler TS, Woodruff TK, Brannigan RE

Abstract

PURPOSE: Survival to reproductive age among men with cancer has steadily increased and yet cancer therapy and cancer itself may carry the risk of infertility. Since 2006, we have used a formalized fertility preservation program with expedited fertility care at our institution. We assessed the impact of this program by comparing the frequency of sperm cryopreservation and patient characteristics before and after its implementation. MATERIALS AND METHODS: Men 18 to 55 years old diagnosed with cancer at our institution from 2002 to 2010 were included in our study. We retrospectively reviewed patient charts to identify those who were offered and subsequently used fertility preservation services before and after program formalization. RESULTS: From 2002 to 2010 at our institution 4,818 men 18 to 55 years old were diagnosed with cancer, of whom 411 were offered fertility preservation consultation and 249 underwent sperm cryopreservation. Since program implementation, the annual number of men receiving fertility preservation consultation and undergoing sperm cryopreservation increased by 2.4 and 2.7-fold, respectively, while the total number diagnosed with cancer remained fairly constant. Upon substratifying patients into the more conventional reproductive age range of 18 to 40 years 23.4% of all men with cancer in this group were offered consultation before formalization vs 43.3% after formalization (p <0.05). The overall sperm use and discard rates were 8.4% and 14.8%, respectively. CONCLUSIONS: A formalized institutional fertility preservation program significantly increased the overall number and percent of male patients with cancer who received fertility preservation consultation and pursued sperm cryopreservation. These increases were seen in men with all types of cancer and across all demographics assessed at our institution.

PMID: 22264454 [PubMed - as supplied by publisher]

Creating a continuum of care: integrating obstetricians and gynecologists in the care of young cancer patients.

Creating a continuum of care: integrating obstetricians and gynecologists in the care of young cancer patients.

Clin Obstet Gynecol. 2011 Dec;54(4):619-32

Authors: Kong BY, Skory RM, Woodruff TK

Abstract

Cancer therapy can be lifesaving but significantly diminish female reproductive potential. This review provides an overview of the deleterious effects of cancer treatments on reproductive function, the fertility preservation options currently available for young women, and the outcomes of pregnancy after cancer treatment. In addition, special considerations for women who are diagnosed with cancer during pregnancy are discussed. To optimize the continuum of care for the patient, new corridors of communication between obstetricians, gynecologists, and oncology specialists must be developed to ensure the best outcomes for the patient, both in terms of cancer treatment and fertility preservation.

PMID: 22031251 [PubMed - in process]

The Gynecologist Has a Unique Role in Providing Oncofertility Care to Young Cancer Patients.

The Gynecologist Has a Unique Role in Providing Oncofertility Care to Young Cancer Patients.

US Obstet Gynecol. 2011 Jan 1;6(1):24-34

Authors: Duncan FE, Jozefik JK, Kim AM, Hirshfeld-Cytron J, Woodruff TK

Abstract

Facing a cancer diagnosis at any age is devastating. However, young cancer patients have the added burden that life-preserving cancer treatments, including surgery, chemotherapy, and radiotherapy, may compromise their future fertility. The possibility of reproductive dysfunction as a consequence of cancer treatment has a negative impact on the quality of life of cancer survivors. The field of oncofertility, which merges the clinical specialties of oncology and reproductive endocrinology, was developed to explore and expand fertility preservation options and to better manage the reproductive status of cancer patients. Fertility preservation for females has proved to be a particular challenge because mature female gametes are rare and difficult to acquire. The purpose of this article is to provide the gynecologist with a comprehensive overview of how cancer treatments affect the female reproductive axis, delineate the diverse fertility preservation options that are currently available or being developed for young women, and describe current measures of ovarian reserve that can be used pre- and post-cancer treatment. As a primary care provider, the gynecologist will likely interact with patients throughout the cancer care continuum. Thus, the gynecologist is in a unique position to join the oncofertility team in providing young cancer patients with up-to-date fertility preservation information and referrals to specialists.

PMID: 21927621 [PubMed - as supplied by publisher]

Nonmalignant Diseases and Treatments Associated with Primary Ovarian Failure: An Expanded Role for Fertility Preservation.

Nonmalignant Diseases and Treatments Associated with Primary Ovarian Failure: An Expanded Role for Fertility Preservation.

J Womens Health (Larchmt). 2011 Aug 9;

Authors: Hirshfeld-Cytron J, Gracia C, Woodruff TK

Abstract Cancer treatments can be detrimental to fertility; recent literature has focused on the efforts of fertility preservation for this patient population. It should be recognized, however, that several nonmalignant medical conditions and therapeutic interventions could be similarly hazardous to fertility. Some of these nonmalignant diseases and their treatments that can adversely impact the reproductive axis are gastrointestinal diseases, rheumatologic disorders, nonmalignant hematologic conditions, neurologic disorders, renal disorders, gynecologic conditions, and metabolic diseases. Their negative effects on reproductive function are only now being appreciated and include impaired ovarian function, endocrine function, or sexual function and inability to carry a pregnancy to term. Complications and comorbidities associated with certain diseases may limit the success of established fertility preservation options. Recent advances in fertility preservation techniques may provide these patients with new options for childbearing. Here, we review several fertility-threatening conditions and treatments, describe current established and experimental fertility preservation options, and present three initiatives that may help minimize the adverse reproductive effects of these medical conditions and treatments by raising awareness of the issues and options: (1) increase awareness among practitioners about the reproductive consequences of specific diseases and treatments, (2) facilitate referral of patients to fertility-sparing or restorative programs, and (3) provide patient education about the risk of infertility at the time of diagnosis before initiation of treatment.

PMID: 21827325 [PubMed - as supplied by publisher]

Animal age, weight and estrus cycle stage impact the quality of in vitro grown follicles.

Animal age, weight and estrus cycle stage impact the quality of in vitro grown follicles.

Hum Reprod. 2011 Jun 13;

Authors: Hirshfeld-Cytron JE, Duncan FE, Xu M, Jozefik JK, Shea LD, Woodruff TK

BACKGROUND Ovarian tissue cryopreservation is an emerging fertility preservation option, and culturing follicles isolated from this tissue to obtain mature gametes may ultimately be the best solution for patients for whom transplantation is contraindicated. It is unclear, however, how patient-specific variables (including age, weight and menstrual cycle stage) impact follicle growth and quality during three-dimensional culture. METHODS We used a mouse model to systematically determine how these variables impact in vitro follicle growth. We characterized metabolic and hormonal profiles of mice at specific ages, weights and cycle stages and secondary follicles from these cohorts were isolated and cultured. We then assessed follicle survival, growth and function, as well as meiotic competence and spindle morphology of the resulting oocytes. RESULTS We found that older mice and mice with increased body weight had higher serum cholesterol, abnormal glucose tolerance and lower levels of circulating Anti-Müllerian hormone compared with younger and leaner controls. Secondary follicles isolated from different cohorts and grown in vitro had indistinguishable growth trajectories. However, the follicles isolated from older and heavier mice and those in diestrus had altered hormone profiles. These follicles contained oocytes with reduced meiotic competence and produced oocytes with greater spindle defects. CONCLUSIONS These results suggest that the original physical environment of the follicle within the ovary can impact its function when isolated and cultured. These findings are valuable as we begin to use in vitro follicle growth technology for a heterogeneous fertility preservation patient population.

PMID: 21669966 [PubMed - as supplied by publisher]

A truncated, activin-induced Smad3 isoform acts as a transcriptional repressor of FSHβ expression in mouse pituitary.

A truncated, activin-induced Smad3 isoform acts as a transcriptional repressor of FSHβ expression in mouse pituitary.

Mol Cell Endocrinol. 2011 Jun 1;

Authors: Kim SY, Zhu J, Woodruff TK

The receptor-regulated protein Smad3 is key player in the signaling cascade stimulated by the binding of activin to its cell surface receptor. Upon phosphorylation, Smad3 forms a heterocomplex with Smad2 and Smad4, translocates to the nucleus and acts as a transcriptional co-activator. We have identified a unique isoform of Smad3 that is expressed in mature pituitary gonadotropes. 5' RACE revealed that this truncated Smad3 isoform is transcribed from an ATG site within exon 4 and consists of 7 exons encoding half of the linker region and the MH2 region. In pituitary cells, the truncated Smad3 isoform was phosphorylated upon activin treatment, in a manner that was temporally distinct from the phosphorylation of full-length Smad3. Activin-induced phosphorylation of Smad3 and the truncated Smad3 isoform was blocked by both follistatin and siRNA-mediated knockdown of Smad3. The truncated Smad3 isoform antagonized Smad3-mediated, activin-responsive promoter activity. We propose that the pituitary gonadotrope contains an ultra-short, activin-responsive feedback loop utilizing two different isoforms of Smad3, one which acts as an agonist (Smad3) and another that acts as an intracrine antagonist (truncated Smad3 isoform) to regulate FSHβ production.

PMID: 21664424 [PubMed - as supplied by publisher]

Gene expression in mouse ovarian follicle development in vivo versus an ex vivo alginate culture system.

Gene expression in mouse ovarian follicle development in vivo versus an ex vivo alginate culture system.

Reproduction. 2011 May 24;

Authors: Parrish E, Siletz A, Xu M, Woodruff TK, Shea L

Ovarian follicle maturation results from a complex interplay of endocrine, paracrine, and direct cell-cell interactions. This study compared the dynamic expression of key developmental genes during folliculogenesis in vivo and during in vitro culture in a three-dimensional alginate hydrogel system. Candidate gene expression profiles were measured within mouse two-layered secondary follicles (2LS), multi-layered secondary follicles (MLS), and cumulus-oocyte complexes (COCs). The expression of 20 genes involved in endocrine communication, growth signaling, and oocyte development was investigated by real-time PCR. Gene product levels were compared between i) follicles of similar stage and ii) COCs derived either in vivo or by in vitro culture. For follicles cultured for 4 days, the expression pattern and the expression level of 12 genes was the same in vivo and in vitro. Several endocrine (Cyp19a1, Inhβa) and growth related genes (Bmp15, Kitl, Tgfβr2) were down-regulated relative to in vivo follicles. For COCs obtained from cultured follicles, endocrine related genes (Inhα and Inhβa) had increased expression relative to in vivo counterparts, whereas growth related genes (Bmp15, Gdf9, Kit) and zona pellucida genes were decreased. However, most of the oocyte specific genes (e.g., Figlα, Jag1, Mater) were expressed in vitro at the same level and with the same pattern as in vivo-derived follicles. These studies establish the similarities and differences between in vivo and in vitro cultured follicles, guiding the creation of environments that maximize follicle development and oocyte quality.

PMID: 21610168 [PubMed - as supplied by publisher]

Zinc Sparks Are Triggered by Fertilization and Facilitate Cell Cycle Resumption in Mammalian Eggs.

Zinc Sparks Are Triggered by Fertilization and Facilitate Cell Cycle Resumption in Mammalian Eggs.

ACS Chem Biol. 2011 Apr 28;

Authors: Kim AM, Bernhardt ML, Kong BY, Ahn RW, Vogt S, Woodruff TK, O'Halloran TV

In last few hours of maturation, the mouse oocyte takes up over twenty billion zinc atoms and arrests after the first meiotic division, until fertilization or pharmacological intervention stimulates cell cycle progression toward a new embryo. Using chemical and physical probes, we show that fertilization of the mature, zinc-enriched egg triggers the ejection of zinc into the extracellular milieu in a series of coordinated events termed zinc sparks. These events immediately follow the well-established series of calcium oscillations within the activated egg and are evolutionarily conserved in several mammalian species, including rodents and nonhuman primates. Functionally, the zinc sparks mediate a decrease in intracellular zinc content that is necessary for continued cell cycle progression, as increasing zinc levels within the activated egg results in the reestablishment of cell cycle arrest at metaphase. The mammalian egg thus uses a zinc-dependent switch mechanism to toggle between metaphase arrest and resumption of the meiotic cell cycle at the initiation of embryonic development.

PMID: 21526836 [PubMed - as supplied by publisher]